Vivlion

🎉 Celebrating a decade of CRISPR screening: Back to 2020 🎉 Breakthroughs in 2020 paved the road for combinatorial #CRISPR #screening. Major breakthroughs came from making new nucleases applicable for pooled combinatorial screening: ▶ The team of John Doench focussed on an enhanced #Cas12a variant from Acidaminococcus (enAsCas12a), which was systematically refined with respect to its cutting and targeting capabilities. The team optimized the gRNA design to improve on-target activity and reduce off-target effects, thereby enhancing AsCas12a’s editing precision across diverse cellular contexts. They also identified alternative DR (direct repeat) sequences that are less prone to recombination than the wild-type sequence, improving their stability and applicability. This finding is essential for delivering multiple gRNAs to a given cell and ensuring the activity. The team managed to make AsCas12a compatible with combinatorial genetic screens and reported an intriguing use case: Synthetic lethality screening in two cancer cell lines. They identified a previously unknown interaction between MARCH5 and WSB2, proving that their approach can indeed elucidate novel cellular dependencies. This was one of the first times that #Cas12 has been shown to perform similarly good as best-in-class #Cas9 libraries, with the major advantage that due to the improved properties of the enzyme, the experimental effort could be reduced by 4-fold.  ▶ In another study, the teams of Jonathan Weissman and Britt Adamson introduced direct-capture #Perturb-seq – a novel sequencing approach in which multiple expressed sgRNA sequences are sequenced directly together with the transcriptome of a single cell. This enabled single-cell CRISPR sequencing with combinatorial CRISPR libraries. The team used their technology to explore interactions between cholesterol biogenesis and DNA repair. They also provided evidence that an individual gene can be more efficiently targeted when multiple sgRNAs are delivered to the cell – an approach which Vivlion has expanded to genomic scale with its #Alexandria fixed-pair library that is available for human and mouse. Combinatorial single-cell screening allows for phenotypic profiling of multiple gene targets in individual cells and thereby overcomes longstanding limitations in uncovering complex gene interactions, e.g., having the power of accelerating both target discovery and drug mode of action studies.

  🎉 Celebrating a decade of CRISPR screening: Back to 2019 🎉 After last highlighting significant technological advances, today we look at results of clinically relevant #CRISPR screens that were published in 2019.  ▶ Analyzing genome-wide silencing screens, the teams of Francisca Vazquez and Adam Bass unveiled the Werner (WRN) helicase as a potential therapeutic target for microsatellite instable (MSI) cancers. Microsatellite instability is a hallmark of e.g. colorectal, endometrial and gastric cancers and is caused by defective DNA mismatch repair. Respective tumors have high mutation rates and limited treatment options. The teams analyzed two large-scale cancer dependency datasets: One coming from project #Achilles (517 cell lines screened with CRISPR/Cas9) and the second from project #DRIVE (398 cell lines screened with an RNAi library). This revealed that inhibiting WRN helicase in MSI cancer cells leads to irreparable DNA damage followed by death, without affecting healthy cells. #DepMap   ▶ In parallel, the teams of Kosuke Yusa and Mathew Garnett also set out to leverage genome-wide CRISPR-Cas9 knockout screens to identify and prioritize potential therapeutic targets. They screened 324 human cell lines from 30 cancer types to reveal key vulnerabilities that can be exploited for cancer treatment. The teams developed an analytical framework to prioritize potential candidates based on their impact on cell fitness, on genomic biomarkers and the drug development potential of the target. Notably, the study identified several previously unrecognized gene dependencies that could serve as promising drug targets – amongst them also the WRN helicase as a synthetic lethal target in MSI tumors. #OpenTargets   💡 These findings showcase that cancer cells often rely on unique sets of genes for survival, suggesting that targeted therapies could be developed to selectively attack cancer cells while sparing normal cells. This precision medicine approach could lead to more effective and less toxic treatments for patients. 💊 Regarding WRN helicase, this dream could soon come true: The selectivity towards cancer cells made it a promising target for synthetic lethality – an exciting result which paved the way for the development of WRN helicase inhibitors that are applied in clinical setting. A first phase I study with such an inhibitor started 2023. Taken together, the 2019 studies identifying WRN helicase as synthetic lethal target represent a major step forward in the fight against hard-to-treat cancers and exemplify the power of large-scale screening data sets in identifying novel cancer vulnerabilities and therapeutic biomarkers.  

In a festive ceremony, Vivlion’s CFO Sönke Bästlein received today the “Honorary Citzenship” of Goethe-Universität Frankfurt University. Sönke is awarded for his significant contributions and his long-standing commitment as a member of University Council. He served on this governing body including the Economic and Financial Committee from 2008 to 2024. With about 45,000 students, 600 professors, 16 faculties, Goethe University is one the largest German universities. Sönke is also an advisory board member of the Goethe Unibator the start-up accelerator of the University, and a lecturer for ‘Equity Governance’ at the Faculty for Economics and Business. Besides him, two other longstanding members of the Council were honored, Gabriele Eick and Prof. Bernhard Zwißler. Congratulations to all awardees!

Our co-founder and advisor Ivan Đikić has been awarded the prestigious #LOEWE-Spitzenprofessur by the Hessian Ministry of Science (HMWK). The funding of around €3 M over five years will support his groundbreaking research program in developing novel therapeutics to degrade disease-relevant proteins. Ivan has dedicated his career to uncovering mechanisms that regulate cellular quality control, pioneering the concept of #ubiquitin as a versatile cellular signal. His recent work focuses on reprogramming the body’s natural degradation pathways — such as the ubiquitin and #autophagy systems — to target harmful proteins or organelles in conditions like #cancer, neurodegeneration and infections. This funding will significantly advance the development of proximity-induced drugs - #PROXIDRUGS – offering new hope for diseases that were previously considered untreatable. Congratulations to Ivan, feeling so proud for you 👏👏👏 #ResearchExcellence #CancerResearch #ProteinDegradation #DrugDevelopment #Innovation https://lnkd.in/ekJ6N29w

🎉 Celebrating a decade of CRISPR screening: Back to 2019 🎉 2019 saw several #CRISPR #screening breakthroughs, and today we’ll focus on two publications that presented major technological advances: ▶ The group of Paul Blainey developed a novel optical pooled screening approach: they conducted a pooled loss-of-function CRISPR screen in millions of cells, followed by image-based phenotyping – assessing protein localization, live-cell dynamics, cell morphology – and then used in situ sequencing to associate the individual genetic perturbation with the observed phenotypes. They aimed at identifying regulators of NF-kB signaling and tested 952 genes in their study. For their proof-of-concept, they chose RelA (p65) translocation as biological endpoint/phenotype and analyzed various fixed human cell lines, revealing known genetic associations. Moreover, they discovered a novel function of the Mediator complex in nuclear retention of p65, negatively impacting pro-inflammatory signaling. This work represents a major improvement of pooled CRISPR screening as it enables the readout of a vast portfolio of dynamic cellular responses, gaining insight into more complex phenotypes.  ▶ In a second study, a team led by Thomas Norman, Luke Gilbert and Jonathan Weissman developed an analytical framework to systematically study how genetic interactions (#GIs) shape diverse cellular phenotypes. Until then, mapping of GIs was mostly based on cell fitness effects, which provides little mechanistic insight into how complex gene networks regulate cell behavior. They introduced the concept of GI “manifolds”, in which all possible phenotypic states of a cell are represented. For this, they used high-throughput CRISPR-based perturbations combined with single-cell transcriptomic profiling, mapping GIs at an unprecedented resolution and data complexity. They focused on systematically perturbing pairs of genes to understand how their interactions influence cellular phenotypes, leveraging the rich phenotypic data derived from single-cell RNA sequencing. This approach captured complex genetic dependencies and co-regulatory mechanisms that cannot be observed through cell fitness GI studies. By employing dimensionality reduction and #manifold learning techniques, they could visualize and interpret GI networks at an unprecedented scale. This work advanced our understanding of cellular systems biology tremendously, demonstrating that GI landscapes can be described by single-cell phenotypes. The findings have significant implication, as they offered a new paradigm for exploring gene function and interactions at a systems level.  

🎉 Celebrating a decade of CRISPR screening: Back to 2018 🎉 Utilizing #CRISPRscreening, three 2018 publications  probe large-scale genetic and protein interactions, contributing to our understanding of cellular machines and complex cellular mechanisms. ▶ A team led by Michael Boutros reported the largest genetic network of cancer cells to date (Benedikt Rauscher et al, Mol Sys Biol 2018). Using a novel computational framework termed MINGLE, they integrated data from 85 screens to extract #syntheticlethal genetic interactions (GIs). These genetic interactions — where the loss of one gene exacerbates the loss of another — are crucial for uncovering disease vulnerabilities that could be therapeutically exploited. They explored more than 2 M gene-background relationships. The study significantly broadened our understanding of cancer-specific gene interactions and highlighted the complexity of genetic networks in cancer. Its findings serve as a resource for identifying potential drug targets and personalized cancer therapies. ▶ Yet another blueprint for mapping the human genetic landscape came from the teams of Jonathan Weissman and Luke Gilbert. They perturbed more than 220k gene pairs by combinatorial #CRISPRi which enabled GI mapping and identified hitherto unknown gene functions (Max Horlbeck et al, Cell 2018). Their work elucidated unexpected relations between pathways involved in cell growth, proliferation and differentiation, plus delivering a high-resolution view of gene function in human biology. ▶ Finally, a team around Cigall Kadoch focused on uncovering the structure and function of protein complexes using data derived from #CRISPR and #RNAi #screening in human cancer cells (Pan, Meyers et al, Cell Syst 2018). They devised functional similarity networks to provide critical insights how proteins function together. Besides reflecting known structures and functions, the networks also revealed new functional groups in complexes that were not yet structurally resolved. Again, the team utilized data from a large number of screens performed in hundreds of cancer cell lines. Then they integrated their functional networks with protein-protein-interaction information, leading to the discovery of novel protein complexes. This work established a new methodology to explore how proteins cooperate in multi-subunit complexes, paving the way for improved drug discovery and a deeper understanding of how molecular complexes function in cellular processes. 🚀 The collective impact of these publications lies in their contribution to creating high-resolution maps of GIs and cellular dependencies, offering powerful tools for basic research and therapeutic applications. They accelerated the use of CRISPR screens as a standard technique for dissecting complex genetic networks, identifying novel drug targets and developing #personalized #therapies, particularly in cancer.

🎉 Celebrating a decade of CRISPR screening: Back to 2018 🎉 Today, we highlight a review and a breakthrough highly relevant to advancing #crispr #screening capabilities. ▶ First the review – if you haven’t done so yet, the piece by John Doench in Nature Reviews Genetics titled “Am I ready for CRISPR? A user's guide to genetic screens” is a must read. It provides a step-by-step roadmap for scientists considering to use the power of #CRISPRscreening in their work, with practical considerations on design, execution and interpretation of CRISPR screening experiments, be it for understanding gene function, drug resistance or complex biological pathways. The review advises CRISPR novices, it demystifies the technical and conceptual aspects and offers critical advice on how to successfully apply #CRISPR #screening to a diverse set of research questions. ▶ Next, a 2018 technological breakthrough: In an editorial letter to Nature Methods, the teams of Luca Pinello and Daniel Bauer introduced CRISPR-SURF, a computational framework for deconvoluting data from CRISPR/Cas9 tiling screens that significantly advanced both functional genomics and regulatory element discovery (Jonathan Hsu et al, Nature Methods 2018). CRISPR-SURF is an abbreviation for Screening of Uncharacterized Region Function – referring to tiling screens that are designed to map and identify functional regulatory elements, e.g. enhancers and promoters, by creating systematic point mutations across large genomic regions. The framework provides a robust statistical framework for analyzing dense CRISPR tiling experiments across large genomic landscapes. It improves the signal to noise ratio by accounting for factors like target sequence properties and screen efficiency, increasing the confidence when discovering regulatory elements, thereby expanding the #searchspace of CRISPR screens to the #noncoding genome. Before this, most CRISPR screens were largely focused on protein-coding genes. CRISPR-SURF became a critical resource for research into complex biological systems and diseases. It paved the way for deeper biological insights into gene regulation, epigenetics and the underlying genetic mechanisms of disease.   Watch out for our next post, where we stay in 2018 and highlight gene interaction mapping to understand complex biology! #analysisiskey

A warm welcome to Simran Rastogi, who started as Associate Scientist at #Vivlion in beginning of October! Simran completed her master's training in molecular biosciences at Universität Heidelberg and DKFZ Deutsches Krebsforschungszentrum. She is specialized in molecular and cell biology and has extensive hands-on experience in different applications of #CRISPR-mediated genome editing. Through several longer internships at Broad Institute of MIT and Harvard, BioMed X Institute and National Center for Tumor Diseases (NCT) Heidelberg, she acquired a wide variety of technologies. Happy to have you on the team, Simran! #CRISPR #PRCISR #Screening #Startup

🎉 Celebrating a decade of CRISPR screening: Back to 2017 (again) 🎉 After highlighting two methodological innovations (CERES & CRISPR-UMI) yesterday, we will now dive into the ever-growing variety of biological applications of CRISPR screening. ▶ Screening for active lncRNA loci: In 2017, a team led by Feng Z. and Eric Lander shed light into the functions of long noncoding RNA (#lncRNA) loci by utilizing a CRISPR activation (#CRISPRa) screen (Joung et al, Nature 2017). Their screen was designed to target more than 10k transcriptional start sites for lncRNAs and identified 11 lncRNA loci to confer resistance to BRAF inhibitors in melanoma cells. Digging deeper into one of these loci, they were able to show how transcriptional activation of this lncRNA locus exerted a dose-dependent effect of four neighboring protein-coding genes, one of which was responsible for the resistance phenotype. This publication highlights how #CRISPRscreening enables the exploration of noncoding regions of the genome with direct implications for human diseases.    ▶ Understanding cancer immunotherapy response: A team around Nicholas Restifo, Shashank Patel and Neville Sanjana explored genetic factors that influence the efficacy of T cell immunotherapies in human cancer, particularly why some tumors resist treatment (Patel et al, Nature 2017). They utilized a genome-wide CRISPR library to mimic loss-of-function mutations occurring in human melanoma, and identified over 100 genes that are essential for the effector function of CD8+ T cells. Amongst them were genes with prominent roles in antigen presentation and interferon-γ signaling. One significant and unexpected discovery was the identification of the apelin receptor (APLNR) as a gene mutated in therapy-refractory patient tumors, and the authors also deciphered the underlying molecular mechanism. Taken together, the study revealed how loss of certain genes, including APLNR, could impair the efficacy of immunotherapies. As such, it is a prominent example for how #CRISPR #screening significantly accelerates research into resistance mechanisms by mimicking diverse genetic backgrounds of tumors in a pooled experiment. The results of this and many similar studies since have guided preclinical efforts to enhance the effectiveness of cancer therapy.

🎉 Celebrating a decade of CRISPR screening: Back to 2017 🎉 From 2017 onwards, CRISPR screening saw multiple technological improvements and an ever-growing variety of applications. Hence, we have chosen to highlight a couple of major publications from 2017 – starting today with two major technological advancements. ▶ Breakthrough in mapping vulnerabilities: The teams of William Hahn and Aviad Tsherniak developed the computational tool CERES and thereby solved a critical issue related to the analysis of genetic dependencies (Meyers et al, Nature Genetics 2017). The tool helps to account for the effects of gene copy number variations (CNVs), which can confound results in CRISPR screens, by correcting for the background gene editing effects caused by these variations. By applying CERES to a dataset of genome-scale CRISPR-Cas9 essentiality screens across 342 cancer cell lines (originating from The Cancer Dependency Map Project), the authors were able to significantly decrease false positive results. This publication marked a milestone in the ability to map critical cancer vulnerabilities, offering a refined approach to identifying genes crucial for cancer proliferation and survival. ▶ Single-cell tracing in pooled CRISPR screens: A team led by Ulrich Elling improved the predictive power of pooled screens by devising the CRISPR-UMI methodology (Michlits et al, Nature Methods 2017). They utilized sgRNA libraries bearing unique molecular identifiers (UMIs) to enable cell lineage tracing. The authors proved the robustness of the new methodology in both negative and positive selection screens and were able to reliably measure genetic perturbations at a single-cell level. CRISPR-UMI thereby overcomes limitations of traditional pooled #CRISPR screens, where results are usually aggregated across populations of cells and confounding effects due to off-target edits can occur. Whilst Perturb-Seq – which was one of our 2016 highlights – focusses on the transcriptomic consequences of individual edits, CRISPR-UMI provides insights into how genetic #perturbations influence the fate of individual cells within a population over time. It can trace the cell’s evolutionary history and its response to genetic changes.   📣 Stay tuned until tomorrow, when we will highlight two novel biological applications of #crisprscreening!