Computational detection of antigen-specific B cell receptors following immunization.

B cell receptors (BCRs) play a crucial role in recognizing and fighting foreign antigens. High-throughput sequencing enables in-depth sampling of the BCRs repertoire after immunization. However, only a minor fraction of BCRs actively participate in any given infection. To what extent can we accurately identify antigen-specific sequences directly from BCRs repertoires? We present a computational method grounded on sequence similarity, aimed at identifying statistically significant responsive BCRs. This method leverages well-known characteristics of affinity maturation and expected diversity. We validate its effectiveness using longitudinally sampled human immune repertoire data following influenza vaccination and SARS-CoV-2 infections. We show that different lineages converge to the same responding Complementarity Determining Region 3, demonstrating convergent selection within an individual. The outcomes of this method hold promise for application in vaccine development, personalized medicine, and antibody-derived therapeutics.

Discovery of SOCS7 as a versatile E3 ligase for protein-based degraders.

Targeted protein degradation (TPD) strategy harnesses the ubiquitin-proteasome system (UPS) to degrade a protein of interest (POI) by bringing it into proximity with an E3 ubiquitin ligase. However, the limited availability of functional E3 ligases and the emergence of resistance through mutations in UPS components restrict this approach. Therefore, identifying alternative E3 ligases suitable for TPD is important to develop new degraders and overcome potential resistance mechanisms. Here, we use a protein-based degrader method, by fusing an anti-tag intracellular antibody to an E3 ligase, to screen E3 ligases enabling the degradation of a tagged POI. We identify SOCS7 E3 ligase as effective biodegrader, able to deplete its target in various cell lines regardless of the POI's subcellular localization. We show its utility by generating a SOCS7-based KRAS degrader that inhibits mutant KRAS pancreatic cancer cells' proliferation. These findings highlight SOCS7 versatility as valuable E3 ligase for generating potent degraders.

Camelid-derived Tcell engagers harnessing human γδ T cells as promising antitumor immunotherapeutic agents.

In the last decade, there has been a surge in developing immunotherapies to enhance the immune system's ability to eliminate tumor cells. Bispecific antibodies known as T cell engagers (TCEs) present an attractive strategy in this pursuit. TCEs aim to guide cytotoxic T cells toward tumor cells, thereby inducing a strong activation and subsequent tumor cell lysis. In this study, we investigated the activity of different TCEs on both conventional alpha-beta (αß) T cells and unconventional gamma delta (γδ) T cells. TCEs were built using camelid single-domain antibodies (VHHs) targeting the tumor-associated antigen CEACAM5 (CEA), together with T cell receptor chains or a CD3 domain. We show that Vγ9Vδ2 T cells display stronger in vitro antitumor activity than αß T cells when stimulated with a CD3xCEA TCE. Furthermore, restricting the activation of fresh human peripheral T cells to Vγ9Vδ2 T cells limited the production of protumor factors and proinflammatory cytokines, commonly associated with toxicity in patients. Taken together, our findings provide further insights that γδ T cell-specific TCEs hold promise as specific, effective, and potentially safe molecules to improve antitumor immunotherapies.

Lysosomal membrane proteins LAMP1 and LIMP2 are segregated in the Golgi apparatus independently of their clathrin adaptor binding motif.

To reach the lysosome, lysosomal membrane proteins (LMPs) are translocated in the endoplasmic reticulum after synthesis and then transported to the Golgi apparatus. The existence of a direct transport from the Golgi apparatus to the endosomes but also of an indirect route through the plasma membrane has been described. Clathrin adaptor binding motifs contained in the cytosolic tail of LMPs have been described as key players in their intracellular trafficking. Here we used the RUSH assay to synchronize the biosynthetic transport of multiple LMPs. After exiting the Golgi apparatus, RUSH-synchronized LAMP1 was addressed to the cell surface both after overexpression or at endogenous level. Its YXXΦ motif was not involved in the transport from the Golgi apparatus to the plasma membrane but in its endocytosis. LAMP1 and LIMP2 were sorted from each other after reaching the Golgi apparatus. LIMP2 was incorporated in punctate structures for export from the Golgi apparatus from which LAMP1 is excluded. LIMP2-containing post-Golgi transport intermediates did not rely neither on its adaptor binding signal nor on its C-terminal cytoplasmic domain.