Abstract: N/A
Genetic modification from a cattle producers perspective
Farmers have made tremendous strides in production and efficiency in crops and livestock in the past 50 years. Production methods and improvements in agronomy have led to major increases in crop yields and the development of hybrids and genetically modified plants have led to even greater improvements. Beef cattle production has also increased and become more efficient through improvements in management techniques and the utilization of performance and genomic testing. Genetic modification has yet to have an impact on beef production. Some producers are wary about the future of GMO’s in the beef industry, however many also look at it as an opportunity to improve sustainability, efficiency and animal welfare to add to a very slim bottom line. The question remains as to how potential GMO animals in the food chain will be viewed by the consumer. Guidance from the USDA and FDA will help resolve this issue however public trust in a safe product is a concern for many producers. Some breed associations including the American Hereford Association have adopted rules to address the registration and identification of genetically modified animals within the breed registry in the future.

Biography: I was raised on a diversified crop and livestock farm in southeastern SD and have been involved in the production of registered Hereford cattle for 50 years. I have been involved in agriculture in several areas having been in agricultural finance for over 20 years, worked in the ethanol industry for 2 years and owned and operated a farm real estate and auction company for over 10 years. I have served on the national board of directors for the American Hereford Association and was President of that board in 2019.

Abstract:
Gene Therapy for children with AADC deficiency
Aromatic L-amino acid decarboxylase (AADC) deficiency is a rare autosomal recessive disorder that causes congenital impairment of dopamine and serotonin synthesis. The disease presents with hypotonia, oculogyric crises (OGC), dystonia, autonomic dysfunction, sleep disruption, and global developmental delay. This study aims to evaluate the safety and efficacy of real-time MR-guided delivery of adeno-associated virus serotype 2 (AAV2)-hAADC to the substantia nigra (SNc) and ventral tegmental area (VTA).
Eight subjects were treated with either a low (Cohort 1) or high (Cohort 2) dose of vector (1.3 x 1011 and 4.2 x 1011 vector genomes) and have been followed up for over 3 years after gene therapy.
Real-time MR data confirmed 70-100% coverage of the SNc/VTA. The surgical intervention was well-tolerated by all subjects. All subjects developed mild to moderate dyskinesia post-gene transfer, peaking in severity between 1-3 months. OGCs completely resolved in 7/8 subjects. Sleep and mood improved dramatically in all subjects. Motor function improved by as measured by GMFM-88 scale; and 6 subjects attained the ability to sit independently and 2 take steps without support. CSF homovanillic acid (HVA) increased in all subjects from less than 10% of the lower limit of normal at baseline, to 24-47% at 6-12 months post-gene transfer, consistent with increased brain dopamine synthesis. 18FDOPA PET demonstrated increased uptake in the midbrain and striatum.
AADC gene transfer to SNc/VTA is well-tolerated, safe, and leads to improvement in disease symptoms and motor function.

Biography: Dr. Bankiewicz earned an M.D. from Jagiellonian University in Kraków and a Ph.D., D.Sc. from the Institute of Neurology and Psychiatry in Warsaw, Poland, as well as the title of Professor by the President of Republic of Poland. He is a member of Polish Academy of Science. Dr. Bankiewicz was a recipient of a Fogarty Fellowship and trained under Dr. Irvin Kopin and Dr. Edward Oldfield at the Surgical Neurology at National Institutes of Neurological Disorders and Stroke at NIH, Bethesda where he remained as a Staff Scientist. Dr. Bankiewicz is a recognized expert in neuro-restorative medicine, including delivery of therapeutics to the brain as well as gene therapy with successful translation of multiple drugs and gene therapies to the clinic. Previous positions included 20 years tenure at University of California San Francisco where he currently holds position of Endowed Chair Kinetics Foundation in Translational Research and Professor of Neurological Surgery. He also held position of Chief of Molecular Therapeutics Section at NIH. Dr. Bankiewicz has both industry and academic experience, is an inventor on numerous patents and has published more than 230 peer-reviewed research articles.
Currently, Dr. Bankiewicz holds academic position at The Ohio State University as a Professor of Neurosurgery and Gilbert and Kathryn Mitchell Endowed Chair. Throughout his career, Dr. Bankiewicz has maintained a strong focus on the development of translational approaches to drug, gene and cell replacement therapies, and he has displayed the ability to synthesize distinct technologies into powerful new approaches to the treatment of serious diseases, including brain cancer, Parkinson’s disease, Huntington’s disease, Alzheimer’s diseases, pediatric neurotransmitter deficiency and lysosomal storage disorders. Dr. Bankiewicz has co- founded MedGenesis Therapeutix, Inc, Voyager Therapeutics, Inc., Brain Neurotherapy Bio, Inc, and has been serving on various Scientific Advisory Boards. Dr. Bankiewicz pioneered and has been conducting two ongoing gene therapy clinical trials in Parkinson’s Disease, trial in Multiple System Atrophy and in pediatric neurotransmitter deficiency. He also invented several devices currently used clinically to administer gene therapy to the brain.

Abstract:
p53 dynamics and the DNA damage response in single cells
The p53 tumor suppressor regulates distinct responses to cellular stresses, including several forms of DNA damage. Such DNA damage is integral to methods of genome editing, and thus p53 regulation plays a critical role in mediating genome editing responses. Furthermore, distinct forms of DNA damage are differentially encoded by p53 dynamics, but the mechanisms by which cells decode p53 dynamics to properly regulate target genes are not well understood. Previously, we showed that p53 target expression dynamics are shaped by the relationship between the target mRNA and protein decay rates and the oscillation frequency of p53 dynamics. Here, we focused on aspects of target transcript synthesis. We determined in individual cells how canonical p53 target gene promoters vary in responsiveness to features of p53 dynamics. Employing a chemical perturbation approach and an optogenetic approach, we independently modulated p53 pulse amplitude, duration, or frequency, and we then monitored p53 levels and target promoter activation in individual cells. We identified distinct signal processing features— thresholding in response to amplitude modulation, a refractory period in response to duration modulation, and dynamic filtering in response to frequency modulation. We then showed that the signal processing features not only affect p53 target promoter activation, they also affect downstream cellular functions. Based on these findings, we determined how p53 dynamic mode-switching shapes dose-dependent responses to DNA damage. Our study shows how p53 target gene promoters can differentially decode features of p53 dynamics to generate distinct responses, providing insight into how perturbing p53 dynamics can be used to control cell fate responses to a wide spectrum of cellular stresses. Such insight may help to improve genome editing efforts.

Biography: Eric Batchelor completed undergraduate training in physics at Villanova University in Villanova, PA. He obtained his M.S. and Ph.D. in Physics from the University of Pennsylvania, Philadelphia, PA, where he studied two-component signal transduction in bacteria in the laboratory of Dr. Mark Goulian. He pursued postdoctoral training in Dr. Galit Lahav’s lab in the Department of Systems Biology at Harvard Medical School, Boston, MA where he studied p53's dynamical response to DNA damage. After completing his postdoctoral training, he started his own lab in the Laboratory of Pathology at the National Cancer Institute of the National Institutes of Health, Bethesda, MD. He recently moved to the Department of Integrative Biology and Physiology at the University of Minnesota Medical School, Minneapolis, MN.

Abstract:
How can Cyobiologists help Genome Writers?
This talk will provide a brief overview of some of the breakthrough cryopreservation technologies being developed in our new NSF Engineering Research Center (ERC) for Advanced Technologies for the Preservation of Biological Systems (ATP-Bio). Our goal is to “stop biological time” and radically extend the ability to bank and transport cells, tissues, organoids, organs, and organisms. Due to the relevance of genetic model organisms to genome writers, I will use the talk to particularly focus on two newly published protocols for the cryopreservation of zebrafish (Danio rerio) and fruit fly (Drosophila melanogaster) embryos.

Biography: John Bischof works in the area of thermal bioengineering with a focus on biopreservation, thermal therapy, and nanomedicine. His awards include the ASME Van Mow Medal, and Fellowships in societies including Cryobiology, JSPS, ASME, and AIMBE. He has served as the President of the Society for Cryobiology and Chair of the Bioengineering Division of the ASME. Bischof obtained a B.S. in Bioengineering from U.C. Berkeley (UCB) in 1987, an M.S. from UCB and U.C. San Francisco in 1989, and a Ph.D. in Mechanical Engineering from UCB in 1992. After a Post-doctoral Fellowship at Harvard in the Center for Engineering in Medicine, he joined the University of Minnesota in 1993. John Bischof is a Distinguished McKnight University Professor, Kuhrmeyer Chair in the Department of Mechanical Engineering, and the Medtronic-Bakken Endowed Chair and Director of the Institute for Engineering in Medicine at the University of Minnesota. John Bischof is also Director of the NSF Engineering Research Center Advanced Technologies for Preservation of Biological Systems (ATP-Bio), which launched on September 1, 2020.

Abstract:

Rapid advances in DNA synthesis techniques have made it possible to engineer diverse genomic elements, pathways, and whole genomes, providing new insights into design and analysis of systems. The synthetic yeast genome project, Sc2.0 is well on its way with the 16 synthetic Saccharomyces cerevisiae chromosomes now >99% completed by a global team. The synthetic genome features several systemic genome modifications but surprisingly, yeast strains with synthetic chromosomes are generally very healthy. Remarkably, the 3D structures of synthetic and native chromosomes are very similar despite the substantial number of changes introduced.
We have automated our big DNA synthesis and assembly pipeline (the GenomeFoundry@ISG), parallelizing the process, and can assembly human genomic regions of 100 kb along with multiple designer synthetic variants thereof. Using precision delivery strategies such as ICE and Big-IN, we can precision deliver DNA segments to stem cells, and use these methods to dissect the genomic “dark matter”, perform transplants of specific human genomic regions to animal genomes, and endow human cells with new capabilities.
Less than 2% of our genome is protein-coding DNA. The vast expanses of non-coding DNA make up the genome’s “dark matter”, where introns, repetitive and regulatory elements reside. Surprisingly, variation between individuals in non-coding regulatory DNA is emerging as a major factor in the genetics of numerous diseases and traits, yet very little is known about how such variations contribute to disease risk. Studying the genetics of regulatory variation is technically challenging, as regulatory elements can affect genes located tens of thousands of bp away, and often, multiple distal regulatory variations, each with a very small effect, combine in an unknown way to significantly modulate the expression of genes.
Genome wide association studies (GWAS) provide crude mileposts for the locations of human genome sequence variations that underlie disease susceptibility. However, they are like a first generation GPS - not too accurate. With the technology we are developing, we should be able to pinpoint the exact variant or variants that are responsible for disease susceptibility. Armed with this knowledge, we can accelerate turning this knowledge into therapies such as gene therapies or new drugs.
A progress report on several collaborative projects on different loci will be provided. Importantly, the Dark Matter Project actively seeks collaboration through its annual “locus nomination” mechanism. A Locus Nomination Board meets and chooses several loci annually for assembly, production of variants, and delivery to the appropriate cell type, typically a mammalian stem cell.

Biography: Jef D. Boeke, Ph.D., D.Sc., founded and directs the Institute for Systems Genetics at NYU Langone Health. From 1985-2014 Dr. Boeke was on the faculty at Johns Hopkins University School of Medicine. He is known for foundational work on mechanistic and genomic aspects of retrotransposition and his lab develops new technologies in genetics, genomics and synthetic biology. He elucidated a major form of mobile DNA, based on reverse transcription of RNA. He coined the term “retrotransposition” to describe this process, common to virtually all eukaryotic genomes and now studied by a worldwide scientific community. His systems-level studies helped elucidate intricate molecular mechanisms involved in retrotransposition in yeasts, mice and humans. In the area of Synthetic Genomics, his research group uses yeast as a platform for exploring the construction of fully synthetic chromosomes for practical and theoretical studies. He leads the international team synthesizing an engineered version of the yeast genome, Sc2.0, the first synthetic eukaryotic genome, and a consortium to explore the design and synthesis of even larger genomes. In 2018, he launched the “Dark Matter Project” (www.thedarkmatterproject.org) designed to better understand the “instruction manuals” that specify how human genes are expressed, using big DNA technology to build mammalian gene loci in yeast and then deliver those loci and their variants to stem cells. This has among other things to a technology to very rapidly design and deploy humanized mouse models. During the SARS-CoV2 pandemic, he led a team of volunteers to develop ultra high throughput coronavirus PCR testing, which led to the formation of the Pandemic Response Lab, the largest COVID-19 testing operation in New York City. Dr. Boeke has founded several biotechnology companies, including Avigen Inc., CDI Labs, Neochromosome, Inc. and the Pandemic Response Laboratory (Reopen Diagnostics, LLC) and serves on a number of Scientific Advisory Boards.

Abstract:
How can transferable biology and breeding contribute to improving food systems and climate change?
The demands of food production, fuels, nutrition, and climate change are going to require that thousands of species undergo genomic selection over the next two decades. We are using machine learning and statistical models of chromatin structure, regulatory grammar, cis-expression, protein stability, and deleterious mutations to improve transferable genome-wide predictions.

Biography: Edward S. Buckler is a USDA-ARS Research Geneticist and adjunct professor in Plant Breeding and Genetics at Cornell University with an educational background in molecular evolution and archaeology. His group's research uses genomic, computational, and field approaches to dissect complex traits and accelerate breeding in maize, sorghum, cassava, and a wide range of other crops. With these technologies applied to over 2000 species, now the Buckler group focuses on exploring ways to re-engineer global agricultural production systems to ensure food security, improve nutrition, and respond to climate change. With the USDA-ARS, he leads an informatics and genomics platform to help accelerate breeding for specialty crops and animals. His contributions to quantitative genetics and genomics were recognized with election to the US National Academy of Sciences and as recipient of the inaugural NAS Food and Agriculture Award.

Abstract:
How did we get here?
We now live in a world where it is not only possible, but relatively simple, to make intentional, targeted modifications in the genome of essentially any biological organism. It was not always like this. For many decades, geneticists relied on naturally occurring sequence variants. Even when the frequency of such variants could be increased in experimental or agricultural organisms by radiation or chemical mutagenesis, their location was essentially random, and any interesting mutation had to be separated from a background of confounding alterations. When gene targeting was demonstrated in the 1980’s, the frequency was so low that its utility was confined to just a few species. The situation began to change in the 1990’s as our understanding of DNA repair processes grew and new molecular tools emerged from basic genetic and biochemical studies. Our current genome editing platforms – ZFNs, TALENs and CRISPR – arose from such studies. The power we currently possess to modify genomes is awesome in both positive and negative respects. While the contributions of genome editing to progress in fundamental biology is exceptional, the projected positive benefits to humanity and the potential misuses have yet to be realized. It is our responsibility to ensure that uses of the technology are developed in ways that equitably engage and benefit the world community.

Biography: Dana Carroll is a distinguished professor in the Department of Biochemistry at the University of Utah School of Medicine. From 2019-2020, he was Interim Director of the Public Impact Program at the Innovative Genomics Institute at the University of California, Berkeley. Dr. Carroll’s research involves genome engineering using targetable nucleases. His lab pioneered the development of zinc-finger nucleases as gene targeting tools, and he continued working with the more recent TALENs and CRISPR/Cas nucleases, with much of the effort focused on optimizing the efficiency of these reagents for targeted mutagenesis and gene replacement. He received the Novitski Prize from the Genetics Society of America in 2012 and the Sober Lectureship Award from the American Society for Biochemistry and Molecular Biology in 2014. Dr. Carroll’s current interests include the societal implications of genome editing. He was a member of the international commission that released a report on Heritable Human Genome Editing in September, 2020. He received his Ph.D. from the University of California, Berkeley, and did postdoctoral research at the Beatson Institute for Cancer Research in Glasgow, Scotland, and at the Carnegie Institution Department of Embryology in Baltimore. He is a member of the US National Academy of Sciences, a fellow of the American Academy of Arts and Sciences and of the American Association for the Advancement of Science.

Abstract:
N/A

Biography:
Janice Chen is the co-founder and CTO of Mammoth Biosciences, a biotechnology company based in the San Francisco Bay Area harnessing a revolutionary gene-editing tool called CRISPR to deliver on the promise of precision medicine across diagnostics and therapeutics. Janice received her PhD from the lab of Nobel Laureate Professor Jennifer Doudna at University of California, Berkeley. She investigated mechanisms of CRISPR proteins and developed technologies leading to multiple papers and patents, and co-invented the programmable CRISPR-based detection technology called DETECTR.
Janice was selected as a Forbes 30 Under 30 in Healthcare, Business Insider's 30 Under 40 in Healthcare, Endpoints Top 20 Women in Biopharma, MIT Technology Review 35 Innovators Under 35 and delivered a TEDx talk on the potential for CRISPR to democratize diagnostics.

Abstract:
Regulatory Considerations for Gene Therapies incorporating Human Genome Editing

Biography:
Dr. Chery received her PhD in Molecular Biology, Cell Biology, and Biochemistry from Brown University. Her PhD thesis characterized a previously unstudied zinc finger protein identified by the lab, elucidating its function in DNA packaging and regulation of acetylation marks implicated in cancers such as breast cancer and leukemia. Previous to her graduate work, she obtained a Bachelor of Science in Chemistry and Bachelor of Arts in Biology from the University of Rochester. Dr. Chery did post-doctoral research at Harvard Medical School/Massachusetts General Hospital where she identified novel small molecules with therapeutic potential for Ebola virus. While there, she collaborated on projects developing small molecules such as antisense oligonucleotides (ASOs) and RNAi as therapeutic modalities for metabolic diseases. Pursuant to this, Dr. Chery became a research fellow at Dana Farber Cancer Institute where she developed a gene therapy approach for Very Early Onset Inflammatory Bowel Disease (VEO-IBD) using CRISPR and viral vector technologies. After her postdoc fellowship, Dr. Chery became a full-time CMC reviewer at the FDA.

Abstract:
Multiplex genome reading and writing & equitable access
Molecular multiplexing (via mixing, not parallelism) aided by native or synthetic barcoding has helped improve the cost and quality of genome reading 20 million-fold. To write well, we generally need to read. Multiplex writing enables: 1) screening libraries of genetic and epigenetic changes, one or a few per cell, one average; 2) Large numbers of edits per cell (per organism strain). Applications include 1a) creation of any cell type via a comprehensive TFome expression library (often in 4 days with 90+% conversion), 1b) CRISPRi/off/on,NHEJ libraries, 2a) 42 germ-line mutations for xenotransplants, 2b) a similar number for cold-resistant elephants to address carbon sequestration, 2c) Many desirable plant traits in one strain (Inari), Genome-wide codon recoding for resistance to all natural viruses (GPW). To achieve equitable access, multiplex is only part of the answer. We should prioritize cost-effective alternatives (e.g. genetic counseling 3000-fold more affordable than gene therapy), and ways to treat both rare and common diseases with a single (albeit multiplexed) treatment, e.g. aging reversal genome writing.

Biography: George Church is Professor of Genetics at Harvard Medical School and Director of  PersonalGenomes.org, which provides the world's only open-access information on human Genomic, Environmental & Trait data (GET). His 1984 Harvard PhD included the first methods for direct genome sequencing, molecular multiplexing & barcoding. These led to the first genome sequence (pathogen, Helicobacter pylori) in  1994 . His innovations have contributed to nearly all "next generation" DNA sequencing methods and companies (CGI-BGI, Life, Illumina, Nanopore). This plus his lab's work on chip-DNA-synthesis, gene editing and stem cell engineering resulted in founding additional application-based companies spanning fields of medical diagnostics ( Knome/PierianDx, Alacris, AbVitro/Juno, Genos, Veritas Genetics ) & synthetic biology / therapeutics ( Joule, Gen9, Editas, Egenesis, enEvolv, WarpDrive ). He has also pioneered new privacy, biosafety, ELSI, environmental & biosecurity policies. He is director of an IARPA BRAIN Project and NIH Center for Excellence in Genomic Science. His honors include election to NAS & NAE & Franklin Bower Laureate for Achievement in Science. He has coauthored 590 papers, 155 patent publications & one book ( Regenesis ).

Abstract:
Using CRISPR/Cas9 genome editing to understand coral symbiosis and bleaching
The symbiosis between corals and dinoflagellate algae is essential to the energetic requirements of coral-reef ecosystems. However, coral reefs are in danger due to elevated ocean temperatures and other stresses that lead to the breakdown of this symbiosis and coral "bleaching". Despite the importance of coral reefs, the molecular basis of how corals maintain a healthy symbiosis and avoid bleaching is poorly understood, in part because of the lack of a tractable genetic model system. The small anemone Aiptasia is symbiotic with algal strains like those in reef-building corals but has many experimental advantages, making it an attractive laboratory model for cnidarian symbiosis. To explore the transcriptional basis of heat-induced bleaching, we used RNAseq to identify genes that are differentially expressed during a time course of heat stress of symbiotic and aposymbiotic Aiptasia strains. We observed a strong upregulation of hundreds of genes at times long before bleaching begins in symbiotic anemones. The putative promoters of these early stress-response genes are enriched for binding sites for the NFkB and HSF1 transcription factors, suggesting that many of these genes share core transcriptional control. The overall expression patterns were similar between the symbiotic and aposymbiotic anemones, indicating that many of the expression changes are not specific to the presence of the algae.
Genetic tools are needed to allow rigorous functional testing of the roles of candidate genes in symbiosis and bleaching. Recently, we have developed methods for knocking down and overexpressing genes of interest in Aiptasia. Meanwhile, we have successfully used the CRISPR/Cas9 technology to create genetic changes in embryos of the coral Acropora millepora. We used this technology to knock out HSF1 and demonstrated its role in coral heat tolerance. Through the establishment of both gain-of-function and loss-of-function methods in both Aiptasia and corals, it will be possible to exploit the year-round spawning of Aiptasia to perform initial tests of gene function in cnidarian-algal symbiosis and then further test the discoveries made using similar technologies in corals.

Biography:
My principal scientific interest is applying molecular genetic approaches to answer long-standing questions of species-species interactions and evolution. My research focuses on the molecular and cellular bases of the symbiosis between corals and their intracellular dinoflagellate algae. This includes understanding how algal cells evade the host’s complex innate immune system to establish symbiosis and how the symbiosis is impacted by cellular stress. The mechanisms of this particular symbiosis are important for two main reasons. First, this symbiosis is critical for the survival of coral reefs, and its breakdown (or “coral bleaching”) due to anthropogenic stressors including climate change is leading to the global decline of coral ecosystems. The loss of these biodiversity hotspots will cause extensive economic and human health damage. Second, it is a dramatic example of an intracellular microbial relationship that benefits the animal host. Such interactions are poorly understood due to the lack of attention and tractable model systems.
Early in my career, I became fascinated by host-microbe interactions while researching coral at the Australian Institute of Marine Science (AIMS). While there, I realized that there was a gap in our understanding of coral genetics, and I saw an opportunity to merge my two passions, molecular and marine biology, to advance this field. Towards this long-term goal, I pursued my Ph.D. in molecular biology in Dr. Craig Miller’s lab. During my Ph.D., I applied novel quantitative genetics, genomics, and transgenic methods to identify DNA changes controlling morphological evolution in natural populations of stickleback fish. This work illustrated the types of genetic changes that generate new traits during vertebrate evolution.
Armed with the molecular training from my Ph.D., I sought to build genetic tools to study symbiosis as a postdoctoral researcher in Dr. John Pringle’s lab at Stanford. During this time, I established genetic methods, such as morpholinos and CRISPR-Cas9, in both corals and a model system for coral biology, the symbiotic anemone Aiptasia (Cleves et al. 2018). This approach has already allowed me to identify and test a heat-responsive genetic pathway that appears to confer thermal tolerance and protect against bleaching (Cleves et al. 2020a; b). Now, my lab focuses on determining the gene regulatory networks that are required for symbiosis.

Moderator, Wednesday Session 1: Genome Editing for Meeting 21st Century Needs

Biography:
Kevin Davies, Ph.D., is the author of EDITING HUMANITY: The CRISPR Revolution and the New Era of Genome Editing (Pegasus Books, 2020). Kevin’s latest book is the riveting story of the development of the Nobel Prize-winning technology for editing genes, driving breakthroughs in science, medicine, and agriculture, while igniting ethical controversies about designer babies and the future of humanity. Kevin won a Guggenheim Fellowship for science writing in 2017.
Kevin is the founding editor of Nature Genetics and currently the Executive Editor of The CRISPR Journal. His previous books include Breakthrough: The Race for the Breast Cancer Gene; Cracking the Genome (translated into 15 languages), an inside account of the race for the Human Genome Project hailed by one reviewer as “A rollicking good tale about an enduring intellectual monument”; and The $1000 Genome, which details the revolution in personalized medicine and consumer genetics. He also collaborated with Nobel laureate Jim Watson and Andrew Berry on DNA: The Story of the Genetic Revolution.

Abstract:
Precision breeding in Agriculture to meet 21st century global needs
Many people in wealthy countries think of agriculture mainly as a source of nutritious food. However, for several hundred million people in low and middle-income countries (LMICs), agriculture is also the main source of income for their families. In the 20th century, investments in agriculture R&D greatly increased farm productivity and contributed to major reductions in poverty and hunger. But globally, this progress has been uneven. Today, regions that are still struggling with high rates of poverty and hunger also are the areas where most people work in smallholder agriculture. Their farm productivity has lagged behind other countries and they now also face a host of new challenges caused by climate change. To meet the 21st century needs, the focus now needs to be on innovations and practices that simultaneously improve productivity, resource efficiency (the conversion of water, land and feed into food), and resilience to stresses like heat, drought, pests and disease.
Many countries are working to achieve these goals via precision agriculture, where farms are managed with the help of new technologies like sensors, satellite imagery and advances in data science to make sure that plants and animals get precisely the resources and treatments they need. Similarly, new technologies have created the opportunity to breed crops and livestock much more precisely. Early breeding programs only relied on phenotypic field data and pedigrees to select new varieties. Low-cost genotyping with SNP arrays has now made the selection process faster and more accurate. There is now also growing interest in the use of gene editing to efficiently develop, in a single generation, more productive, sustainable genetic varieties that are exactly aligned with local conditions.
There remain key technical challenges with product development through gene editing. They include insufficient knowledge of the genetic architecture of traits and the fact that most performance traits are controlled by many genes, each contributing only a small effect. Nevertheless, there are already promising gene editing projects that can lead to high-performing, locally-adapted crop varieties and livestock breeds resistant to harmful diseases and tolerant to stressors like heat.
Other key obstacles remain around public acceptance and global access. Public acceptance of gene editing demands that scientists do the hard work of providing evidence that the products of gene editing are safe—for humans, the environment, and, when livestock are involved, for the animals themselves. Meanwhile, the benefits of this technology must be available equally to farmers everywhere, not just to those in wealthy countries. These advances could be particularly important for smallholder farming communities in LMICs where agriculture innovation is inseparable from economic progress.

Biography:
Alfred de Vries works at the Bill & Melinda Gates Foundation as Senior Program Officer for Animal Production. He leads the Foundation’s efforts in R&D for Animal Production (genetics, reproduction, feed) aimed at increasing livestock productivity in Sub-Saharan Africa and South-Asia. Alfred has extensive experience in animal breeding across many geographies from his time at international breeding companies (CRV, Topigs Norsvin and PIC). He had management positions in R&D, technical service and operations. He obtained his MSc and PhD degrees in Animal Sciences from Wageningen University and holds a Global Certificate in Management from INSEAD.

Abstract:
Pioneering Precision in Gene Editing in Pisces
Pisces like the zebrafish (Danio rerio) are amazing model systems for genome engineering technology development. Using the rapid iteration possible from a 1 mm diameter single cell system, the Ekker lab has recently developed three new tools: 1) MENdel algorithm for predicting MMEJ-based precision INDEL edits 2) DonorGuide modified HDR substrate and localization method for SNP and INDEL edits and 3) FusXTBE TALE Base Editor and TALE Writer informatics tool for the rapid programming of mitochondrial DNA. Each of these approaches share two outcomes: a) enhanced precision in genome writing and b) work cross-platform, from zebrafish in vivo to human cells in vitro. Although primarily tested in vertebrates, these tools when assessed show robust function in other systems as well, supporting the longer-term vision of moving from ‘genome vandalism’ to reproducible and predictable programming of the code of life.

Biography:
Dr. Ekker is Dean of Graduate School of Biomedical Sciences, Director of the Office of Entrepreneurship and Professor of Biochemistry and Molecular Biology at the Mayo Clinic. Dr. Ekker has been conducting genome engineering for 30 years. Dr. Ekker is Editor-in-Chief of the Zebrafish journal and was the Founding President of the Genome Writers Guild genome engineering society. Dr. Ekker is CEO of LifEngine Technologies Inc, co-founder of LifEngine Animal Health Laboratories Inc, co-Founder of DGI (acquired by Immusoft), and is Chairman of the Board and co-Founder of InSciEd Out Foundation, a non-profit focused on science education reform for a healthier world. Dr. Ekker received bachelor of science degrees (Genetics and Developmental Biology, Electrical Engineering) from the University of Illinois Urbana-Champaign where he conducted genome science work with the molecular evolutionist Dr. Carl Woese. Dr. Ekker earned a PhD in Molecular Biology and Genetics at the Johns Hopkins University and Howard Hughes Medical Institute. Dr. Ekker was the founding Director of the Arnold and Mabel Beckman Center for Transposon Research (now called the Center for Genome Engineering) at the University of Minnesota. Dr. Ekker’s laboratory has been continuously funded by the NIH for over 20 years and has co-authored over 170 publications. The Ekker laboratory has used zebrafish to pioneer the development of genome engineering tools including transposons, morpholino antisense oligonucleotides and targeted genome editing methods spanning nuclear and mitochondrial programming in human cells and other organisms.

Abstract:
Eco-cisgenic
Using ecotechnologies to edit the genes of wild organisms can effectively address serious environmental problems, but real-world attempts may fail due to mistrust or excessive regulatory risk. Instead of immediately building CRISPR-based gene drive systems that may not be sufficiently localized or evolutionarily stable and are unlikely to receive near-term regulatory approval, we are working to suppress the fecundity of invasive rodents by engineering males to disrupt genes required for female viability or fertility. The resulting rodents cannot readily spread to areas where they are not wanted, and could be made more efficient by adding a gene drive at a later date. Our work is overseen by the City of Cambridge Biosafety Committee and aims to one day conduct field trials on the MIT campus, but we also sought outside perspectives. By building relationships with the Maori of Aotearoa New Zealand, where the government is likely to favor use of whatever technologies for rodent controls that we develop, we learned of an approach that appears to build trust and support in communities across the world.

Biography: MIT Professor Kevin Esvelt leads the Sculpting Evolution Group in exploring evolutionary and ecological engineering. The creator of a synthetic ecosystem to rapidly evolve molecular tools, he is best known for inventing CRISPR-based “gene drive” systems capable of single-handedly editing wild species. Esvelt and his colleagues chose to publicly describe the technology and highlight the need for safeguards before testing it and demonstrating reversibility in the laboratory; his lab has since outlined numerous localization methods. The author of several relevant patents, he has called for gene drive to remain a not-for-profit technology. An outspoken advocate of sharing research plans to accelerate discovery and improve safety, Esvelt has led efforts to ensure that all ecological editing research is not only open but community-guided, and his team is currently working closely with communities in Massachusetts, Aotearoa New Zealand, and Uruguay on diverse ecological editing projects. His laboratory seeks to safeguard biotechnology against mistrust and misuse by pioneering new ways of visibly working with communities, inventing novel precautions, accelerating reliable countermeasure development, and applying cryptographic methods enable secure and universal DNA synthesis screening.

Biography: A Fellow of the Society of Actuaries and an Enrolled Actuary who works for Foster & Foster, Inc. in Naperville, IL. He has nearly 25 years of experience consulting employers on retirement plan issues. Jason earned a B.S. degree in actuarial science from the University of Iowa. He is the father of a boy, Will, with Adrenoleukodystrophy (ALD).

Abstract:
Creating Value for Farmers, Scientists and Society
Climate change, the COVID-19 pandemic, and price spikes of commodity grains have combined to limit progress on food security in the past 5 years after years of improvement. Climate change is predicted to have a greater impact on tropical regions around the world than in temperate zones, especially in those areas where smallholder farmers dominate production and are often using decades-old crop varieties. The share of income spent on food in low-income countries remains stubbornly high in select countries, at up 40 to 50%. COVID-19 has disrupted value chains around the world. The price of commodities has spiked upwards more regularly in the past 20 years, often due to relatively mild weather-related events, like drought or flooding, in bread baskets of the world. The good news is that improvements in agricultural productivity at the smallholder farmer level have a proven track record of reducing poverty and increasing resilience. And providing a steady stream of new genetics and improved agronomics for both the major crops and less-improved crops of the world is one of the best means of improving resilience and adaptation to, and even mitigating, climate change. How do we go about delivering more value to farmers, scientists, and society, and resume even greater progress on food security goals? Maintaining unencumbered sharing of germplasm and genetic sequence data, through publicly funded seed and data banks is a start. Capacity building of scientific, agriculture extension, and regulatory biosafety organizations in regions most at risk helps build social acceptance of and political will for innovation. Collaboration across geographies and between the public and private sectors, utilizing combined expertise, experience, and technology to deliver improved varieties has the potential to move more technology forward, more quickly, than if these organizations were working alone. A recent example outside the world of agriculture of a highly successful collaboration is the development of the COVID19 vaccine. Initial sequencing of the virus and variants occurred in labs around the world and was submitted to open access databases, allowing rapid development of diagnostic tools and eventually leading to vaccines in record time. Tremendous value was created through collaboration and capacity building. Making progress on hunger, malnutrition, and climate change is no less of a challenge which will only be met through a similar level of cooperation.

Biography: N/A
Jim Gaffney grew up on a farm in southwest Minnesota where he became familiar with the challenges and opportunities of corn, soybeans, and farrow-to-finish hog farming. He is a product of the land-grant university system, matriculating at the University of Minnesota, South Dakota State University, and University of Florida. In between undergraduate and graduate school, Jim ventured to Cameroon, Central Africa, as a member of the United States Peace Corps, where he worked at a technical agriculture school and in surrounding villages. After receiving a PhD in agronomy from the University of Florida, Jim spent the next 25 years of his career in the crop protection and seed industry, in research and development, technical services, marketing, and regulatory. While at Corteva Agriscience, Jim was able to translate his passion for African agriculture into a number of development projects to expand the use of industry technology and practices to the public sector and for the benefit of under-utilized crops. Corn and soybeans have been displaced by tef and cassava as Jim’s favorite crops. Jim is now employed by the United States Agency for International Development as a General Development Officer for Private Sector Engagement, where he devotes 100% of his time to the development and delivery of innovation, technology, and improved genetics to smallholder farmers.

Abstract:
Unlocking LoxP to Track Genome Editing In Vivo
The development of CRISPR associated proteins, such as Cas9, has led to increased accessibility and ease of use in genome editing. However, additional tools are needed to quantify and identify successful genome editing events in living animals. We developed a method to rapidly and quantitatively monitor gene editing activity non-invasively in living animals that also facilitates confocal microscopy and nucleotide level analyses. Here we report a new CRISPR “footprinting” approach to activate luciferase and fluorescent proteins in mice as a function of gene editing. This system is based on experience with our prior Cre-detector system and is designed for Cas editors able to target LoxP including gRNAs including SaCas9 and ErCas12a. These CRISPRs cut specifically within LoxP, an approach that is a departure from previous gene editing in vivo activity detection techniques that targeted adjacent stop sequences. In this sensor paradigm, CRISPR activity was monitored non-invasively in living Cre reporter mice (FVB.129S6(B6)-Gt(ROSA)26Sortm1(Luc)Kael/J and Gt(ROSA)26Sortm4(ACTB-tdTomato,-EGFP)Luo/J, which will be referred to as LSL and mT/mG throughout the paper) after intramuscular or intravenous hydrodynamic plasmid injections, demonstrating utility in two diverse organ systems. The same genome-editing event was examined at the cellular level in specific tissues by confocal microscopy to determine the identity and frequency of successfully genome-edited cells. Further, SaCas9 induced targeted editing at efficiencies that were comparable to Cre recombinase demonstrating high effective delivery and activity in a whole animal. This work establishes genome editing tools and models to track CRISPR editing in vivo non-invasively and to fingerprint the identity of targeted cells. This approach also enables similar utility for any of the thousands of previously generated LoxP animal models.

Biography: N/A

Protein-DNA conjugation for genome and cellular engineering applications

Biography:
Wendy Gordon is currently an Associate Professor in the Department of Biochemistry, Molecular Biology, and Biophysics at University of Minnesota and a Pew Biomedical Scholar. She received her PhD in Physical Chemistry from University of Chicago and performed postdoctoral work in the area of Notch signaling/structural biophysics with Steve Blacklow at Harvard Medical School. Her lab focuses on developing technologies to study how cells sense and respond to altered mechanical forces at a molecular level during disease pathogenesis and studies cell-surface receptor proteolysis as a mechanism of communicating mechanical stimuli. Her lab also uses HUH-endonucleases, which form covalent bonds with ssDNA, to develop mechanobiology tools as well as application to genome engineering. Outside of science, she likes to hang out with her family and ride her Peloton bike.

Biography:
Perry Hackett passed through Stanford (BS, Physics), the University of Colorado (PhD Biophysics & Genetics) and postdoctoral stints in virology at the Max Planck Institute for Cell Biology (Germany) and the University of California, San Francisco before coming to the University of Minnesota in 1980 where he is a professor of Genetics, Cell Biology and Development, a co-founder of the Center for Genome Engineering and Founder/Director of the Biotechnology Initiative for Innovation, Collaboration and Entrepreneurship. His research since 1964 has focused on molecular genetics and genome engineering. The Sleeping Beauty Transposon System was invented in his lab for non-viral gene delivery to vertebrate genomes and is in clinical trials for human CAR T-cell therapy. He is a co-founder of three genome engineering companies - Discovery Genomics (gene therapy), Recombinetics (genetically modified large animals), and NovoClade (genetics-based pest control).

Abstract:
N/A

Biography:
Jennifer Hamilton is a Jane Coffin Childs postdoctoral fellow working with Dr. Jennifer Doudna to advance CRISPR-based gene editing therapeutics for use in humans. Dr. Hamilton did her PhD training in virology at the Icahn School of Medicine at Mount Sinai. Advised by Dr. Peter Palese, she studied host responses to influenza virus infection and the engineering of RNA viruses for novel uses, such as killing cancerous cells.
She is now leveraging her background in virology to co-opt viral infection strategies to more effectively deliver CRISPR-Cas9 tools to disease- relevant cells. Since the beginning of the COVID-19 pandemic, Dr. Hamilton has also co-lead the technical development of the Innovative Genomics Institute’s SARS-CoV-2 testing lab, which provides free clinical testing for first responders and underserved populations in the Bay Area, as well as asymptomatic screening for the UC Berkeley campus.

Abstract:
Unpacking the public discours around advances in food &agriculture
The survival of our species has always depended on advances in food and agriculture. If few people dispute this statement, then why do we have so much conflict, confusion, and distrust in discussions around food and agriculture? The good news is our global food and agricultural systems are the best they have been in the history of humanity. More people have access to safe and nutritious food than ever before. But at the same time, there are still huge problems: Too many people go hungry every day, and issues like obesity are real and daunting.
While it is healthy to be skeptical, if people are presented with overwhelming evidence that should alleviate that skepticism and they still don’t change their minds, then they are no longer skeptics; they have become denialists. And that is not a helpful position. At a time when misinformation has lead to both the Capitol riots on January 6th, and large numbers of people denying the Covid-19 vaccine, please join us with a conversation with Scott Hamilton Kennedy, as he discusses how to deal with misinformation, and even change people’s minds through the power of science and storytelling.
We are witnessing the rise of a “post-truth” era where “alternative facts” may threaten our ability to innovate and thus survive one of the biggest challenges humans have ever faced: How are we going to feed over nine billion people by 2050? And while there isn’t one perfect road that will get us there, the rigorous application of science and storytelling is the best tool we have to help us chart a safe course through an uncertain future.

Biography:
Academy Award nominee Scott Hamilton Kennedy is a writer, director, producer, cameraman, and editor.
His documentary work includes the Oscar nominated The Garden, and Independent Spirit Award Nominee OT: our town which tells the underdog story of the attempt to produce Thornton Wilder’s Our Town as first play performed by students in twenty years at Dominguez High in Compton, California. Among its many awards, OT: our town received a Human and Civil Rights award from the National Education Association (NEA).
Scott’s latest documentary, narrated by the esteemed astrophysicist and science communicator Neil deGrasse Tyson, is FOOD EVOLUTION, which, through resetting the GMO controversy, highlights the importance of using the scientific method to help everyone – from parents to politicians – make better decisions. FOOD EVOLUTION was both one of the most controversial and well-received documentaries of 2017, garnering a rare 100% on Rotten Tomatoes, where the LA Times wrote: "Calm, careful, potentially revolutionary, FOOD EVOLUTION is an iconoclastic documentary on a hot-button topic,” and the NY Times: "FOOD EVOLUTION posits an inconvenient truth for organic boosters to swallow: In a world desperate for safe, sustainable food, GMOs may well be a force for good." It has been screened on Capitol Hill, the National Academy of Sciences, the European Parliament, the FAO, and many more.

Abstract:
Preclinical Considerations for Gene Therapies Featuring Genome Editing Applications
Due to the advances in genome editing (GE) technology over the past decade, interest in its use as a therapeutic modality for the treatment of human disease has been increasing. However, the introduction of GE technologies in gene therapy products raises unique concerns that include off-target editing, chromosomal translocations, and genomic instability. Thus, the translation of these gene therapy products to clinical trials requires comprehensive characterization of product risks and how they can be mitigated. This presentation will describe the regulatory framework to preclinically assess the safety and activity profiles of genome-editing or edited therapeutic products to enable administration in early phase clinical trials. This presentation will provide a general overview of CBER/OTAT considerations for preclinical proof of-concept and safety assessment of these novel gene therapy products.

Biography: Dr. Shana Hardy serves as a Pharmacology/Toxicology Regulatory Review Scientist in the Office of Tissues and Advanced Therapies (OTAT) in the U.S. Food and Drug Administration (FDA) Center for Biologics Evaluation and Research (CBER). This office evaluates and approves innovative biological products with curative potential. Dr. Hardy reviews toxicological and pharmacological data submitted to support investigational cell and gene therapies. Her areas of expertise include oncology-based therapies, genome-editing technologies, and edited cell products. Since serving as a reviewer at the FDA, Dr. Hardy has received two Division of Clinical Evaluation and Pharmacology/ Toxicology (DCEPT) Excellence in Scientific Review Awards. Prior to joining FDA, Dr. Hardy was a National Academies of Sciences, National Research Council Postdoctoral Fellow at the National Institute of Standards and Technology (NIST). Her research focused on establishing a reliable measurement assay and infrastructure for monitoring the on-target and off-target activity of genome editing technologies. Dr. Hardy completed her PhD in Molecular Pharmacology at Purdue University, where her research focused on the characterization of the tyrosine kinase SYK molecular activity in breast cancer epithelial-to-mesenchymal plasticity and metastasis. She earned her BS and MS in biology at Tuskegee University.

Abstract:
Benefits and pitfalls of genetic biocontrol methods for invasive reptiles in the Greater Florida Everglades
Invasive reptiles, and notably Burmese pythons, are detrimental to the ecosystem and the restoration of the Greater Everglades, a RAMSAR site of international significance. There is increased focus and funding by the Federal and State governments to better manage this species, and to expand research on possible detection and control techniques. Invasive pythons have been implicated in the severe loss of mammal populations in southern Florida, with greater than 90% reduction reported in some species. Due to their cryptic coloration and behavior, visual detection and trapping to aid in the management of these giant constrictors is severely limited by a less than 1% detection rate.
Without effective detection and control efforts, the invasive python population continues to expand northward, and genetic evidence has been indicated for rapid adaptation to the colder Florida climate. Environmental DNA (eDNA) methods have been employed to detect python eDNA in the Loxahatchee National Wildlife Refuge since 2014. This area is further north than the population has been physically sighted or is expected to be, based on biological assessment. Due to the expanding detrimental environmental impacts to the Florida ecosystems and lack of options for detection and control of Burmese pythons, all alternative biocontrol options are being explored.
Advances in synthetic biology have led to the development of novel biocontrol techniques ranging from CRISPR-modulated artificial gene drives, RNAi-based toxicants, to the use of viral vectors – all of which enable species-specific targeting. These methods are not dependent on human detection, which is currently the only effective way to remove these invasive species. Moreover, some genetic biocontrol approaches have the potential to self-propagate, thus imparting landscape management solutions with little to no risk to non-target organisms. However, while potentially powerful, these technologies are complex and require a large degree of proper permitting, regulation, engagement, communication, and logistical facilitation. As part of our interdisciplinary teams, social scientists will be engaged to characterize stakeholder concerns, levels of support for the various methods, and communication plans.
Our current investigations into the responsible implementation of these innovative tools is focused on assessing their feasibility, without any proposed release of genetic material or live animals. Initial work centers on cataloging the state of the science and the generation of genetic and genomic sequences that could facilitate population suppression. Research in the control of pests and invasive species is rapidly advancing, but most of the work has been limited to addressing invasive mammals and disease in arthropods. More work in reptilian models will be essential to develop tools to assess and measure the associated risks and projected effectiveness of various modes of genetic biocontrol.
Last, the likelihood is high that genetic biocontrol projects will be proposed by other public or private entities, and the Department of the Interior may be called upon to review or advise on proper protocols. Our involvement in this rapidly advancing field will keep us at the forefront of these new technologies and provide the expertise necessary to become an authority capable of providing collaboration, review, and risk assessment.

Biography: Dr. Maggie Hunter directs the Conservation Genetics laboratory at the U.S. Geological Survey, Wetland and Aquatic Research Center and is faculty at the University of Florida. She works to integrate genetic tools with management and monitoring efforts for imperiled species recovery and management of invasive species. Her research utilizes methods such as conservation genetics and genomics, environmental DNA (eDNA), and immune and disease assessment in invasive and imperiled species. She is particularly focused in improving the uptake of science into practical applications in wildlife management. Most recently her work has focused on genetic biocontrol methods to control invasive species in the Florida landscape. She is a co-founder of the Group on Earth Observations Biodiversity Observation Network (GEO BON) Genetics Composition Working Group. She is also the co-chair for the UN Environments Program International Union for Conservation of Nature, Conservation Genetics Specialist Group in North America.

Abstract: N/A
Investigations of the cellular mechanisms of base editing
Base editors (BEs) are double-strand break (DSB)-free genome editing agents that install point mutations with high efficiency and specificity. Because of their reliance on uracil and inosine DNA damage intermediates, it has been hypothesized that BEs are more ubiquitous than precision DSB-reliant genome editing methods, which require DNA repair processes that are only active during certain phases of the cell cycle. I will describe in my talk the first systematic study of the cell cycle-dependence of base editing via cell synchronization experiments. These experimends reveal that nickase-derived BEs (which introduce DNA backbone nicks opposite the uracil or inosine base) function independently of the cell cycle, while non-nicking BEs are highly dependent on cell cycle status. Additionally, I will describe our analysis of the cell cycle-dependence of DNA damage repair genes in single-cell RNAseq datasets. A comparison of these analyses with our synchronization data suggests that nickase-derived BEs rely on the long-patch base excision repair pathway to install point mutations.

Biography:
Alexis received her B. S. degree in chemistry from the University of California, Berkeley in December of 2008. She then joined the lab of Jacqueline K. Barton at the California Institute of Technology for her doctoral studies. While at Caltech, she worked as an NSF Graduate Research Fellow on the design, synthesis, and study of DNA mismatch-binding metal complexes and received her Ph.D. in 2014. She pursued postdoctoral work as a Ruth L. Kirschstein NIH Postdoctoral Fellow in the laboratory of David R. Liu, where she developed base editing, a new approach to genome editing that enables the direct, irreversible chemical conversion of one target DNA base into another in a programmable manner, without requiring double-stranded DNA backbone cleavage. Alexis joined the Department of Chemistry and Biochemistry at the University of California at San Diego in 2017, where her lab develops and applies new precision genome editing techniques to the functional genomics field.

Abstract:
Translating Polymeric Vehicles Between Ribonucleoprotein and Plasmid DNA Cargoes: Do the Same Design Rules Apply?
Synthetic polymers are rapidly emerging as promising alternatives to viral delivery of therapeutic nucleic acid payloads such as DNA, RNA and ribonucleoproteins (RNP). Whether structure-function trends that are identified for a specific nucleic payload can be reapplied to other cargo types remains an unresolved question. In this work, we identified effective vehicles for plasmid DNA (pDNA) by screening a chemically diverse polymer library consisting of 43 unique structures. We discovered that a polymer previously identified for RNP delivery (P38) also emerged as the lead structure during screening studies for pDNA delivery, suggesting considerable overlap in the design rules for RNP and pDNA payloads. Through statistical learning tools we identified structural drivers for internalization, toxicity and transfection efficiency for both RNP and pDNA payloads and established that pDNA delivery is less reliant on hydrophobic interactions than RNPs. In addition to mediating transient transfection efficiently, P38 also co-delivered RNPs and pDNA donors to effect homology-directed repair (HDR) at higher rates than JetPEI. Our approach yielded a versatile transfection reagent that delivers multi-modal cargoes to accomplish challenging therapeutic objectives such as HDR.

Biography:
Ramya obtained her B.E. (Hons.) in chemical engineering from BITS Pilani, India, and her PhD in chemical engineering at the University of Michigan, Ann Arbor, advised by Prof. Joerg Lahann. At Michigan, she received a Rackham Predoctoral fellowship, the Procter & Gamble Team Innovation award, and the Richard & Eleanor Towner Prize for creative and innovative teaching. Ramya is also an ACS PMSE Future Faculty awardee. Currently, Ramya is a postdoctoral associate at the University of Minnesota, Twin Cities where she has been developing materiomics workflows to accelerate polymeric vector discovery at Prof. Theresa Reineke’s group for the past three years. In January 2022, Ramya will begin her independent career as an Assistant Professor in the Department of Chemical Engineering at the Colorado School of Mines. Her lab will develop materiomics frameworks to discover (1) polymeric gene delivery vehicles and (2) polymer coatings that regulate biomolecular interactions transpiring at interfaces between living systems and synthetic materials.

Abstract: N/A
Gene Editing, Public Trust, and Societal Responsibility
Biotech developers are concerned about the future of gene editing having experienced the contentious history of first-generation GM foods. They have also expressed desires to do better with public engagement in gene-editing innovation, hoping to engender future public trust and acceptance. The framework of Responsible Research and Innovation (RRI) may provide a pathway forward to act on their desires for greater public legitimacy. However, in the U.S. there has also been reluctance to incorporate RRI practices into biotechnology innovation systems given the significant and real barriers posed to researchers, developers, and regulators for doing so. This talk will address two primary and related areas: factors leading to public trust in gene editing innovation systems and opportunities and challenges for incorporating RRI practices into biotechnology research and development. A body of social and policy sciences research will be reviewed to examine 1) factors that lead to public trust in and acceptance of emerging technologies; 2) stakeholder attitudes towards RRI principles and practices; 3) barriers to implementing RRI in biotech innovation systems; and 4) proposed governance models to incorporate the principles of RRI into gene editing innovation systems. This work will be synthesized to suggest lessons and opportunities for moving forward with gene editing in more productive and societally responsive ways.

Biography:
Dr. Jennifer Kuzma is the Goodnight-NCGSK Foundation Distinguished Professor in the School of Public and International Affairs, and co-founder and co-director of the Genetic Engineering and Society (GES) Center at NC State University. Kuzma’s research interests involve the integration of social, policy, and natural sciences for the governance of emerging technologies, including risk analysis, public perceptions and trust, and policy processes. She has received several awards for her research and policy contributions. In 2014, she received the SRA Sigma Xi Distinguished Lecturer Award for recognition of her contributions to the field of risk analysis, and in 2017-2018 she was awarded the Fulbright Canada Research Chair in Science Policy at the University of Ottawa. In 2019 she was elected a Fellow of the American Association for the Advancement of Science (AAAS) for distinguished translational work in bridging the bench and society, advancing anticipatory governance of new technologies, and contributions to methods for oversight policy analysis. In 2020, she received the NC State Alumni Association Outstanding Research Award and was elected to the Research Leadership Academy. Kuzma has held several local, national and international leadership positions, including a member of World Economic Forum Global Futures Council on Technology, Values and Policy; the U.S. National Academy of Sciences’ Committee on Preparing for Future Biotechnology Products, Society for Risk Analysis (SRA) Council Member and Secretary, chair of the Gordon Conference on Science & Technology Policy, Member of the US FDA Blood Products Advisory Committee, AAAS-American Bar Association National Council of Scientists and Lawyers, AAAS Societal Implications Section X Electorate Nomination Committee, MN Governor’s Bioscience Advisory Committee, and a Member of the UN WHO-FAO Expert Group for Nanotechnologies in Food and Agriculture. She is interviewed frequently in the media for her work and expertise in technology policy, including the New York Times, Science, The Scientist, Nature, NPR, Washington Post, Scientific American, Boston Globe, PBS Nova, Wired, and ABC and NBC News. Prior to her current position, Kuzma was associate professor of science and technology policy at the Humphrey School of Public Affairs, University of Minnesota (2003-2013). Here she was area and degree program chair of Science, Technology and Environmental Policy; Associate and Interim Director of the Center for Science, Technology, and Public Policy; co-PI of the NSF-IGERT for Risk Analysis for Introduced Species; and Associate Director of the Initiative for Renewable Energy and the Environment. Before that (1997-2003), she served as program director and study director for several U.S. National Academy of Sciences reports related to biotechnology governance and bioterrorism and as AAAS Risk Policy Fellow at the USDA Office of Risk Assessment and Cost Benefit Analysis. She obtained her Ph.D. in biochemistry from the University of Colorado Boulder in 1995. Here she discovered bacterial isoprene production and holds the first patent for methods for bacterial production of isoprene (bioisoprene). Her postdoctoral work was completed at the Rockefeller University in New York where she worked on plant drought and salinity tolerance mechanisms, leading to an article in the journal Science.

Abstract:
Non-Viral Approach Enhances Large Transgene Insertion Efficiency for B-Cell Engineering
B cells have the unique ability to differentiate into long-lived plasma cells, producing a large quantity of protein (aka antibody) for many years, and even decades. These characteristics have generated great interest in the use of B cells as a novel cell-based therapy for protein delivery to treat enzymopathies. We previously developed and optimized a robust protocol for engineering B cells using CRISPR/Cas9 and a recombinant AAV (rAAV) vector, namely viral-based engineering approach. Moving forward, we tested this approach for engineering B cells to express a therapeutic enzyme to treat an enzymopathy. We chose mucopolysaccharidosis type I (MPS I) as a disease model. MPS I is a genetic disease caused by mutations in the gene responsible for producing the a-L-iduronidase (IDUA) enzyme. We constructed a cassette comprising a therapeutic cDNA of IDUA that is co-expressed with RQR8 (an engineering marker that is not normally express in B cells) driven by MND promoter. Due to the large cargo size of this cassette, the homology arms (HAs) were 250 base pairs (bp) to meet rAAV vector capacity (4.7 kb). The 250bp HAs in this rAAV produced low knock-in (<5%) frequency when compared to our previous reports using 1kb HAs (>50%). Our finding emphasizes a major drawback associated with the AAV viral-based approaches. In addition, immune responses leading to clearance of the rAAV-transduced cells has been reported. Furthermore, rAAV production is costly and turn-around time takes much longer when compared to plasmid DNA. These bottlenecks prompted us to explore a non-viral strategy using a plasmid donor as an alternative approach. To test the feasibility of using the non-viral approach, we introduced pMax GFP plasmid in B cells, resulting in 60% GFP-positive B cells. Next, we tested CRISPR/Cas9 mediated HDR using a nanoplasmidTM as a DNA donor molecule. We tested 3 different mechanisms to insert SA-GFP; HDR, HDR/HMEJ, and HMEJ. Surprisingly, we observed similar engineering frequencies in all 3 mechanisms, although the HMEJ approach severely affected B-cell viability (~30% viability) when compared to HDR (~40%). However, B cells from all methods started to rebound after 4 days. Moving forward we cloned the IDUA-RQR8 cassette with 250-bp HAs targeting the IgH locus into a nanoplasmidTM. Surprisingly, we observed up to 15% engineering efficiency using this approach, which was higher than the viral-based approach (5%). This data suggests that our non-viral approach appears to be particularly advantageous for large cargo integration with better engineering efficiency in B cells.

Biography: Kanut Laohrawee is a Ph.D. candidate in the Molecular Cellular Developmental Biology and Genetics program, University of Minnesota. He received his master's of Biological Sciences focused on using adeno-associated viral vector for in vivo gene therapy to treat a mouse model of mucopolysaccharidosis type II. He's currently training in Dr. Branden Moriarity 's lab, Department of Pediatrics, Medical School. As a Ph.D. student in Dr. Moriarity's lab, Kanut's work focuses on using CRISPR/Cas9 system to engineer primary human B cells to express a therapeutic enzyme and used the engineered cells as a novel, cell-based, enzyme-replacement therapy for a mouse model of hurler syndrome.

GWG President, Moderator, Friday Session 3: Powering the future - what does the future hold for GE?

Biography:
David Largaespada, PhD, is a full professor in the Departments of Pediatrics and Genetics, Cell Biology, and Development and the Associate Director for Basic Research in the Masonic Cancer Center at University of Minnesota. He is an authority on mouse genetics, gene modification, and cancer genes. He received his BS in Genetics and Cell Biology from the University of Minnesota, Twin Cities in 1987 and his PhD in Molecular Biology with Dr. Rex Risser at the University of Wisconsin-Madison in 1992. He did a post-doctoral fellowship at the National Cancer Institute working with world-renowned geneticists Dr. Nancy Jenkins and Dr. Neal Copeland, where the Leukemia and Lymphoma Society of America awarded him a post-doctoral fellowship. He joined the faculty of the University of Minnesota in late 1996. Dr. Largaespada currently holds the Hedberg Family/Children’s Cancer Research Fund Chair in Brain Tumor Research. He was awarded the American Cancer Society Research Professor Award in 2013, the highest award given by the ACS.

Abstract:
Gene-edited Pigs are Resistant to Multiple Major Epidemics
Every year, major livestock contagious diseases cause billions of significant economic losses in animal husbandry, seriously endangering human food supply and food safety. Especially, in the process of highly intensive pig farming, the disease problem is more prominent, mainly including PRRSV, ASF, TGEV and so on. Genetically Modification can greatly speed up genetic progress in the development of new disease-resistant breeding materials. Focusing on the major serious pig diseases, we used gene editing to generate new materials of pig breeding with more comprehensive disease resistance. We generated double-gene-knockout (DKO) pigs harboring edited knockout alleles for known receptor proteins CD163 and pAPN and show that DKO pigs are completely resistant to genotype 2 PRRSV and TGEV. We found no differences in meat-production or reproductive-performance traits between wild-type and DKO pigs but detected increased iron in DKO muscle. Additional infection challenge experiments showed that DKO pigs exhibited decreased susceptibility to porcine deltacoronavirus (PDCoV), thus offering unprecedented in vivo evidence of pAPN as one of PDCoV receptors. Beyond showing that multiple gene edits can be combined in a livestock animal to achieve simultaneous resistance to two major viruses, our study introduces a valuable model for investigating infection mechanisms of porcine pathogenic viruses that exploit pAPN or CD163 for entry. For minimally invasive gene modification, we further optimized the key sites of the interaction between pAPN protein and TGEV. The Immortal Pig epithelial (IPI-2I) cell lines with 69bp deletion of the 16 exon of pAPN, the 736, 738 and 736+738 amino acid were accurately edited respectively. TGEV challenge showed that TGEV infection in IPI-2I-PE cells with precise editing at site 736 of pAPN was significantly decreased, but not completely resistant to TGEV. IPI cells accurately edited at site 736+738 of pAPN can effectively resist TGEV infection. These data provide a theoretical basis for a more comprehensive understanding of the interaction mechanism between TGEV and pAPN and the preparation of accurate editing pigs.

Biography:
Kui Li has worked in the field of molecular genetics and animal breeding for 37 years, he served as a full Professor at Huazhong (Central China) Agricultural University, Institute of Agricultural Genomics at Shenzhen and Institute of Animal Sciences at Beijing of Chinese Academy of Agricultural Sciences (CAAS), respectively, since 1996. He was a visiting scholar at University of Sydney in Australia from 1997 to 1998, and a full-time visiting Professor at Cornel University in USA from 2001 to 2003. He served as the member of National GMO safety evaluation committee of China, and he was the host of major national GM projects from 2008 to 2020. He has been actively promoting the research and application of gene-edited animals in China. He has led his group generated Transgenic and gene editing pigs for food and medical use including PRRSV resistance gene-edited pigs (CD163), CD163 and pAPN double gene-edited pigs, MSTN (Myostatin) gene-edited pigs, Fat1 transgenic pigs, Phytase gene transgenic pigs, gene-edited pigs for Xenotransplantation, Multiple gene editing and translation into diabetes and cardiovascular disease models. He has published more 200 international papers in English.

Abstract:
Tuning Genes, Genomes, and Traits in Plants and Agriculture
Genome editing is being lauded as a revolutionary technology that will invigorate plant breeding for both mainstream and underutilized crops. However, crop domestication and improvement is founded on genetic complexity that goes well beyond gene mutations with dramatic changes in crop adaptation and improvement. We are dissecting the mechanisms and principles underlying quantitative trait variation, by applying genome editing to investigate the architectures of the regulatory DNA sequences that control the activity and phenotypic outputs from key developmental genes. We have found that these regulatory regions are highly sensitive to genetic changes and can serve as “tunable” transcriptional control regions, which can be manipulated to create novel genetic and quantitative trait variation that goes beyond what nature has provided. I will present several case studies from our work on tomato and related nightshade species that open the door to rapid improvement of orphan crops to enhance crop diversity and sustainability.

Biography:
Zach Lippman is a Professor of Plant Biology at Cold Spring Harbor Laboratory (CSHL) and a Howard Hughes Medical Institute (HHMI) Investigator. His research group integrates genetics, development, genomics, and genome editing to study flowering and flower production in nature and agriculture. Taking advantage of natural and induced variation in these processes in tomato and related Solanaceae plants, Lippman’s group has shown how stem cell proliferation and maturation underlie diversity in vegetative and reproductive shoot systems. Identifying the genes and mechanisms underlying this diversity have led to broader exploration on the roles of structural variation, gene regulation, and epistasis in development, domestication, and breeding. Based on these discoveries, Lippman is developing and applying innovative concepts and tools for crop improvement. His contributions to plant genetics and genome editing were recognized with receiving a MacArthur Fellowship and the US National Academy of Sciences Award in Food and Agriculture.

Abstract:
Harnessing genomics to fast- track genetic improvement in aquaculture
Aquaculture must be enhanced to meet the growing demand for seafood because natural fisheries has reached sustainable production since mid-1980s. Overfishing will eventually deplete our natural resources that otherwise should be renewable. In the last 30 years, major progress has been made with world aquaculture, making it grown to a level now accounting for approximately 50% of all seafood consumption. With a growing world population, declining natural fisheries and perhaps increasing demand for seafood per capita, it is apparent that aquaculture will soon become the primary source of fish and shellfish for human consumption. In this presentation, progress of genome research, especially whole genome sequencing, identification of “beneficial” or “harmful” genes with accurate sequence information, development of genome editing technology, as a proof of concept, will be reviewed. However, the bottleneck continues to be the lack of information for beneficial and harmful genes, and their expression and regulation in relation to performance and production traits. A set of key aquaculture performance and production traits, such as growth rate, feed conversion efficiency, disease resistance, processing yield, and harvestability, among other traits, have essentially not been tackled. Given the challenges of aquaculture with large number of species, difficulties for trait studies with the aquatic environment, and low funding, the challenges are hugely daunting. However, the science and technology framework is ready to fast-track genetic improvement in aquaculture, upon greater levels of public understanding and support.

Biography: A native of China, Dr. Zhanjiang (John) Liu has worked around the world as a scientist, educator, and leader. His areas of research expertise include aquaculture, biotechnology, genetics, genomics, and bioinformatics.
Dr. Liu is serving as Interim Provost and Vice Chancellor for Academic Affairs at Syracuse University. As the chief academic officer of the university, he oversees the internal affairs of the university and all the academic programs in its colleges and schools, as well as research. Working with the academic deans and faculty in the schools and colleges, the provost ensures the delivery of high quality academic programs and learning experiences for students. The Provost also works with senior vice presidents of other divisions, and various academic supporting units to advance the mission of the university. A “first-generation student,” he obtained his BS degree and graduated with distinction from the Northwest A&F University, majoring in Plant Protection in 1981. He obtained his MS and Ph.D. degree from Plant Pathology, and Molecular and Cellular Biology, in 1985 and 1989, respectively, both from University of Minnesota. He worked in the Institute of Human Genetics of the University of Minnesota Medical School as a research faculty and then as Director for R&D at National Biosciences, Inc. before he joined the faculty of Auburn University in 1995, where he worked through the ranks of assistant professor, associate professor, full professor, and distinguished Alumni Professor. He served as center/institute director, associate dean for research in the College of Agriculture, assistant director of the Alabama Agricultural Experiment Station, associate provost, and associate vice president for research at Auburn University. In 2017, he was recruited to serve as the Vice President for Research at Syracuse University, and in November 2019, he was appointed as Interim Provost and Vice Chancellor for Academic Affairs at Syracuse University.
As an international authority in the area of aquaculture genomics and bioinformatics, Liu has trained more than 100 Ph.D. students and postdocs, obtained more than 80 grants totaling more than $50 million, and published four books and more than 330 papers and book chapters. He was elected a Fellow of AAAS in 2007 and a Fellow of the World Aquaculture Society in 2017.

Abstract:
FDA’s approach to oversight of low risk IGAs in Animals
Dr. Lombardi will provide an overview of FDA’s science- and risk-based oversight of Intentional Genomic Alterations (IGAs) in Animals, including describing low risk IGAs in animals, for which FDA does not expect an approval application prior to marketing based on a determination of low risk. Dr. Lombardi will walk through the data review process FDA uses to determine whether an IGA is low risk and will present a case study outlining data expectations demonstrating that an IGA is low risk to humans, animals, and the environment for use as an animal model of disease. This information is intended to help developers understand the evaluation process for determining whether IGAs in animals are low risk. FDA will also share its future plans for further stakeholder engagement and provide an update regarding the GWG outreach session planned on 7/28 as well as other opportunities for stakeholders to share their feedback with CVM.

Biography: Dr. Heather Lombardi is the Director of the Division of Animal Bioengineering and Cellular Therapies in the Office of New Animal Drug Evaluation at the Food and Drug Administration’s Center for Veterinary Medicine (CVM). This Division reviews intentional genomic alterations in animals and animal cell, tissue, and cell- or tissue-based products. She has participated in many center and agency efforts to bring innovative products to market in a predictable and efficient manner, such as the Innovation Exploration Team (IVET), numerous technology teams and working groups, and through development of the Veterinary Innovation Program (VIP). Dr. Lombardi has developed guidance and policy specific to the regulation of new and emerging technologies. Dr. Lombardi holds a PhD in biological chemistry from the University of Pennsylvania and a BS in chemistry (biochemistry track) from the University of North Carolina at Chapel Hill

Moderatore Thursday Session 4: Session Four: Bio-entrepreneurship & early to mid-career researchers in genome engineering

Biography:
Mary is a Senior Manager in the Venture Center of UMN's Technology Commercialization office. She has an MBA from the University of St. Thomas, 25 years of high tech and medical device industry experience at 3M, Imation and Medtronic plus professional consulting and startup experience. She coaches startups and develops and executes programs to commercialize UMN technology. Programs include the University's only equity-based seed funding program, the Walleye Tank life science pitch competitions with Mayo Clinic, and Minnesota Ventures, an investor event to get Minnesota startups funded.

Abstract:
The Origin of Synthetic Species with Engineered Genetic Incompatibility
Speciation constrains the flow of genetic information between populations of sexually reproducing organisms. Gaining control over mechanisms of speciation may enable new strategies to manage wild populations of disease vectors, agricultural pests, and invasive species. Additionally, such control would provide safe biocontainment of transgenes and threshold independent gene drives.
We have developed a method for synthesizing speciation events in sexually reproducing organisms termed Engineered Genetic Incompatibility (EGI). An EGI strain is homozygous for well tolerated mutations in the promoter regions of dosage sensitive genes and a programmable transcriptional activator (e.g. dCas9-VPR) targeted to the wild-type promoter sequence. Mating between EGI and organisms with the wild-type promoter sequence results in offspring where the transcriptional activator binds to the wild-type sequence and drives lethal overexpression of the target gene.
We previously demonstrated EGI in the yeast Saccharomyces cerevisiae. Here we show that EGI is highly effective in a multicellular organism. Multiple EGI strains of D. melanogaster were engineered that are both 100% reproductively isolated from the wild-type and each other. Further, EGI can be tuned so that offspring between incompatible populations die at specific life-stages which has implications for population control programs.

Biography: Dr Maciej Maselko is a CSIRO Synthetic Biology Future Science fellow and group leader in Applied BioSciences at Macquarie University in Sydney, Australia. Prior to joining Macquarie University in 2019, Maciej received a PhD in Molecular Biology at Oregon State University where he worked on antiviral compounds followed by a postdoc at the University of Minnesota in Dr Mike Smanski's lab where he developed a method to engineer speciation events in sexually reproducing organisms. Maciej's research primarily focuses on insect biotechnology and genetic pest control systems.

Abstract:
The Origin of Synthetic Species with Engineered Genetic Incompatibility
Speciation constrains the flow of genetic information between populations of sexually reproducing organisms. Gaining control over mechanisms of speciation may enable new strategies to manage wild populations of disease vectors, agricultural pests, and invasive species. Additionally, such control would provide safe biocontainment of transgenes and threshold independent gene drives. We have developed a method for synthesizing speciation events in sexually reproducing organisms termed Engineered Genetic Incompatibility (EGI). An EGI strain is homozygous for well tolerated mutations in the promoter regions of dosage sensitive genes and a programmable transcriptional activator (e.g. dCas9-VPR) targeted to the wild-type promoter sequence. Mating between EGI and organisms with the wild-type promoter sequence results in offspring where the transcriptional activator binds to the wild-type sequence and drives lethal overexpression of the target gene. We previously demonstrated EGI in the yeast Saccharomyces cerevisiae. Here we show that EGI is highly effective in a multicellular organism. Multiple EGI strains of D. melanogaster were engineered that are both 100% reproductively isolated from the wild-type and each other. Further, EGI can be tuned so that offspring between incompatible populations die at specific life-stages which has implications for population control programs.

Biography: Dr Maciej Maselko is a CSIRO Synthetic Biology Future Science fellow and group leader in Applied BioSciences at Macquarie University in Sydney, Australia. Prior to joining Macquarie University in 2019, Maciej received a PhD in Molecular Biology at Oregon State University where he worked on antiviral compounds followed by a postdoc at the University of Minnesota in Dr Mike Smanski's lab where he developed a method to engineer speciation events in sexually reproducing organisms. Maciej's research primarily focuses on insect biotechnology and genetic pest control systems.

Abstract:
Self-cutting and integrating plasmids (SCIP) as an Inexpensive and Scalable Screening Method for Screening Genomic Integration Sites
Here, we investigated self-cutting and integrating CRISPR-Cas9 plasmids (SCIPs) as easy-to-use gene editing tools that insert themselves at CRISPR-guided locations. SCIPs demonstrated similar expression kinetics and gene disruption efficiency in mouse (EL4) and human (Jurkat) cells, with stable integration in 3-6% of transfected cells. As proofs-of-concept, we have used SCIP to 1) insert a gene fragment encoding tdTomato into the CD69 locus of Jurkat cells, thereby creating a cell line that reports T cell activation, and 2) insert a chimeric antigen receptor (CAR) gene into the TRAC locus. To expand on these studies we have established a high throughput methodology to generate SCIP plasmids targeting a range of genomic targets in Jurkat T cells. Although we see a wide range of integration efficiencies, we observe consistent transgene expression in all sites tested. As with our previous observations, transgene expression seems to correlate with the efficiency of sgRNA genome editing. We also have performed proof of concept experiments to demonstrate SCIP activity in pluripotent stem cells. Thus, we have thus demonstrated that SCIPs function as simple, efficient, and programmable tools useful for generating gene knock-out/knock-in cell lines that are amenable to high-throughput genome engineering.

Biography:

Abstract:
Yeast avatar for anti-infective drug screening
Antimicrobial resistance is a global health emergency. Despite this, the number of new antibiotics in development is limited and there is a desperate need for new pipelines to identify novel antibiotic candidates. We have developed a yeast-based drug-screening platform to be used in high-throughput compound screens to identify molecules with antimicrobial activity. We call this the yeast avatar platform. The overall strategy is to engineer the model organism Saccharomyces cerevisiae so its growth depends on the expression of sets of enzymes transplanted from human pathogens. By introducing entire enzymatic pathways, focusing on those that are essential in bacteria, fungus and plants but not in humans, the resulting yeast strains will provide an easy system for high-throughput drug screening and the opportunity to identify compounds with high therapeutic indices.

Biography: Leslie Mitchell is a co-founder of Neochromosome, Inc., a biotech startup whose disruptive chromosome synthesis and engineering technologies are changing the world of synthetic biology. Leslie has worked extensively on chromosome and genome engineering in both yeast and mammalian systems and helped lead the international Synthetic Yeast Genome Project, Sc2.0, aiming to build a designer yeast genome from scratch. Leslie completed her PhD at the University of Ottawa and a postdoc that started at Johns Hopkins University School of Medicine and ended at New York University Langone Health.

Abstract:
In vivo delivery of Cas9 ribonucleoprotein and donor DNA with gold nanoparticles
Cas9 based therapeutics have the potential to revolutionize the treatment of genetic diseases. However, safe and effective methods for delivering Cas9 protein, gRNA and donor DNA need to be developed before the therapeutic potential of CRISPR based therapeutics can be realized. In this presentation, I will describe a non-viral Cas9 delivery vehicle, termed CRISPR-Gold, which can induce homology directed DNA repair (HDR) in vivo by directly delivering Cas9 protein, gRNA, and donor DNA. CRISPR-Gold is composed of gold nanoparticles assembled with the Cas9/gRNA ribonucleoprotein (RNP) complex, donor DNA, and an endosomal disruptive polymer. We have been able to demonstrate that CRISPR-Gold can correct the DNA mutation that causes Duchenne muscular dystrophy (DMD) in mdx mice via HDR, with an efficiency of 5.4% after an intramuscular injection and with minimal levels off-target DNA damage. In addition, CRISPR-Gold was able to deliver Cas9 RNP into the brain, after an intracranial injection, and rescue mice from FMR1 related autism, via deletion of the mGluR5 gene via non-homologous end joining (NHEJ). CRISPR-Gold is a non-viral delivery vehicle that can generate HDR and NHEJ in vivo and has potential for treating muscle and neurological diseases

Biography: Dr. Niren Murthy is a professor in the Department of Bioengineering at the University of California at Berkeley. Dr. Murthy’s scientific career has focused on the molecular design and synthesis of new materials for drug delivery and molecular imaging. The Murthy laboratory developed the hydrocyanines in 2009, which are now one of the most commonly used probes for imaging reactive oxygen species and commercially available from multiple sources. The Murthy laboratory has developed several new nanoparticulate technologies for drug delivery, such as the polyketals, which have been used by numerous laboratories to enhance the delivery of small molecules and proteins. The Murthy laboratory has been recently focused on developing non-viral delivery vehicles that can deliver Cas9 protein, gRNA and Donor DNA in vivo. Dr. Murthy received the NSF CAREER award in 2006, and the 2009 Society for Biomaterials Young Investigator Award.

Wednesday Keynote Speaker. American novelist, nonfiction author, filmmaker, political strategist

Biography:
Shawn Otto speaks to audiences worldwide about the scientific foundations of democracy and the causes and dangers of anti-science authoritarianism. He is author of the award-winning nonfiction book The War on Science, which predicted the rise of anti-science authoritarians and the threat they pose to democracy. The book has been called "a game changer, and probably the most important book you'll read this year." He has advised candidate science debate efforts in many countries.
He also an award-winning screenwriter and novelist, including writing and co-producing the Academy Award-nominated movie House of Sand and Fog, and the LA Times Book Prize finalist literary crime novel, Sins of Our Fathers.
Otto was awarded the IEEE-USA National Distinguished Public Service Award for his work elevating science in American public dialogue. He is cofounder and producer of the US presidential science debates at ScienceDebate.org and the only person to get Donald Trump to answer science questions during the 2016 presidential campaign.
He lives in Minnesota with his wife, Rebecca Otto, the former Minnesota State Auditor and candidate for Governor, in a solar- and wind-powered green home he designed and the couple built with their own hands.

Abstract:
The power of echoes: How HLA anchors a rewritten futureFor decades, caregivers have pioneered an unsung front of personalized medicine: tissue transplant. But key shortcomings still let most patients die without well matched tissue. Genome editing can help solve such challenges (while raising new ones). But even as we begin editing our own cells to stay healthy, data pooled by millions of good-willed tissue volunteers, worldwide, can drive further discoveries on many fronts of health science, long into that bright future.

Biography:
Genomicist Nathan Pearson has long teamed with fellow scientists, coders, caregivers, and layfolk — from historian Henry Louis Gates, Jr., to journalist Carl Zimmer, and even rocker Ozzy Osbourne — to explore what DNA says about our health and history. Trained at Stanford and the University of Chicago, he led collaborative efforts at the New York Genome Center, before founding the personal immunogenomics company Root, to honor tissue donor volunteers with free, grounded personal insights from their own HLA genes. He writes at http://genomena.com, and tweets as @GenomeNathan.

Abstract:
The (Not So) Secret Life of a Science Writer
You had just published the best research ever and wanted the whole world to know about it. But the New York Times never called, and the one reporter you talked to did not write a story. Why? Many factors influence a reporter’s decision to cover a particular paper, scientist, or field. I will talk about those factors and about ways to influence those decisions. I will also discuss the editing process, the types of stories and media outlets, and the challenges science journalists face in doing their work. Using my own career as an example, I will cover how the rise of social media and misinformation and the financial pressures publications now face have conspired to make my job ever harder and may one day make it disappear.

Biography: Elizabeth Pennisi is a senior correspondent with Science with more than 25 years experience helping to shape the journal's news section and online news content. She writes about biology, focusing primarily on genomics, evolution, microbiology, and organismal biology, with a smattering of ecology and behavior thrown in. She joined the staff of Science in 1996 and added editing to her job duties in 2007. She has an undergraduate degree in biology from Cornell University and a master's degree in science writing from Boston University. In addition to Science, her byline has appeared in Science News--where she won the James T. Grady-James H. Stack Award for Interpreting Chemistry for the Public—The Scientist, and United Press International.
Although she loves being a science writer, Liz is most passionate about the outdoors and in particular about whitewater kayaking and outrigger canoeing.

Abstract:
Genome Editing of Cells to Create More Effective and Safer Cell Based Medicines

Biography: Matthew Porteus MD, PhD is the Sutardja Chuk Professor of Definitive and Curative Medicine and a Professor in the Department of Pediatrics, Institute of Stem Cell Biology and Regenerative Medicine and Maternal-Child Health Research Institute at Stanford. His primary research focus is on developing genome editing as an approach to cure disease, particularly those of the blood (such as sickle cell disease) but also of other organ systems as well. He received his undergraduate degree at Harvard in History and Science where his honors thesis studied the recombinant DNA controversy of the 1970s. He then completed his MD and PhD training at Stanford, clinical training in Pediatric Hematology/Oncology at Boston Children’s Hospital, and post-doctoral research training with Noble Laureate David Baltimore at CalTech. He works as an attending physician on the Pediatric Hematopoietic Stem Cell Transplant service at Lucile Packard Children’s Hospital where he cares for children under going bone marrow transplantation for both malignant and non-malignant diseases. His goal is to combine his research and clinical interests to develop innovative curative therapies. He served on the 2017 National Academy Study Committee of Human Genome Editing and currently serves on the Scientific Advisory Board for WADA on Cell and Gene Doping and the NIH NexTRAC advisory committee evaluating the emergence of new technologies.

Abstract:
Base Editing Strategies for Hemoglobin Variants
Hemoglobin (hHBB) variants are found throughout human populations. Over 1000 naturally occurring hemoglobin variants of single amino acid substitutions have been reported. HBB variants alter hemoglobin structure and biochemical properties and can have insignificant to severe physiological effects. The sickle cell variant (Hb S) in the homozygous state is responsible for sickle-cell anemia and is a form of severe hemoglobinopathy. Other hemoglobin variants such as Hb C and Hb E are mostly benign unless coinherited with another abnormal hemoglobin, in which case, serious hemoglobinopathies occur. Hb SC accounts for about 30% of sickle cell disease in the United States and more than 50% in parts of West Africa. Patients with Hb E/B-thalassemia account for approximately 50% of the cases of severe beta-thalassemia. Here, we present highly efficient base editing strategies for correcting both Hb C and Hb E hemoglobin variants in HUDEP2 cells. Moreover, we show high rates of editing in Cd34+ patient-derived Hb SC cells. Additionally, we have created and characterized humanized animal models of Hb SC and Hb EE that mimic the pathophysiology of the corresponding disease. We are using these cell and animal models to test and optimize base editing as a therapeutic option for correcting these hemoglobinopathies.

Biography: Shondra Pruett-Miller, Ph.D. is an Assistant Member in the Department of Cell and Molecular Biology, the Founding Director of the Center for Advanced Genome Engineering (CAGE), and the Associate Director of Shared Resoucres for the Comprehensive Cancer Center at St. Jude Children’s Research Hospital in Memphis, TN. Shondra completed her Ph.D. in Cell and Molecular Biology from The University of Texas Southwestern Medical Center in August 2008. While at UT Southwestern, she worked in Matthew Porteus’ lab on the optimization of zinc finger nucleases for use in mammalian cells. After graduate school, she was recruited to Sigma-Aldrich as a Senior Scientist in R&D working on their CompoZr ZFN technology. In 2012, she returned to academia as the Founder and Director of the Genome Engineering and iPSC Center (GEiC) at Washington University School of Medicine in St. Louis. In 2017, she joined the Faculty at St. Jude Children’s Research Hospital where she established and is directing the Center for Advanced Genome Engineering (CAGE). Shondra has overseen the creation of over 1000 custom edited clonal cell lines, more than 250 custom edited preclinical animal models, and over 75 custom pooled gRNA screens.

Abstract:
Development of tools for multiplexed plant genome engineering
Cas12a is a promising genome editing system for targeting AT-rich genomic regions. Comprehensive genome engineering requires simultaneous targeting of multiple genes at defined locations. To expand the targeting scope of Cas12a, we screen Cas12a orthologs and identify Mb2Cas12a with high editing efficiency at relaxed PAM sites. To enable large-scale genome engineering, we compare 12 multiplexed Cas12a systems and identify a potent system that exhibits nearly 100% biallelic editing efficiency with the ability to target as many as 16 sites in rice. This system will enable simultaneous knockout of many genes in plants within one generation. To achieve simultaneous activation of multiple genes, we develop a CRISPR-Act3.0 system through systematically exploring different effector recruitment strategies and various transcription activators based on deactivated SpCas9. The CRISPR-Act3.0 system results in four- to six-fold higher activation than the state-of-the-art (2nd generation) CRISPRa systems. With CRISPR-Act3.0, robust activation of up to seven genes in a metabolic pathway is achieved in rice. In addition, the CRISPR-Act3.0 allows simultaneous modification of multiple phenotypes in Arabidopsis, which are stably transmitted to the T3 generations. Altogether, these new CRISPR systems are useful additions to the plant genome engineering toolbox, aiding simultaneous loss-of-function and gain-of-function studies.

Biography:
Dr. Yiping Qi received B.S. in Microbiology from Nankai University, M.S. in Biochemistry and Molecular Biology from Shanghai Jiao Tong University, and PhD in Plant Biology from University of Minnesota, Twin Cities. From 2009 to 2013, he conducted postdoc research on plant genome engineering with ZFN and TALEN at Dr. Dan Voytas lab. Dr. Qi started his first lab in 2013 at East Carolina University in North Carolina. He is currently an Associate Professor at University of Maryland, College Park. His lab has developed multiple CRISPR systems for plant genome editing and transcriptome regulation. His recent recognitions include NSF Plant Genome Early Career Award, FFAR New Innovator in Food and Agriculture Research Award, SIVB Young Scientist Award, UMD-AGNR Faculty Research Award, and University of Maryland Invention of the Year in Life Science Category.

Abstract:
Efficient polymer-mediated delivery of gene editing payloads through combinatorial design, parallelized experimentation, and machine learning
parallelized experimentation, and machine learning Chemical delivery vehicles such as cationic polymers are tailorable alternatives to viral vectors for the delivery of genome-editing payloads. Clinical translation hinges on developing methods to rapidly explore vast chemical design spaces and derive structure–function relationships governing delivery performance. Our group has discovered a polymer for efficient intracellular ribonucleoprotein (RNP) delivery through combinatorial polymer design and parallelized experimental methods. A library of 43 chemically diverse polymers have been synthesized via combinatorial polymerization, realizing systematic variations in physicochemical properties. Cationic monomer building blocks have been selected that varied in their pKa values, size, and lipophilicity of their substituents. Comonomers of varying hydrophilicity were also incorporated, enabling elucidation of the roles of protonation equilibria (drives RNP binding and release) and hydrophobic–hydrophilic balance in the polymer vehicle properties and performance. The biological performance of RNP delivery was screened via image cytometry enabling rapid discovery of a high performing “hit” polymer formulation that outperforms all commercial transfection reagents screened, achieving nearly 60% editing efficiency via nonhomologous end-joining. Structure–activity relationships underlying editing efficiency, cellular toxicity, and RNP uptake were examined via machine learning to uncover the physicochemical basis of high performance of the hit polymer formulation. Although cellular toxicity and RNP uptake were solely determined by polymer-RNP complex size distribution and protonation degree, respectively, these two design parameters were found not to enhance editing efficiency. Instead, polymer hydrophobicity and the Hill coefficient, a parameter describing cooperativity-enhanced polymer deprotonation, were found to be critical determinants of editing efficiency. Combinatorial synthesis and high-throughput characterization methodologies coupled with data science approaches enabled the rapid discovery of a polymeric vehicle that would have otherwise remained inaccessible to chemical intuition. The statistically derived design rules elucidated in this work have promise to guide the synthesis and optimization of future polymer libraries tailored for specific tissues and therapeutic applications of RNP-based genome editing.

Biography:
Theresa M. Reineke is a Distinguished McKnight University Professor in the Department of Chemistry at The University of Minnesota. She also holds graduate faculty appointments in the Departments of Chemical Engineering/Materials Science and Pharmaceutics. She received a B.S. Degree from the University of Wisconsin-Eau Claire, a M.S. Degree from Arizona State University, and a Ph.D. from The University of Michigan. She then received a National Institutes of Health Postdoctoral Fellowship to further her research background at the California Institute of Technology prior to beginning her independent faculty career. Her research group is focused on enabling fundamental and applied technology advancements of polymers in the fields of gene therapy and genome editing, drug delivery, and sustainability. She has published over 160 peer-reviewed manuscripts, holds numerous patents, and manages a large group of researchers funded by several corporate, private foundation, and national funding agency grants. Reineke has received several awards, including the 2009 National Institutes of Health Director’s New Innovator Award, 2012 Outstanding New Investigator Award from the American Society of Gene and Cell Therapy, 2017 Carl S. Marvel Creative Polymer Chemistry Award from the American Chemical Society Division of Polymer Chemistry, and in 2018 was awarded the DuPont Nutrition and Health Sciences Excellence Medal. She is also a founding Associate Editor of ACS MacroLetters and currently on the Editorial Advisory Boards of the peer-reviewed journals Biomacromolecules, Bioconjugate Chemistry, Polymer Chemistry, and ACS Applied Polymer Materials.

Abstract:
NK cells: next generation cell therapies for cancer
Dr. Rezvani will discuss a new frontier in NK cell therapeutics: engineering NK cells with chimeric antigen receptors. She will discuss the opportunities and challenges of NK cell CAR engineering, and present pre-clinical and early phase clinical data on cord blood-derived NK cells expressing CD19 CAR and IL-15 to enhance their in vivo persistence in patients with relapsed or refractory blood cancers. In addition, she will discuss novel strategies for the gene editing of CAR NK cells to enhance their function by targeting immune checkpoints. Finally, she will discuss the approach of precomplexing NK cells with an anti-CD16 bispecific antibody targeting cancer targets to redirect their specificity, thus providing a rapid approach to translate NK cells with CAR-like characteristics to the clinic.

Biography: Katy Rezvani M.D, PhD is the Sally Cooper Murray Chair in Cancer Research, Professor of Medicine, Chief of Section for Cellular Therapy, Director of Translational Research and Director of the GMP Facility at MD Anderson Cancer Center. She also serves as the Executive Director of the Adoptive Cell Therapy Platform at MD Anderson. Her research laboratory focuses on the role of natural killer (NK) cells in mediating immunity against hematologic and solid tumors. The goal of this research is to understand mechanisms of tumor-induced NK cell dysfunction and to develop strategies to genetically engineer NK cells in order to enhance their in vivo anti-tumor activity and persistence. Findings from Dr. Rezvani’s lab have led to the approval and funding of several investigator-initiated clinical trials of NK cell immunotherapy in patients with hematologic malignancies and solid tumors, as well as the first-in-human clinical trial of off-the-shelf CAR-transduced cord blood NK cells in patients with relapsed/refractory lymphoid malignancies. Dr. Rezvani’s work is supported by multiple grants from the National Cancer Institute, the Leukemia and Lymphoma Society, the American Cancer Society, Stand Up to Cancer and the Cancer Prevention & Research Institute of Texas (CPRIT). Dr. Rezvani completed her medical training at University College London, followed by Fellowships of the Royal College of Physicians and the Royal College of Pathologists of the United Kingdom, a Ph.D. in Immunology from Imperial College London and postdoctoral studies at the National Institutes of Health.

Abstract:
The FusX TALE Base Editor (FusXTBE) for rapid mitochondrial DNA programming ofhuman cellsin vitroand zebrafish disease models in vivo
Mitochondria play an important but still largely mysterious role in human physiology, as demonstrated by the enormous biological variation and diverse disorders in patients with mitochondrial disease. Understanding how mitochondria function in normal biology and how human mitochondrial DNA variations contribute to health and disease has been hampered by few effective approaches to manipulate mtDNA and a lack of existing animal models. To circumvent RNA delivery issues to mitochondria, a new, CRISPR-free, TALE-derived base editor was developed that can induce C to T (or G to A) mtDNA sequence changes in human cells and in initial mouse model work. To enhance accessibility of this new editing tool and to enable rapid testing in new in vivo models, we developed this next generation FusX TALE Base Editor (FusXTBE). The FusX system is the first one-step TALE assembly system (Ma et al., 2016) that has been used to make an array of functional TALENs that work from flies to fish to human cells. Here we expand the FusX assembly system to use the initial mitochondrial base editor to facilitate broad-based access to TALE mitochondrial base editing technology. A de novo in silico design tool was also developed to assist in mapping potential base editing sites in human and zebrafish mitochondrial genomes and is also suitable for the analysis of any mtDNA genome. FusXTBE was demonstrated to function with comparable activity to the initial base editor in human cells in vitro. Zebrafish embryos were used as a pioneering in vivo test system, and we show injected animals with FusXTBE harbor edits in mtDNA loci with over 90% editing efficiency in F0 animals, the first example of majority mtDNA heterplasmy induction in any system. Gene editing specificity as precise as a single nucleotide was observed in vivo for a protein-coding gene. We have also adopted a new, non-destructive genotyping protocol for single animal mtDNA analyses, enabling rapid biological functional genomics applications downstream. FusXTBE is a new gene editing toolkit for exploring important unanswered questions in mitochondrial biology and genetics.

Biography:

Abstract:
Use of Entire Human Gut Microbiota to Cure Human Diseases without Drugs
Clostridium (now Clostridioides) difficile-associated disease (CDAD) is the major known cause of antibiotic-induced diarrhea and colitis, and the disease is thought to result from persistent disruption of commensal gut microbiota and dysbiosis. Bacteriotherapy, by way of intestinal microbiota transplantation (IMT), can be used to treat recurrent CDAD, which is thought to reestablish the normal colonic microflora. However, limitations of conventional microbiologic techniques have, until recently, precluded testing of this idea. We have used 16S rRNA gene sequencing, metagenomic, and metabolomic approaches to characterize the bacterial composition of the colonic microflora in patients suffering from recurrent CDAD before and after treatment by IMT from a healthy donor. Although the patient's residual colonic microbiota, prior to therapy was deficient in members of the bacterial divisions: Firmicutes and Bacteriodetes, IMT had a dramatic impact on the composition of the patient's gut microbiota. Change in gut bacterial composition of a recipient towards that of the host was accompanied by resolution of the patient's symptoms. The striking similarity of the recipient's and donor's intestinal microbiota following after bacteriotherapy suggests that the donor's bacteria quickly occupied their requisite niches resulting in restoration of both the structure and function of the microbial communities present. We sought to overcome many of the barriers in our clinical IMT program by using standardized frozen fecal material as well as freeze dried capsule material from a screened standard donor. Standardization of material preparation significantly simplified the practical aspects of IMT without loss of efficacy in clearing recurrent CDI – the overall success rate was 96% with frozen materials and 80% with encapsulated materials. More recent studies have shown that durable clinical success is dependent upon the speed of engraftment of donor microbiota and production of secondary bile acids, as well as the increased relative abundances of Lachnospiraceae, Bacteroidaceae, and Porphyromonadaceae. Current studies are investigating the use of IMT for other medical conditions and changes to formulations leading to increased efficacy.

Biography:
Dr. Sadowsky received his undergraduate degree in the Department of Bacteriology at the University of Wisconsin-Madison and received his Ph.D. in Microbiology at the University of Hawaii in 1983, focusing on microbial ecology and molecular biology. He has worked for private industry (Allied Corp) and the federal government (USDA-ARS) and joined the faculty at the University of Minnesota in 1989, where he is currently a Distinguished McKnight University Professor in two departments and a member of 10 graduate faculties. Since 2009 he has served as Director of the BioTechnology Institute (BTI), a collaboration between the College of Biological Sciences and the College of Science and Engineering, established to promote interdisciplinary research and outreach in biotechnology, microbial engineering, microbial processing, and fermentation development
Dr. Sadowsky has authored or coauthored more than 409 articles in scientific journals, was elected a fellow of the American Academy of Microbiology in 1999, and a fellow of the American Association for the Advancement of Science in 2008.
In addition to his teaching and research efforts, Dr. Sadowsky was Director of Graduate Studies for the Microbial Engineering Program and has been Director of Graduate Studies for the Microbial Ecology Minors Program for 30 years. He serves on several editorial boards is founding Editor-In-Chief of one of ASMs newest journals – Microbiology Spectrum. He also served on the ASM Branch Organizing Committee and currently serves on the Journals Committee.
Research efforts in his laboratory are directed towards understanding the microbial ecology of host-microbe interactions using new intestinal microbiota transplantation (IMT) technology, which his lab helped to develop and standardize. He has been using DNA sequencing, metagenomics, metabolomics, and computational methods to provide valuable insights and practical knowledge about how microorganisms in the human GI tract are related to human health at the biochemical level. While his lab has focused on C. difficile associated diarrheal disease, he is also involved in several other projects using FMT, including autism spectrum disorder, ulcerative colitis, diabetes, and metabolic syndrome. Dr. Sadowsky’s research efforts are very diverse. He also studies the structure and function of bacteria in soils, water, and in invasive aquatic species using metagenomic approaches.

Abstract:
The (R)evolution of Indigenous Food Systems of North America
Oglala Lakota Chef Sean Sherman, founder of the company The Sioux Chef, is decolonizing our food system. From growing up on Pine Ridge to an epiphany on a beach in Mexico, Chef Sean Sherman shares his journey of discovering, reviving, and reimagining Native cuisine. He also co-founded the nonprofit NATIFS (North American Traditional Indigenous Food Systems) with the mission to bring back Indigenous education and food access to the tribal communities.

Biography:
Chef Sean Sherman, Oglala Lakota, born in Pine Ridge, SD, has been cooking across the US and Mexico over the past 30 years, and has become renowned nationally and internationally in the culinary movement of indigenous foods. His main focus has been on the revitalization and evolution of indigenous foods systems throughout North America. Chef Sean has studied on his own extensively to determine the foundations of these food systems to gain a full understanding of bringing back a sense of Native American cuisine to today’s world. In 2014, he opened the business titled, The Sioux Chef as a caterer and food educator in the Minneapolis/Saint Paul area. He and his business partner Dana Thompson also designed and opened the Tatanka Truck, which featured 100% pre-contact foods of the Dakota and Minnesota territories.
In October 2017, Sean was able to perform the first decolonized dinner at the James Beard House in Manhattan along with his team. His first book, The Sioux Chef’s Indigenous Kitchen was awarded the James Beard medal for Best American Cookbook for 2018 and was chosen one of the top ten cookbooks of 2017 by the LA Times, San Francisco Chronicle as well as the Smithsonian Magazine. Also that year, Chef Sean was selected as a Bush Fellow, as well as receiving the 2019 Leadership Award by the James Beard Foundation. The Sioux Chef team continues with their mission to help educate and make indigenous foods more accessible to as many communities as possible through the recently founded nonprofit North American Traditional Indigenous Food Systems (NATIFS). Through this entity, Sean sees this vision as even more relevant in the time of COVID. Educating the world on localizing food systems is critical and we believe that we can leverage NATIFS to expedite this mission. Learn more at www.natifs.org.

Abstract:
Using genetic engineering to combat invasive species
Recently developed tools for Precise Genome Engineering have ushered in a number of novel proof-of-concept technologies for pest control. Termed ‘genetic biocontrol’, these approaches allow researchers to essentially convert the pest organism into a pesticide. Released genetically engineered biocontrol agents would spread deleterious genes into the pest population, leading to a local eradication. This talk will introduce a strategy for genetic biocontrol developed in the Smanski Lab and discuss its translation into applied organisms.

Biography:
Michael Smanski is an Associate Professor at the University of Minnesota in the Department of Biochemistry, Molecular Biology, and Biophysics and the BioTechnology Institute. Dr. Smanski received a B.S. degree in Biochemistry and Cell Biology from the University of California – San Diego and a Ph.D. in Microbiology from the University of Wisconsin, where he deciphered the genetic and biochemical basis for antibiotic production in Streptomyces. As an HHMI Fellow of the Damon Runyon Cancer Research Foundation, he worked with Chris Voigt at MIT to develop a pipeline for engineering complex genetic systems, including antibiotic biosynthetic pathways. His current lab is develop enabling technologies to engineer biology in ways that address challenges in medicine, agriculture, and the environment. He has been recognized for his research accomplishments with the Dale F. Frey Award for Breakthrough Scientists (2015), DARPA Young Faculty Award (2017), University of Minnesota McKnight Land Grant Professorship (2019), and DARPA Director’s Fellowship (2019).

Abstract:
Engineering soil microbes for Agriculture
Joyn Bio is an ag-biotech company engineering microbes to reduce the environmental impact of agriculture. Joyn’s engineered microbes serve as the next class of ag solutions with sustainability and efficacy at the forefront. During this talk, we’ll review how our ability to engineer soil microbes correlates with host selection, and how DNA writing in these microbes can be leveraged to create novel crop protection solutions for agriculture.

Biography:Brynne Stanton is the Head of Strategic Innovation and co-founder of Joyn Bio, an ag-biotech company engineering microbes to reduce the environmental impact of agriculture. Stanton oversees the development of the next class of ag solutions with sustainability and efficacy at the forefront. Previously, Stanton spent five years at Ginkgo Bioworks and propelled the company from bootstrapping with government grants to become the world’s premier synbio company.

Abstract:
High-throughput Off-Target Detection for Adoptive Cellular Therapy Research
The CRISPR/Cas9 system demonstrates unparalleled editing efficiency but suffers from concerns related to target site specificity. Even though high-fidelity Cas9 mutants have been developed with increased specificity, interrogation of off-target effects remains necessary for many applications. Empirical determination of off-target effects (OTEs) continues to be a compulsory exercise as in silico prediction tools generally suffer from low specificity or low sensitivity. Here we present a modified high-throughput version of a previously reported, unbiased OTE detection methodology to empirically interrogate OTEs in a cellular environment. This assay requires relatively low amounts of genomic DNA, and the initial library prep is based on enzymatic rather than mechanical shearing of genomic DNA without loss of sensitivity. Furthermore, we can detect OTEs with greater sensitivity using chemically stabilized gRNAs and can more accurately detect OTEs through optimized alignment strategies in the data analysis. This optimized nomination strategy is employed to explore the off-target profile of guideRNAs against targets commonly used in researching adoptive cellular therapies. For this, we selected >120 guideRNAs and determined the off-target profile using Jurkat cells stably expressing Cas9 as a model system. This Cas9 off-target nomination strategy is followed by multiplexed target enrichment by amplification and next-generation sequencing including data analysis. The rhAmpSeq for CRISPR system allows for rapid quantification and analytically sensitive detection of editing events to confirm bona fide OTEs.

Biography: Rolf Turk is a Senior Staff Scientist in the Molecular Genetics research group at Integrated DNA Technologies. Rolf obtained a master’s degree in science at the University of Amsterdam. He continued his training as a graduate student at the Center for Human and Clinical Genetics at the Leiden University Medical Center. His research revolved around identifying the molecular mechanisms behind different forms of muscular dystrophy. After obtaining his Ph.D., he continued his research in this field as a post-doctoral researcher at the University of Iowa. Since his employment at IDT in the fall of 2015, Rolf has contributed to the development of RNA-guided endonucleases and optimization of delivery strategies. Additionally, he focuses on off-target nomination and confirmation methodologies.

Abstract: N/A

Biography: Mark Walton was appointed Chief Technology Officer in August 2019. Prior to taking on the CTO role, he served as Vice-President for R&D and Regulatory Affairs, and the Global Director of Regulatory Affairs. Before joining AquaBounty, Dr. Walton held positions as President of ViaGen, a leading animal cloning company, Chief Business Officer for Recombinetics, the company that has developed Polled Holstein cattle using TALEN gene editing technology, and Executive Vice-President for Research and Technology at RiceTec, a global hybrid rice seed company. He began his career in agriculture biotechnology in the 1980s and in 1990 started Linkage Genetics, the first DNA testing company to provide molecular marker services to plant and animal breeders. Mark is deeply involved in the on-going discussions between industry and governments on the regulation of genetically engineered livestock and played an active role in obtaining the Cloning Risk Assessment and the Guidance on Regulation of Genetically Engineered Animals from the US Food and Drug Administration. He earned a Ph.D. in Agronomy from the University of Nebraska – Lincoln.

Abstract:
Clinical Application of CRISPR Edited Tumor Infiltrating Lymphocytes in Gastrointestinal Cancer
Neoantigen-specific tumor infiltrating lymphocytes (TIL) have shown promise clinically but fail to consistently elicit durable tumor regression. The intracellular checkpoint CISH negatively regulates T cell receptor (TCR) signaling and adoptive transfer of Cish-/- T cells achieves pronounced and durable tumor regression in vivo in mice. We identified that CISH is preferentially expressed in human TIL compared to T cells from peripheral blood, and at the single-cell transcriptome level CISH expression is inversely correlated with markers of T cell activation; suggesting that CISH is a key regulator of TIL response to neoantigens. To further characterize CISH function in human T cells and determine whether TIL neoantigen reactivity could be enhanced by CISH inhibition, we developed a CRISPR/Cas9-based strategy to knockout (KO) CISH in human T cells with high-efficiency and without detectable off-target editing as measured by amplicon sequencing of computationally predicted off-target loci and unbiased GUIDE-Seq in human T cells. In peripheral blood T cells, CISH KO enhanced proliferation, cytokine polyfunctionality, and cytotoxicity in vitro. To determine if CISH KO similarly enhanced TIL function, we developed a clinical-scale, cGMP-compliant manufacturing process for CISH disruption in primary human TIL. In cGMP process validation runs we achieved highly efficient CISH KO at the genomic and protein levels (>90%) without detectable off-target editing while maintaining high viability and expansion (>1000-fold). Compared to WT controls, CISH KO TIL exhibited increased cytokine-dependent proliferation and enhanced TCR avidity and neoantigen recognition. Strikingly, we identified examples where neoantigen reactivity was lost during rapid expansion (REP) in control cultures but retained in the CISH KO TIL product. Similarly, we found that CISH KO was capable of unmasking reactivity against common TP53 mutations. Intriguingly, hyperactivation in CISH KO TIL did not increase differentiation, suggesting that CISH KO may uncouple activation and differentiation pathways. Based on these preclinical studies, we recently initiated a human clinical trial at the University of Minnesota evaluating CISH edited neoantigen-reactive TIL in patients with treatment refractory metastatic gastrointestinal cancer (NCT04426669). To date, three patients have received TIL infusion without evidence of acute adverse toxicity related to the cellular product. Clinical updates will be presented.

Biography:
Dr. Beau Webber is an Assistant Professor in the Department of Pediatrics, Division of Hematology and Oncology at the University of Minnesota. He graduated from the University of Wisconsin- LaCrosse in 2007 with a BS in Cellular and Molecular Biology and conducted his Ph.D. studies at the University of Minnesota where he studied the embryonic development of hematopoietic stem cells. As a postdoctoral fellow in the Hematology, Oncology, and Transplantation program at the University of Minnesota, Dr. Webber developed advanced strategies for genetic modification of human lymphohematopoietic and pluripotent stem cells for cancer immunotherapy and correction of inherited diseases. Dr. Webber joined the Department of Pediatrics Faculty as an Assistant Professor in 2017. Dr. Webber’s laboratory is focused on synergizing genome engineering, stem cell biology, and adoptive cellular therapy to develop novel treatments for genetic disease and cancer. Research projects in the lab currently fall into two broad areas: the application of genome engineering to develop improved cell-based immune and gene therapies, and the development of “bottom-up” cancer models using human pluripotent stem cells.

Abstract:
Engineering Health for Human's Best Friend
Current drug development paradigms require extensive study in lab animal models before a new entity is ever used in humans. Of course, most of these candidates are tested on lab beagles who are born and bred as brave souls that help advance human medicine. What if there was another way?
In this talk, we will discuss how LEAH Labs uses gene writing to bring novel therapies to our canine companions. We aim to flip the script towards utilizing spontaneous disease in companion animals to de-risk and accelerate human cell therapy development, all the while providing novel options to pets in need.
As a proof of concept, we are first focused on translating the breakthrough technology CAR-T cell therapy to dogs with B cell lymphoma, the clinical analog of human CAR-T curable non-Hodgkin's lymphoma. In the future, we aim to partner with academic and pharma institutions to trial next-gen human cell therapy strategies in spontaneous disease in dogs, giving pets a chance at life while gleaning actionable clinical data for human studies.
And, as a company born from the roots of the Genome Writers Guild, we also ponder the use of gene editing to cure monogeneic diseases in dogs that result from millennia of genetic drag during the creation and maintenance of the 195 AKC recognized dog breeds in the world today. May pet dogs be a suitable species for germline editing, and all the promise and apprehension that comes with it?
We believe that the rapid advances in gene writing, cell therapy, and gene therapy bring our furry best friends to the forefront of these important questions, and we are excited to build our vision within the ethics, and with the blessing from, the GWG community.

Biography:
Wesley A. Wierson PhD, founder and CEO of LEAH Labs, is a life sciences entrepreneur working to apply his expertise in gene editing technology to solve consumer level problems in society. He studied microbiology as an undergraduate, where he was first exposed to gene editing technology through undergraduate research. As a PhD student, he developed gene editing technology that permits the integration of exogenous genes into the genome with precision, thus giving cells new functions, allowing scientists to study biology and genetics and to engineer living therapies. Wes founded LEAH Labs in late 2018 to bring living therapies to dogs. LEAH Labs is first focused on translating the breakthrough, FDA approved cell therapy called "CAR-T cell therapy" to dogs with B cell lymphoma. Not only does this approach bring the potential to disrupt a decades old standard of care for our pets, LEAH Labs also aims to trial novel cell therapies in canine models of spontaneous disease to give dogs a chance at life while derisking and accelerating human drug development.

Abstract:
Cas9 Based Gene Therapy for Angelman Syndrome
Angelman syndrome (AS) is a severe neurodevelopmental disorder caused by mutation or deletion of the maternally-inherited UBE3A allele. In neurons, the paternally-inherited UBE3A allele (patUBE3A) is silenced in cis by a long non-coding RNA called UBE3A-ATS. As part of a systematic screen, we found that Cas9 can be used to unsilence paternal Ube3a in cultured mouse and human neurons when targeted to Snord115 genes, small nucleolar RNAs (snoRNAs) that are clustered in the 3´ region of Ube3a-ATS. A short Cas9 variant and guide (gRNA) targeting ~75 Snord115 genes was packaged into an adeno-associated virus (AAV) and administered to embryonic and early postnatal AS mice, when the therapeutic benefit of restoring Ube3a is predicted to be greatest. This early treatment unsilenced patUbe3a throughout the brain for at least seventeen months and rescued anatomical and behavioral phenotypes in AS mice. Genomic integration of the AAV vector into Cas9 target sites caused premature termination of Ube3a-ATS at the vector-derived polyA cassette, or when integrated in the reverse orientation, by transcriptional collision with the vector-derived Cas9 transcript. Our study shows that targeted genomic integration of a gene therapy vector can restore patUBE3A function throughout life, providing a path towards a disease-modifying treatment for a syndromic neurodevelopmental disorder.

Biography:
Dr. Wolter completed his PhD in the lab of Dr. Marco Mangone at Arizona State University, studying evolutionary trends and targeting principles in microRNAs. He is currently a Distinguished Post-Doctoral Fellow of Gene Therapy at the University of North Carolina Chapel Hill, where he is broadly interested in molecular deficiencies in neurodevelopmental disorders. In the lab Dr. Jason Stein, Dr. Wolter uses libraries of genetically diverse primary human neural progenitor lines to study how common genetic variation affects cellular traits associated with neuropsychiatric and neurodevelopmental disorders. Dr. Wolter also leads a diverse team of research studying underlying molecular changes in ARSACS, a pediatric onset neurodegenerative disorder. Lastly, in the lab of Dr. Mark Zylka, he is exploring how CRISPR/Cas9 can be used to treat Angelman syndrome, a severe neurodevelopmental disorder.

Abstract:
Stepping Forward: Regulatory Approaches that Encourage Innovation
We face unprecedented challenges in agriculture including responding to the mounting hunger and nutrition insecurity crisis from a global pandemic, while conserving and safeguarding finite and rapidly diminishing natural resources. USDA is committed to transforming America’s food system with a greater focus on more resilient local and regional food production, fairer markets for all producers, ensuring access to healthy and nutritious food in all communities, building new markets and streams of income for farmers and producers using climate smart food and agricultural practices. Genome editing, along with a broad range of other tools, provide an opportunity for scientists, breeders, farmers, and ranchers to adapt to and mitigate climate change, enable sustainable farmer and rancher income, and provide additional benefits for society, including healthier and more resilient crops and livestock, while reducing agriculture’s impact on the environment. Genome editing can only help us achieve these goals if global regulatory and policy approaches allow their use in agricultural breeding programs.
Farmers and ranchers know what works best for their farms. If empowered to do so, many will choose effective options, including disease-resistant crops and animals produced with genome editing. Farmers and ranchers need a full toolbox of existing and innovative options, including tools that reduce use of pesticides and antibiotics in agriculture. Genome editing, in concert with other existing technologies, can supercharge efforts in combatting agricultural pests, adapting to climate change, increasing productivity of farms, improving animal welfare, and reducing environmental impacts. However, farmers and ranchers will not be able to access these tools unless flexible approaches are in place that encourage innovation and allow safe products to be used on farms.
The livelihoods of people all along the agricultural value chain depend upon on our regulatory and policy choices. The global regulatory landscape for products of genome editing is rapidly evolving, with an increasing number of countries focusing more on characteristics of products and whether they could be achieved by conventional breeding, rather than the technologies used to create them. Regulatory approaches and how they are applied have a tremendous impact in determining what products are developed and who can afford to use these new biotechnologies. We need to step forward and continue the momentum towards regulatory approaches that encourage innovation.

Biography: Dr. Diane Wray-Cahen is the Senior Advisor for Animal Health and Production, and Animal Products for the U.S. Department of Agriculture (USDA), Office of the Chief Scientist, where she is on long-term detail from the USDA Foreign Agricultural Service (FAS). At FAS, she served as a senior science advisor for agricultural biotechnologies for over 10 years, focusing on scientific and regulatory developments and their potential impact on agricultural innovation and trade policy. She also coordinated efforts with other like-minded countries to encourage informed and rational responses to new agricultural biotechnologies and to promote science-based and risk-proportionate regulatory approaches. Dr. Wray-Cahen has worked extensively with international colleagues on educational outreach and communication efforts regarding innovative agricultural technologies and associated issues, including sustainable intensification. She leads USDA’s international outreach efforts for animal biotechnologies, collaborating with livestock and breeding associations, public and private sector researchers, and regulatory officials in other countries. The on-going series of international and regional workshops on regulatory approaches for animal biotechnology has engaged researchers, breeders, regulators, and policy makers in more than 65 countries since 2011.
Prior to moving into the realm of science and trade policy, Dr. Wray-Cahen spent over 16 years conducting swine and dairy cattle research in the United States and the United Kingdom. Her research career focused on both agricultural applications and development of large animal biomedical models. Her work encompassed a mix of basic and applied research, including dairy metabolism research at the British AFRC Institute of Grassland and Environmental Research and the University of Reading (UK) and swine nutrition and metabolism research at the USDA Agricultural Research Service’s Beltsville Agricultural Research Center. She developed baby pig models for neonatal nutrition at the USDA Children's Nutrition Research Center and spent 8 years working with large animal models for biomedical applications at the HHS Food and Drug Administration’s Center for Medical Devices and Radiological Health. She also served as a lead scientist in the US Pandemic Influenza and Emerging Diseases Program within Biomedical Advanced Research and Development Authority. Dr. Wray-Cahen earned her B.S. and Ph.D. in Animal Science from Cornell University.

Abstract:
Genome Modified Animal for Xenotransplantation

Biography: Luhan Yang is a scientific entrepreneur who pioneered genome editing technology and is passionate about changing the world for better. She is leading the efforts of creating human transplantable organs and cells using genome editing approach at eGenesis ( Cambridge, USA) and Qihan ( Hangzhou, China).

Abstract:
Modular assembly of AAV-antibody composites for receptor-mediated gene delivery
Modern gene therapy is limited by lack of specific delivery vectors, rather than lack of therapeutic targets. Viral vectors currently used in clinical trials, such as adeno-associated virus (AAV), infect a broad range of tissues naturally targeted by the wild-type virus. In the clinic, this can result in ectopic expression in undesirable cell types and/or low expression in target tissues. To remedy this, engineering infection to be antigen-specific has been a goal of the field for almost 30 years. Past engineering efforts have grafted a targeting moiety (i.e. scFv, DARPin) into the protein capsid. While this method has resulted in targeted infection, each new antigen requires a new engineered capsid. We have developed a method of creating antigen-specific AAVs using a single engineered capsid containing a DNA binding domain (HUH tag). These HUH-AAVs can then be attached to DNA-conjugated antibodies with high efficiency.We have shown that a single HUH-AAV can be retargeted to generically any antigen; each of the antibodies we have used thus far has worked as expected in immortalized and primary cells. I will present data showing targeted delivery of fluorescent markers to multiple subtypes of cells in vivo and ex vivo.

Biography: N/A