Genetic Technologies and New Gene Design for Curative Therapies @Stanford - @parkerici - @arcinstitute
Stanford, CA
Based in United States
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Roth_Lab retweeted
In the latest issue! Synthetic transcription factors designed by domain recombination enhance CAR T cell antitumor function dlvr.it/TVdDXq
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This has been a huge effort from Oliver, everyone in the Roth Lab and the Satpathy Lab, and an amazing group of collaborators. None of this type of work is possible without high risk-high reward funding, especially from the @arcinstitute, the @NIH Director ‘s New Innovator Award, and @PICI, @Burroughs Wellcome, and the @Weill Foundation
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Much more functional and mechanistic details about individual top DESynR AP-1 TFs in the full manuscript! But it is exciting to propose that the human genomes natural set of 20,000 genes may not be the only “human” genes that can be productively manipulated when studying human biology, building cellular therapies, or predictively designing optimal genetic perturbations for desired tasks. (9/10)
nitter.net/Satpathology/status/20…
Human protein-coding genes evolved from old parts - reconfigured domains from ancestral genes. We can screen short-sequence evolution at scale using mutagenesis, CRISPR, etc, but we had no way to do the same for domain-level evolution.
Happy to share our work developing DESynR genes today in @CellCellPress: scalable synthesis and functional screening of thousands of new human genes, built from old parts. DESynR transcription factors outperform native factors and dramatically improve CAR T cell function.
A team effort led by the incredible Oliver Takacsi-Nagy! @parkerici @CancerResearch @TheMarkFdn
cell.com/cell/fulltext/S0092…
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Importantly, it was the unique combination of existing natural domains into a new combined DESynR human AP-1 TF, rather than overexpression of each domain individually or each contributing natural AP-1 TF individually, that programmed the full extent of improved functional behavior. (8/10)
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We focused on the AP-1 family of basic lucine zipper TFs - they are both some of the most highly expressed in the genome (think Jun, Fos), as well as enriched in previous knockout and overexpression hits from prior natural gene screening in human T cell models. There are 24 natural human AP-1 TFs. We built over 13,000 new “DESynR” AP-1 TFs that the human genome could have constructed from existing AP1 TF domains. (3/10)
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Eukaryotic genomes most commonly acquire new genes through duplication of existing genes followed by recombination. We set out to experimentally mimic and accelerate this process, building, integrating into a primary human T cell’s genome, and functionally testing over 10,000 new human transcription factors in a single experiment. (2/10)
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Transcription Factors are central to cellular identity and function. The human genome currently contains ~1600 of them. But could there be productive human transcription factors that have not evolved? Out today in Cell, a tremendously talented graduate student Oliver Takacsi-Nagy @Oli_TN, together with the @Satpathology Lab, determined to find out. (1/10)
authors.elsevier.com/sd/arti…
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This has been a long running collaborative effort, run by exceptional future PhD students Courtney Kernick and Lauren Chow, and an entire team between Brian Shy’s and Greg Allen’s Labs at @UCSF and @GladstoneInst and our group @Stanford and @arcinstitute (10/11)
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More broadly beyond ex vivo electroporation based T cell and iPSC editing, we characterized head-to-head toxicity, delivery, editing efficiency, and manufacturability properties for a comprehensive series of linear and circular, single stranded and double stranded DNA templates, hopefully of use as a reference across applications requiring intracellular DNA delivery. (9/11)
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Building from this observation, we initiated a far ranging series of technical optimizations to enable much larger DNA integrations. We now present GLIDE-editing, a genetic engineering method using small DNA “helper” plasmids, mRNA-encoded nucleases, and sequence design optimizations among other improvements which allows for efficient ultra-large DNA integrations in primary human T cells. (5/11)
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In a close long term collaboration with the Shy and Allen labs at UCSF, we set out to comprehensively determine the ideal DNA template format for enabling much larger DNA integrations. Linear and circular ssDNA both worked well for smaller ~1.5 kb integrations, and linear ssDNA seemed most promising. (3/11)
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