Improving CAR-T immunotherapy for lymphoma by fine-tuning avidity

The nuclear envelope protects the genome from mechanical stress during processes such as migration, division, and compression , but how it buffers forces at the scale of DNA remains unclear. Here, we utilize optical tweezers to show that a multivalent protein–DNA co-condensate containing the nuclear envelope protein LEM2 and the DNA-binding protein BAF shield DNA beyond its melting point at 65 pN. Under load, their collective assembly induces an unconventional DNA stiffening effect that provides mechanical reinforcement, dependent on the intrinsically disordered region (IDR) of LEM2. Within cells, these components form an elastic surface hydrogel at the nuclear periphery, visible as a continuous surface by cryo-electron tomography. Disruption of this surface hydrogel increases DNA damage and micronuclei formation during nuclear deformation. Together, this work expands the functional repertoire of condensates, revealing a load-responsive nuclear surface hydrogel at the mesoscale that mitigates mechanical stress.

Translocations involving FGFR2 gene fusions are common in cholangiocarcinoma and gastric carcinoma and predict response to FGFR kinase inhibitors. However, response rates and durability are limited due to the emergence of resistance, typically involving FGFR2 kinase domain mutations, and to sub-optimal dosing, relating to adverse drug effects.

This webcast will present new work showing that the vast majority of such alterations retain the extracellular domain (ECD), potentially enabling highly selective targeting of the FGFR2 ECD using biotherapeutics.

To improve on the activity of traditional bivalent monotopic antibodies, the Sellers lab systematically generated biparatopic antibodies targeting distinct epitope pairs in FGFR2 ECD, and identified antibodies that effectively block signaling and malignant growth driven by FGFR2-fusions.

These antibodies robustly blocked proliferation and colony formation in FGFR2-fusion driven cholangiocarcinoma and demonstrated robust in vivo anti-tumour activity. In vivo activity was marked by significant antibody-mediated downregulation of FGFR2 and in turn this was associated with robust lysosomal internalization enacted by the two biparatopics. In vitro, the biparatopic antibodies demonstrated activity against FGFR inhibitor resistant alleles of FGFR2. The internalization properties of the antibodies also make them suitable for exploration as antibody-drug conjugates

Structural Maintenance of Chromosome (SMC) complexes, cohesin, condensin and Smc5/6, play a fundamental role in genome organization, facilitating chromosome compaction, segregation, and DNA repair. Despite their essential functions, the mechanisms by which the complexes interact with different DNA substrates and influence topological transitions remain not fully understood. Using the LUMICKS C-Trap, we have employed single-molecule approaches to analyze the behavior of purified SMC complexes, focusing on yeast cohesin and human SMC5/6, on different DNA substrates, including double-stranded (dsDNA) and single-stranded DNA (ssDNA). Additionally, we have used a quadrupole optical trap to bridging by SMC complexes and their effect on DNA decatenation by Topoisomerase IIα (Top2A). These findings provide new insights into the fundamental properties and requirements of cohesin and Smc5/6’s interaction with DNA substrates, as well as their ability to bridge two independent DNAs.

What are the rules of membrane tension propagation in cells? For proper function, cells need to link short-range biochemical signaling events with long-range integration of cell physiology. Forces transmitted through the plasma membrane are thought to serve as this globally integrator. However, conflicting observations have left the field divided as to whether cell membranes support or resist tension propagation. This discrepancy likely originates from the use of exogenous forces that may not accurately mimic endogenous forces. We overcome this complication by leveraging optogenetics to directly control localized actin-based protrusions or actomyosin contractions while simultaneously monitoring the propagation of membrane tension using dual-trap optical tweezers. Surprisingly, actin-driven protrusions and actomyosin contractions both elicit rapid global membrane tension propagation, whereas forces applied to cell membranes alone do not. We present a simple unifying mechanical model in which mechanical forces that engage the actin cortex drive rapid, robust membrane tension propagation through long-range membrane flows.

Genome replication and gene expression are carried out by macromolecular machines that exist at nanometer scale and generate piconewton forces. Challenged by the hierarchical chromatin organization and omnipresent thermal fluctuations, these DNA-based machines still accomplish their tasks with remarkable efficiency and accuracy. We leverage single-molecule techniques, particularly correlative fluorescence and force microscopy (smCFFM), to probe the dynamics and mechanics of replication, transcription, and chromatin machinery. These investigations have yielded new insights into the principles of genetic and epigenetic inheritance.