Cellular senescence, characterized by stable cell cycle arrest, plays an important role in aging and age-associated pathologies. Eliminating senescent cells rejuvenates aged tissues and ameliorates age-associated diseases. Here, we identified that natural killer group 2 member D ligands (NKG2DLs) are up-regulated in senescent cells in vitro, regardless of stimuli that induced cellular senescence, and in various tissues of aged mice and nonhuman primates in vivo. Accordingly, we developed and demonstrated that chimeric antigen receptor (CAR) T cells targeting human NKG2DLs selectively and effectively diminish human cells undergoing senescence induced by oncogenic stress, replicative stress, DNA damage, or P16 ^INK4a overexpression in vitro. Targeting senescent cells with mouse NKG2D-CAR T cells alleviated multiple aging-associated pathologies and improved physical performance in both irradiated and aged mice. Autologous T cells armed with the human NKG2D CAR effectively delete naturally occurring senescent cells in aged nonhuman primates without any observed adverse effects. Our findings establish that NKG2D-CAR T cells could serve as potent and selective senolytic agents for aging and age-associated diseases driven by senescence.

The mechanism by which inflammasome activation is modulated remains unclear. In this study, we identified an AIM2-interacting protein, the E3 ubiquitin ligase HUWE1, which was also found to interact with NLRP3 and NLRC4 through the HIN domain of AIM2 and the NACHT domains of NLRP3 and NLRC4. The BH3 domain of HUWE1 was important for its interaction with NLRP3, AIM2, and NLRC4. Caspase-1 maturation, IL-1β release, and pyroptosis were reduced in Huwe1-deficient bone marrow–derived macrophages (BMDMs) compared with WT BMDMs in response to stimuli to induce NLRP3, NLRC4, and AIM2 inflammasome activation. Furthermore, the activation of NLRP3, NLRC4, and AIM2 inflammasomes in both mouse and human cells was remarkably reduced by treatment with the HUWE1 inhibitor BI8622. HUWE1 mediated the K27-linked polyubiquitination of AIM2, NLRP3, and NLRC4, which led to inflammasome assembly, ASC speck formation, and sustained caspase-1 activation. Huwe1-deficient mice had an increased bacterial burden and decreased caspase-1 activation and IL-1β production upon Salmonella, Francisella, or Acinetobacter baumannii infection. Our study provides insights into the mechanisms of inflammasome activation as well as a potential therapeutic target against bacterial infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has been detected in almost all organs of coronavirus disease-19 patients, although some organs do not express angiotensin-converting enzyme-2 (ACE2), a known receptor of SARS-CoV-2, implying the presence of alternative receptors and/or co-receptors. Here, we show that the ubiquitously distributed human transferrin receptor (TfR), which binds to diferric transferrin to traffic between membrane and endosome for the iron delivery cycle, can ACE2-independently mediate SARS-CoV-2 infection. Human, not mouse TfR, interacts with Spike protein with a high affinity (K _D ~2.95 nM) to mediate SARS-CoV-2 endocytosis. TfR knock-down (TfR-deficiency is lethal) and overexpression inhibit and promote SARS-CoV-2 infection, respectively. Humanized TfR expression enables SARS-CoV-2 infection in baby hamster kidney cells and C57 mice, which are known to be insusceptible to the virus infection. Soluble TfR, Tf, designed peptides blocking TfR-Spike interaction and anti-TfR antibody show significant anti-COVID-19 effects in cell and monkey models. Collectively, this report indicates that TfR is a receptor/co-receptor of SARS-CoV-2 mediating SARS-CoV-2 entry and infectivity by likely using the TfR trafficking pathway.

Background: Glioblastoma (GBM) is recognized as among the most aggressive forms of brain tumor. Patients typically present with a five-year survival rate of less than 6% with traditional surgery and chemoradiotherapy, which calls for novel immunotherapies like chimeric antigen receptor T (CAR-T) cells therapy. In response to endoplasmic reticulum (ER) stress in multiple tumor cells including GBM, the glucose-regulated protein 78 (GRP78) expression increases and the protein is partially translocated to the cell surface, while it is restricted to the cytoplasm and the nucleus in normal cells. Methods: In this study, to target the cell surface GRP78 (csGRP78), CAR-T cells based on its binding peptide were generated. In vitro two GBM cell lines and glioma stem cells (GSCs) were used to confirm the localization of csGRP78 and the cytotoxicity of the CAR-T cells. In vivo a GBM xenograft model was used to assess the killing activity and the safety of the CAR-T cells. Results: We confirmed the localization of csGRP78 at the cell surface of two GBM cell lines (U-251MG and U-87MG) and in GSCs. Co-culture experiments revealed that the CAR-T cells could specifically kill the GBM tumor cells and GSCs with specific IFN-γ release. Furthermore, in the tumor xenograft model, the CAR-T cells could decrease the number of GSCs and significantly suppress tumor cell growth. Importantly, we found no obvious off-target effects or T cell infiltration in major organs following systemic administration of these cells. Conclusions: The csGRP78 targeted CAR-T cells efficiently kill GBM tumor cells and GSCs both in vitro and in vivo, and ultimately suppress the xenograft tumors growth without obvious tissue injuries. Therefore, our study demonstrates that csGRP78 represents a valuable target and the csGRP78-targeted CAR-T cells strategy is an effective immunotherapy against GBM.

• CD6-CAR-T cells based on the extracellular domain of CD6 exerts a cytotoxic effect by targeting CD166, not CD318. • CD6-CAR-T cells were highly efficacious against CD166-positive tumor cells. • CD6-CAR-T cells show potent cytotoxicity targeting CRC CSCs.

Background: The receptor tyrosine kinases TAM family (TYRO3, AXL, and MERTK) are highly expressed in multiple forms of cancer cells and tumor-associated macrophages and promote the development of cancers including pancreatic tumor. Targeting TAM receptors could be a promising therapeutic option. Methods: We designed a novel CAR based on the extracellular domain of growth arrest-specific protein 6 (GAS6), a natural ligand for all TAM members. The ability of CAR-T to kill pancreatic cancer cells is tested in vitro and in vivo, and the safety is evaluated in mice and nonhuman primate. Results: GAS6-CAR-T cells efficiently kill TAM-positive pancreatic cancer cell lines, gemcitabine-resistant cancer cells, and cancer stem-like cells in vitro. GAS6-CAR-T cells also significantly suppressed the growth of PANC1 xenografts and patient-derived xenografts in mice. Furthermore, these CAR-T cells did not induce obvious side effects in nonhuman primate or mice although the CAR was demonstrated to recognize mouse TAM. Conclusions: Our findings indicate that GAS6-CAR-T-cell therapy may be effective for pancreatic cancers with low toxicity.