Biology subjects

Ferber, D.

Publications and source records attributed to Ferber, D..

5 recordsLinked to original sources

bioRxiv2026

Evolutionary algorithms accelerate de novo design of potent Nectin-4-specific cancer biologics

Recent advances in AI-based structural biology have made de novo protein binder design increasingly effective, yet performance remains highly target dependent. For instance, the cancer surface antigen Nectin-4, an immunoglobulin-like cell adhesion protein, proved particularly challenging for RFdiffusion-based minibinder generation, yielding substantially fewer high-quality candidates than related targets. To address this bottleneck, we integrated an evolutionary genetic algorithm (GA) with AI-driven design. GA selection with tunable stringency was coupled with diversification via partial diffusion or direct sequence editing, enabling efficient exploration of sequence-structure space and rapid enrichment of promising candidates. This AI-GA pipeline quickly produced large and diverse minibinder panels with very good in silico quality metrics and is compatible with inputs from multiple design algorithms. Pooled, large-scale experimental screening identified highly stable Nectin-4 minibinders with single-digit nanomolar down to subnanomolar affinities. Lead binders were further engineered into Nectin-4-specific flow cytometry detection reagents and potent bispecific T cell engagers, demonstrating functional activity beyond binding. Together, these results show that evolutionary refinement can unlock challenging targets and accelerate de novo protein design for next-generation cancer biologics.

bioengineering↗

bioRxiv2025

Inhibition of NLRP3 by a CNS-penetrating indazole scaffold

Low-grade inflammation is a hallmark of ageing and a key cause of age-related impairments and diseases1. The NOD-like receptor NLRP3 senses a variety of danger signals and environmental insults, resulting in pro-inflammatory response, inflammasome formation and pyroptosis2,3. Its aberrant activation has been linked to many acute and chronic diseases ranging from atherosclerosis to Alzheimers disease and cancer, making NLRP3 an attractive therapeutic target4,5. Here we report the discovery, characterization, and structure of an indazole-based NLRP3 antagonist, BAL-1516, which potently inhibits inflammasome formation in monocytes and microglia. The cryo-electron microscopy structure of BAL-1516 bound to NLRP3 reveals a previously undescribed compound binding site at a surface groove of the nucleotide-binding domain with contacts to the FISNA and WHD subdomains. The characteristic feature of BAL compound binding is the formation of three hydrogen bonds to the peripheral {beta}-strand of the triple-ATPase; two from the indazoles nitrogen atoms and a third from the compounds linker region. Additional phenyl and thiazole moieties render the compound hydrophobic, allowing excellent blood-brain barrier penetration. The compound binding site is highly specific for NOD-like receptors, and the optimized compound BAL-1516 is able to directly bind mouse NLRP3 despite two conservative residue changes in the binding interface. The BAL compounds represent a first-in-class family of NLRP3 inhibitors, providing a broad design space, including covalent and degradative properties, for the development of NLRP3-directed therapeutics. The innate immune system contains cytosolic proteins that sense cellular stress caused by bacterial, viral and fungal infections or sterile inflammation, to control cellular integrity2. NLRP3 is a well-studied member of the nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) that is involved in the activation of the inflammasome, a multiprotein complex that mediates inflammation6. Upon detection of stress or pathogen-associated signals, NLRP3 triggers the activation of caspase-1, which leads to the production of pro-inflammatory cytokines such as IL-1{beta} and IL-18, driving inflammatory responses and ultimately pyroptotic cell death. In the context of neuroinflammation, NLRP3 plays a significant role in the pathogenesis of various neurodegenerative diseases, including Alzheimers disease, Parkinsons disease, and multiple sclerosis7. Research into targeting NLRP3 signalling with CNS-penetrating molecules holds potential for developing therapeutic strategies to alleviate neuroinflammatory conditions and to slow the progression of neurodegenerative diseases.

immunology↗

bioRxiv2025

Development of AI-designed protein binders for detection and targeting of cancer cell surface proteins

Artificial intelligence (AI)-based protein design opens new avenues for the rapid generation of new research tools and therapeutics, but experimental validation lags behind the computational design throughput. Here, we present a scalable workflow for the discovery and validation of AI-designed protein binders. Leveraging the RFdiffusion protein design pipeline with a custom filter for stable alpha-helical bundle folds, we construct libraries of thousands of AI-binders against cancer-associated surface proteins. Mammalian cell-surface and phage display screening yield multiple high-affinity PD-L1 binders but fewer hits for CD276 (B7-H3) and VTCN1 (B7-H4), reflecting the target-dependent efficiency of RFdiffusion in generating high-quality designs. Using our experimentally validated AI-designed binder libraries, we benchmark freely available structure prediction models. We find that interface predicted template modelling (ipTM) scores by Chai-1 with ESM embedding correlate well with experimental success and even predict deleterious effects of binding interface mutations. To demonstrate the versatility of AI-binders as research tools, we deploy them in CAR-T cells and also assemble them with fluorophore-labeled streptavidin into tetravalent quattrobinders, which achieve antibody-comparable staining of endogenous PD-L1 by flow cytometry. With high production yields and accessible structural models, AI-designed quattrobinders are versatile and cost-effective research tools amenable to community-driven validation and optimization.

bioengineering↗

bioRxiv2024

Discovery of a Potent and Selective Inhibitor of Human NLRP3 with a Novel Binding Modality and Mechanism of Action

The NLRP3 inflammasome is an intracellular protein complex that causes inflammation via the release of IL-1{beta} and pyroptosis. NLRP3 activation is associated with many age-related inflammatory diseases, and NLRP3 inhibition is a promising therapeutic strategy. We previously performed a DNA encoded library screen to identify novel NLRP3 binding molecules. Herein we describe the characterization of BAL-0028 as a potent and specific inhibitor of NLRP3 signaling. Notably, BAL-0028 is a poor inhibitor of mouse NLRP3 but inhibits human and primate NLRP3 with nanomolar potency. Using cellular and biochemical analyses we demonstrate that BAL-0028 binds to the NLRP3 NACHT domain at a site that is distinct from the MCC950 binding pocket. Using humanized NLRP3 mice we show that a derivative of BAL-0028 inhibits NLRP3 activation in vivo in a peritonitis model. Finally, we demonstrate that BAL-0028 inhibits select hyperactive NLRP3 mutations associated with autoinflammatory diseases more potently than does MCC950. BAL-0028 thus represents a new modality for NLRP3 inhibition in inflammatory diseases. SUMMARYNLRP3 is a target for anti-inflammatory therapies and can be inhibited by the tool compound MCC950. We describe the characterization of a new small molecule inhibitor of NLRP3 BAL-0028 that has a distinct mechanism of action and binding site.

immunology↗

bioRxiv2021

Langerhans islets induce anti-tumor immunity at the expense of glycemic control and predict chemotherapy response in pancreatic cancer

Induction of anti-tumor immunity in pancreatic ductal adenocarcinoma (PDA) is an unresolved challenge. Systematic investigation of the microenvironment of primary pancreatic tumors revealed a role of endocrine Langerhans islets in the coordination of immune activation. We found that intratumoral {beta}-cells, regulated via STAT3, secrete C-C motif chemokine ligand 27 (CCL27) and thereby promote a TH1 phenotype in the microenvironment resulting in an enhanced T cell infiltration and prolonged patient survival. The local effect can be abrogated in a patient-based human explant model by inhibition of the CCL27 receptor CCR10. This defense mechanism is paralleled by an impaired metabolic function of Langerhans islets with reduced insulin levels resulting in a dysregulation of glycemic control in patients. Based on these findings, screening of PDA cases (n= 2264) led to the identification of type 2 diabetes mellitus (T2DM) and extractable glycated haemoglobin (HbA1c) levels as response markers for neoadjuvant chemotherapy with fluorouracil, leucovorin, irinotecan and oxaliplatin (FOLFIRINOX). Collectively these data provide insights into the interconnection of T2DM and PDA, and link declining glycemic control to therapeutic efficacy, which can be utilized as a tool for clinical decision-making and improve patient management.

cancer biology↗