Jenkins, T. P. | Search bioRxiv

bioRxiv2026

De novo designed single-domain antibodies protect against lethal cobra venom neurotoxicity in vivo

Generative protein design can now rapidly produce de novo binders with high affinity and functional activity against a wide range of targets, including lethal snake venom toxins. However, so far most reported successes rely on new-to-nature scaffolds with limited therapeutic precedent. Single-domain antibodies (VHHs) offer a clinically validated alternative scaffold that can bind and neutralize long-chain -neurotoxins, which are some of the most lethal components in snake venoms. Here we compare three recently established de novo design models with VHH-design capabilities (Germinal, RFantibody, and BoltzGen) for their ability to generate VHHs against the neurotoxin -cobratoxin from the monocled cobra (Naja kaouthia). Using standardized model inputs and evaluation criteria based on AlphaFold3 interface confidence (ipTM) and RMSD self-consistency, we find that Germinal was the only method to generate designs passing stringent in silico criteria for experimental testing. We therefore performed a larger Germinal design campaign employing three different VHH frameworks and experimentally validated 46 designs in vitro. Of these, 42 expressed as soluble proteins and we identified four binding hits derived from two of the three tested frameworks. Of the four binders, two lead candidates were further characterized and demonstrated high affinity (KDs of 4.1 nM and 10.8 nM), monomeric behavior and low polyreactivity, indicating favorable biophysical and developability properties, as well as functional toxin neutralization in vitro. To assess their therapeutic potential we investigated their ability to protect against -cobratoxin toxicity in vivo. Both candidates fully protected mice after -cobratoxin challenge, with 100% survival compared to a lethal control. One candidate also retained notable neutralization capacity against whole venom of Naja kaouthia with a survival of 56%, while the other protected 22% when tested in a rescue setting. Together, we demonstrate that de novo VHH design can generate high affinity single-domain antibodies with in vivo protection against lethal cobra venom neurotoxicity, and provide practical insights into method- and framework-dependent performance.

bioengineering↗

bioRxiv2018

Exclusive dependence of IL-10Rα signalling on intestinal microbiota homeostasis and control of whipworm infection

The whipworm Trichuris trichiura is a soil-transmitted helminth that dwells in the epithelium of the caecum and proximal colon of their hosts causing the human disease, trichuriasis. Trichuriasis is characterized by colitis attributed to the inflammatory response elicited by the parasite while tunnelling through intestinal epithelial cells (IECs).\n\nThe IL-10 family of receptors, comprising combinations of subunits IL-10R, IL-10R{beta}, IL-22R and IL-28R, modulates intestinal inflammatory responses. Here we carefully dissected the role of these subunits in the resistance of mice to infection with T. muris, a mouse model of the human whipworm T. trichiura. Our findings demonstrate that whilst IL-22R and IL-28R are dispensable in the host response to whipworms, IL-10 signalling through IL-10R and IL-10R{beta} is essential to control caecal pathology, worm expulsion and survival during T. muris infections. We show that deficiency of IL-10, IL-10R and IL-10R{beta} results in dysbiosis of the caecal microbiota characterised by expanded populations of opportunistic bacteria of the families Enterococcaceae and Enterobacteriaceae. Moreover, breakdown of the epithelial barrier after whipworm infection in IL-10, IL-10R and IL-10R{beta}-deficient mice, allows the translocation of these opportunistic pathogens or their excretory products to the liver causing organ failure and lethal disease. Importantly, bone marrow chimera experiments indicate that signalling through IL-10R and IL-10R{beta} in haematopoietic cells, but not IECs, is crucial to control worm expulsion and immunopathology. These findings are supported by worm expulsion upon infection of conditional mutant mice for the IL-10R on IECs. Our findings emphasize the pivotal role of systemic IL-10R signalling on immune cells in promoting microbiota homeostasis and maintaining the intestinal epithelial barrier, thus preventing immunopathology during whipworms infections.\n\nAuthor summaryThe human gut is home to millions of bacteria, collectively called the microbiota, and also to parasites that include whipworms. The interactions between gut cells, the microbiota and whipworms define conditions for balanced parasitism. Cells lining the gut host whipworms but also interact with gut immune cells to deploy measures that control or expel whipworms whilst maintaining a barrier to prevent microbial translocation. Whipworms affect the composition of the microbiota, which in turn impacts the condition of the gut lining and the way in which immune cells are activated. In order to avoid tissue damage and disease, these interactions are tightly regulated. Here we show that signalling through a member of the IL-10 receptor family, IL-10R, in gut immune cells is critical for regulating of these interactions. Lack of this receptor on gut immune cells results in persistence of whipworms in the gut accompanied by an uncontrolled inflammation that destroys the gut lining. This tissue damage is accompanied by the overgrowth of members of the microbiota that act as opportunistic pathogens. Furthermore, the destruction of the gut barrier allows these bacteria to reach the liver where they cause organ failure and fatal disease.

immunology↗