Next generation chimeric antigen receptor (CAR) T-cell therapies hold promise in providing a curative solution for solid tumors. Nonetheless, the significant clinical hurdle of on-target off-tumor toxicity (OTOT) remains a major downside to the advancement of CAR T-cell therapies.
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.
DNA crosslinks block DNA replication and are repaired by the Fanconi Anemia pathway. The FANCD2-FANCI (D2-I) protein complex is central to this process as it initiates repair and is also known to play a more general role in DNA repair and in protecting stalled replication forks from unscheduled degradation.
Here, using single-molecule imaging, we investigate the behaivior of D2-I and provide a unified molecular mechanism that reconciles the roles of D2-I in recognition and protection of stalled replication forks in multiple DNA repair pathways.
Commercial CAR-T therapies still suffer from severe limitations, as majority of patients fail to achieve complete response and ultimately relapse.
Watch this Webinar to discover Prof. Marco Ruella’s team have adopted a novel CAR-T avidity screening method to improve safety and exhaustion profile, leading to 100% clinical response in a phase I trial.
Mechanistic issues limit the effectiveness of many current cancer-targeting antibody therapies, with monospecific antibodies often hindered by receptor dimerization and activation. Biparatopic antibodies, which bind to two unique non-overlapping epitopes, offer a promising solution with stronger binding, more potent antagonism, and higher specificity.
Next generation chimeric antigen receptor (CAR) T-cell therapies hold promise in providing a curative solution for solid tumors. Nonetheless, the significant clinical hurdle of on-target off-tumor toxicity (OTOT) remains a major downside to the advancement of CAR T-cell therapies.
Interestingly, 8 out of 10 CAR T lead candidates fail after preclinical development. This suggests that current preclinical in vitro assays insufficiently predictive. This status quo could worsen as the field moves into tackling more challenging targets such as solid tumors, which require more sophisticated next generation designs like dual-targeting, logic-gating or BiTE-secreting CARs. However, implementing Cell Avidity analysis at an early stage in the drug development process can help to identify superior lead candidates and improve (pre)clinical correlation, as shown by this study.
Precisely manipulating genetic material at the single molecule level is gaining importance across life sciences – and so do the tools that allow researchers to do exactly that. The C-Trap system combines single molecule fluorescence microscopy with optical tweezers to manipulate DNA, allowing researchers to directly observe and track molecular events as they occur. Designing and creating specific DNA constructs is crucial for maximizing the potential of single molecule studies. In this application note we introduce the powerful combination of cutting edge biochemistry and single-molecule visualization methods to increase throughput and maximize the results gained from each individual measurement.
Dive into the cutting-edge fusion of nuclear extracts and dynamic single-molecule (DSM) analysis in LUMICKS’ C-Trap. This approach revolutionizes how DNA-protein interactions are studied, offering unmatched biological relevance and accessibility. Biology researchers eager to unravel the complexities of nuclear processes will find this application note a game-changer. Explore how the C-Trap elevates molecular biology research to unprecedented heights.