Targeting of the alphav beta3 integrin complex by CAR-T cells leads to rapid regression of diffuse intrinsic pontine glioma and glioblastoma.

BACKGROUND: Diffuse intrinsic pontine glioma (DIPG) and glioblastoma (GBM) are two highly aggressive and generally incurable gliomas with little therapeutic advancements made in the past several decades. Despite immense initial success of chimeric antigen receptor (CAR) T cells for the treatment of leukemia and lymphoma, significant headway into the application of CAR-T cells against solid tumors, including gliomas, is still forthcoming. The integrin complex alphav beta3 (αvß3) is present on multiple and diverse solid tumor types and tumor vasculature with limited expression throughout most normal tissues, qualifying it as an appealing target for CAR-T cell-mediated immunotherapy. METHODS: Patient-derived DIPG and GBM cell lines were evaluated by flow cytometry for surface expression of αvß3. Second-generation CAR-T cells expressing an anti-αvß3 single-chain variable fragment were generated by retroviral transduction containing either a CD28 or 4-1BB costimulatory domain and CD3zeta. CAR-T cells were evaluated by flow cytometry for CAR expression, memory phenotype distribution, and inhibitory receptor profile. DIPG and GBM cell lines were orthotopically implanted into NSG mice via stereotactic injection and monitored with bioluminescent imaging to evaluate αvß3 CAR-T cell-mediated antitumor responses. RESULTS: We found that patient-derived DIPG cells and GBM cell lines express high levels of surface αvß3 by flow cytometry, while αvß3 is minimally expressed on normal tissues by RNA sequencing and protein microarray. The manufactured CAR-T cells consisted of a substantial frequency of favorable early memory cells and a low inhibitory receptor profile. αvß3 CAR-T cells demonstrated efficient, antigen-specific tumor cell killing in both cytotoxicity assays and in in vivo models of orthotopically and stereotactically implanted DIPG and GBM tumors into relevant locations in the brain of NSG mice. Tumor responses were rapid and robust with systemic CAR-T cell proliferation and long-lived persistence associated with long-term survival. Following tumor clearance, TCF-1+αvß3 CAR-T cells were detectable, underscoring their ability to persist and undergo self-renewal. CONCLUSIONS: These results highlight the potential of αvß3 CAR-T cells for immunotherapeutic treatment of aggressive brain tumors with reduced risk of on-target, off-tumor mediated toxicity due to the restricted nature of αvß3 expression in normal tissues.

mTORC1 Enhances Early Phase Ribosome Processivity.

During translation elongation, the ribosome serially adds amino acids to a growing polypeptide over many rounds of catalysis. The ribosome remains bound to mRNAs over these multiple catalytic cycles, requiring high processivity. Despite its importance to translation, relatively little is known about how mRNA sequences or signaling pathways might enhance or reduce ribosome processivity. Here, we describe a metric for ribosome processivity, the ribosome density index (RDI), which is readily calculated from ribosomal profiling data. We show that ribosome processivity is not strongly influenced by open-reading frame (ORF) length or codon optimality. However, we do observe that ribosome processivity exists in two phases and that the early phase of ribosome processivity is enhanced by mTORC1, a key translational regulator. By showing that ribosome processivity is regulated, our findings suggest an additional layer of control that the cell can exert to govern gene expression.