Biology subjects

Sigoillot, F.

Publications and source records attributed to Sigoillot, F..

6 recordsLinked to original sources

bioRxiv2026

Scalable platform for cellular and biochemical screening of combinatorial small-molecule libraries in droplets

A new pattern of hit discovery is emerging with the rise of chemocentric modalities, such as chemical induced proximity (CIP). Synthesis and functional screening of dynamic, purpose-built libraries, such as E3 ligase biased libraries for targeted protein degradation (TPD), is redefining the druggable landscape. However, this new paradigm arguably lies beyond the remit of conventional screening methods which have been optimized for static, diversity-oriented libraries. To address this gap, we developed microfluidic compound screening in droplets (MicDrop), a scalable platform where purpose-built DNA-encoded one-bead one-compound library members are individually tested at high concentrations in pico-liter sized droplets for either biochemical or cellular function. A proof-of-concept CRBN library demonstrated the robustness of the workflow through an IKZF3 degradation screen which discovered an unexpected succinimide-based IKZF3 degrader, while reproducing the relative potency ranking of reference compounds. Screening of a prospective VHL library for CDO1 recruitment and degradation demonstrated that orthogonal assays gave an informative consensus hit list and an actionable machine-learning (ML) recruitment model. These findings establish a blueprint for intentional discovery of chemical inducers of proximity, while producing reliable data to accelerate ML-based design-make-test-analyze (DMTA) cycles in pursuit of new therapeutics. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=106 SRC="FIGDIR/small/730915v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@12de3cborg.highwire.dtl.DTLVardef@1c6618dorg.highwire.dtl.DTLVardef@12ea42eorg.highwire.dtl.DTLVardef@11cea77_HPS_FORMAT_FIGEXP M_FIG C_FIG

Cell Biology↗

bioRxiv2023

Reinstating targeted protein degradation with DCAF1 PROTACs in CRBN PROTAC resistant settings

Targeted protein degradation (TPD) of neo-substrates with proteolysis targeting chimeras (PROTACs) or molecular glues has emerged as a key modality in exploring new biology as well as designing new drug candidates where catalytic inhibition is neither efficacious nor an option. TPD is mediated through harnessing E3 ligases and redirecting them to ubiquitinate de novo target proteins for subsequent proteasomal degradation. Until recently, E3 ligase chemical matter available for mediating TPD has been limited to a relatively low number of ligases, considering that over 600 E3 ligases are encoded by the human genome. In addition, the most utilized ligase for TPD approaches, CRBN, has been observed to be downregulated in settings of acquired resistance to immunomodulatory inhibitory drugs (IMiDs). IMiDs are molecular glues that target IKZF transcription factors to CRBN for degradation. Resistance is potentially accelerated by non-essentiality of CRBN for cell viability. Here we investigated if the essential E3 ligase receptor DCAF1 can be harnessed for TPD utilizing a potent, non-covalent DCAF1 binder. We show that this binder, selective for the CRL4DCAF1 E3 ligase complex, can be functionalized into an efficient DCAF1-BRD9 PROTAC. Chemical and genetic rescue experiments confirm specific degradation via the CRL4DCAF1 E3 ligase. We further highlight the versatility of DCAF1 for TPD by developing a DCAF1-dasatininb PROTAC targeting multiple cytosolic and membrane bound tyrosine kinases. We expand these findings towards Brutons tyrosine kinase (BTK) selective PROTACs and through extensive optimization and characterization efforts share key observations that led to a potent and selective DCAF1-BTK PROTAC (DBt-10). Finally, with this PROTAC DBt-10, we show rescue of BTK degradation in a BTK-dependent, CRBN-degradation-resistant cell line and provide a rationale for E3 ligase swap to overcome CRBN mediated resistance.

biochemistry↗

bioRxiv2023

Drug-induced eRF1 degradation promotes readthrough and reveals a new branch of ribosome quality control

Suppression of premature termination codons (PTC) by translational readthrough is a promising strategy to treat a wide variety of severe genetic diseases caused by nonsense mutations. Here, we present two novel and potent readthrough promoters - NVS1.1 and NVS2.1 - that restore substantial levels of functional full-length CFTR and IDUA proteins in disease models for cystic fibrosis and Hurler syndrome, respectively. In contrast to other readthrough promoters that affect stop codon decoding, the NVS compounds stimulate PTC suppression by triggering rapid proteasomal degradation of the translation termination factor eRF1. Our results show that this occurs by trapping eRF1 in the terminating ribosome, causing ribosome stalls and subsequent ribosome collisions, activating a novel branch of the ribosome-associated quality control (RQC) network that involves the translational stress sensor GCN1 and the catalytic activity of the E3 ubiquitin ligases RNF14 and RNF25.

molecular biology↗

bioRxiv2021

Multi-Species Phenotypic Screening across Disease Models of Mucolipidosis Type IV

Invertebrate model organisms (the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster) are valuable tools to bridge the gap between traditional in vitro discovery and preclinical animal models. Invertebrate model organisms are poised to serve as better disease models than 2D cellular monocultures for drug discovery, as well as easier and more cost-effective to scale up than 3D organoids/assembloids or co-cultures. A strength of model organisms is the opportunity to probe conserved biology such as lysosomal function and autophagy in a physiological setting. However, invertebrate models are not without pharmacokinetic and pharmacodynamic challenges, such as poor tissue penetration and confidence in a compounds mechanism of action. To confront those challenges, we took advantage of the Novartis mechanism-of-action box (MoA Box), a compound library of well-annotated and drug-like chemical probes. Curious as to how the MoA Box, comprised of chemical probes optimized for mammalian targets, would fare in an invertebrate setting we screened the MoA Box across three different models of the lysosomal storage disease mucolipidosis Type IV (MLIV). MLIV is caused by mutations in the lysosomal transient receptor potential ion channel mucolipin-1 (TRPML1) resulting in hyper-acidic lysosomes and dysregulated autophagy. The overlap of screening hits from worm, fly, and patient fibroblast screens identified cyclin-dependent kinase (CDK) inhibition as an evolutionarily conserved disease modifier and potential drug repurposing strategy. Summary statementA trio of phenotypic screens across Drosophila, C. elegans, and H. sapiens models of mucolipidosis IV was performed and identified overlapping hits including cyclin-dependent kinase inhibitors.

cell biology↗

bioRxiv2019

In-Depth Characterization and Validation in BRG1-Mutant Lung Cancers Define Novel Catalytic Inhibitors of SWI/SNF Chromatin Remodeling

Members of the ATP-dependent SWI/SNF chromatin remodeling complexes are among the most frequently mutated genes in cancer, suggesting their dysregulation plays a critical role. The synthetic lethality between SWI/SNF catalytic subunits BRM/SMARCA2 and BRG1/SMARCA4 has instigated great interest in targeting BRM. Here we have performed a critical and in-depth investigation of novel dual inhibitors (BRM011 and BRM014) of BRM and BRG1 in order to validate their utility as chemical probes of SWI/SNF catalytic function, while obtaining insights into the therapeutic potential of SWI/SNF inhibition. In corroboration of on-target activity, we discovered compound resistant mutations through pooled screening of BRM variants in BRG1-mut cancer cells. Strikingly, genome-wide transcriptional and chromatin profiling (ATAC-Seq) provided further evidence of pharmacological perturbation of SWI/SNF chromatin remodeling as BRM011 treatment induced specific changes in chromatin accessibility and gene expression similar to genetic depletion of BRM. Finally, these compounds have the capacity to inhibit the growth of tumor-xenografts, yielding important insights into the feasibility of developing BRM/BRG1 ATPase inhibitors for the treatment of BRG1-mut lung cancers. Overall, our studies not only establish the feasibility of inhibiting SWI/SNF catalytic function, providing a framework for SWI/SNF therapeutic targeting, but have also yielded successful elucidation of small-molecule inhibitors that will be of importance in probing SWI/SNF function in various disease contexts.

cancer biology↗

bioRxiv2019

A genome-wide CRISPR screen identifies ZCCHC14 as a host factor required for hepatitis B surface antigen production

GRAPHICAL ABSTRACT\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC=\"FIGDIR/small/718940v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (45K):\norg.highwire.dtl.DTLVardef@1782e32org.highwire.dtl.DTLVardef@120f111org.highwire.dtl.DTLVardef@1b2ad50org.highwire.dtl.DTLVardef@1907bc1_HPS_FORMAT_FIGEXP M_FIG C_FIG SUMMARYA hallmark of chronic hepatitis B virus (CHB) infection is the presence of high circulating levels of non-infectious small lipid HBV surface antigen (HBsAg) vesicles. Although rare, sustained HBsAg loss is the idealized endpoint of any CHB therapy. A novel small molecule RG7834 has been previously reported to inhibit HBsAg expression by targeting terminal nucleotidyltransferase protein 4A and 4B (TENT4A and TENT4B). In this study, we describe a genome-wide CRISPR screen to identify other potential novel host factors required for HBsAg expression and to gain further insights into the mechanism of RG7834. We report more than 60 genes involved in regulating HBsAg and identified novel factors involved in RG7834 activity, including a zinc finger CCHC-type containing 14 (ZCCHC14) protein. We show that ZCCHC14, together with TENT4A/B, stabilizes HBsAg expression through HBV RNA tailing, providing a potential new therapeutic target to achieve functional cure in CHB patients.

microbiology↗