Advancing T cell-based immunotherapies through targeted engineering with CRISPR-Cas9

T cell-based immunotherapies such as chimeric antigen receptor T (CAR-T) cell therapy have undoubtably revolutionized the treatment of cancer. However, the broad effectiveness of CAR-T cell therapy is hindered by several unresolved problems, most notably a lack of therapeutic efficacy in treating solid tumor cancers. A second challenge stems from the widespread use of viral vectors in CAR-T manufacturing, which poses safety risks to patients receiving treatment. Here, we showed that genome editing with CRISPR-Cas9 can be used to overcome both of these issues. As the solid tumor microenvironment (TME) is known to be metabolically suppressive, we devised a single-step editing method to enhance the metabolism and effector function of CAR-T cells. This approach combined CRISPR-mediated homology-directed repair with a gene-trap approach to link CAR integration with simultaneous deletion of a metabolic gene of interest. For proof-of-concept, we targeted the folate receptor alpha (aFR) CAR to the locus of the essential autophagy gene ATG5, and showed that editing at ATG5 could be achieved with a high level of on-target specificity. Functionally, deletion of ATG5 led to alterations in glucose and glutamine metabolism and enhanced CAR-T cell efficacy under nutrient-restricted conditions in vitro and in vivo. To address the safety concerns associated with viral transduction, we developed a process for nonviral manufacturing of clinical-grade CAR-T cells for B-cell malignancies. This approach used electroporation of a Cas9 ribonucleoprotein complexed with a linear double-stranded DNA template to facilitate site-specific insertion of a CD22 CAR at the T cell receptor alpha chain (TRAC) locus. In vitro, nonviral CD22 CAR-T cells exhibited comparable antitumor activity to lentiviral CD22 CAR-T cells. thereby establishing feasibility of our nonviral manufacturing process. Taken together, the results of these studies highlight the broad applicability of CRISPR-Cas9 as a tool for engineering safer, more effective T cell-based immunotherapies for patients with cancer.Graduate2025-11-1

Hunter-gatherer sea voyages extended to remotest Mediterranean islands

The Maltese archipelago is a small island chain that is among the most remote in the Mediterranean. Humans were not thought to have reached and inhabited such small and isolated islands until the regional shift to Neolithic lifeways, around 7.5 thousand years ago (ka)1. In the standard view, the limited resources and ecological vulnerabilities of small islands, coupled with the technological challenges of long-distance seafaring, meant that hunter-gatherers were either unable or unwilling to make these journeys2-4. Here we describe chronological, archaeological, faunal and botanical data that support the presence of Holocene hunter-gatherers on the Maltese islands. At this time, Malta's geographical configuration and sea levels approximated those of the present day, necessitating seafaring distances of around 100 km from Sicily, the closest landmass. Occupations began at around 8.5 ka and are likely to have lasted until around 7.5 ka. These hunter-gatherers exploited land animals, but were also able to take advantage of marine resources and avifauna, helping to sustain these groups on a small island. Our discoveries document the longest yet-known hunter-gatherer sea crossings in the Mediterranean, raising the possibility of unknown, precocious connections across the wider region.Full Tex

Autophagy Regulation of Metabolism Is Required for CD8+ T Cell Anti-tumor Immunity

Summary: Autophagy is a cell survival process essential for the regulation of immune responses to infections. However, the role of T cell autophagy in anti-tumor immunity is less clear. Here, we demonstrate a cell-autonomous role for autophagy in the regulation of CD8+ T-cell-mediated control of tumors. Mice deficient for the essential autophagy genes Atg5, Atg14, or Atg16L1 display a dramatic impairment in the growth of syngeneic tumors. Moreover, T cells lacking Atg5 have a profound shift to an effector memory phenotype and produce greater amounts of interferon-γ (IFN-γ) and tumor necrosis factor α (TNF-α). Mechanistically, Atg5−/− CD8+ T cells exhibit enhanced glucose metabolism that results in alterations in histone methylation, increases in H3K4me3 density, and transcriptional upregulation of both metabolic and effector target genes. Nonetheless, glucose restriction is sufficient to suppress Atg5-dependent increases in effector function. Thus, autophagy-dependent changes in CD8+ T cell metabolism directly regulate anti-tumor immunity. : DeVorkin et al. show that loss of autophagy enhances CD8+ T-cell-mediated rejection of tumors. Mechanistically, suppression of autophagy shifts T cells to a glycolytic phenotype and causes a reduction in S-adenosylmethionine. As a consequence, autophagy-deficient T cells transcriptionally reprogram immune response genes to an effector memory state. Keywords: autophagy, CD8+ T cells, anti-tumor immunity, glycolysis, lactate, SAM, methylatio

The GOGREEN survey: constraining the satellite quenching time-scale in massive clusters at z ≳ 1

International audienceWe model satellite quenching at z ~ 1 by combining 14 massive (1013.8 halo/M⊙ 15) clusters at 0.8 quench), accounts for quenching in our simulated satellite population both at the time of infall by using the observed coeval field quenched fraction and after infall by tuning τquench to reproduce the observed satellite quenched fraction versus stellar mass trend. This model successfully reproduces the observed satellite quenched fraction as a function of stellar mass (by construction), projected cluster-centric radius, and redshift and is consistent with the observed field and cluster stellar mass functions at z ~ 1. We find that the satellite quenching time-scale is mass dependent, in conflict with some previous studies at low and intermediate redshift. Over the stellar mass range probed (M⋆ > 1010 M⊙), we find that the satellite quenching time-scale decreases with increasing satellite stellar mass from ~1.6 Gyr at 1010 M⊙ to ~0.6-1 Gyr at 1011 M⊙ and is roughly consistent with the total cold gas (HI + H2) depletion time-scales at intermediate z, suggesting that starvation may be the dominant driver of environmental quenching at z ⋆ > 1011 M⊙) are quenched prior to infall