The proper regulation of ICOS and ICOS ligand (ICOSL) has been shown to be essential for maintaining proper immune homeostasis. Loss of either protein results in defective humoral immunity, and overexpression of ICOS results in aberrant Ab production resembling lupus. How ICOSL is regulated in response to ICOS interaction is still unclear. We demonstrate that a disintegrin and metalloproteinase (ADAM)10 is the primary physiological sheddase of ICOSL in mice and humans. Using an in vivo system in which ADAM10 is deleted only on B cells, elevated levels of ICOSL were seen. This increase is also seen when ADAM10 is deleted from human B cell lines. Identification of the primary sheddase has allowed the characterization of a novel mechanism of ICOS regulation. In wild-type mice, interaction of ICOS/ICOSL results in ADAM10-induced shedding of ICOSL on B cells and moderate ICOS internalization on T cells. When this shedding is blocked, excessive ICOS internalization occurs. This results in severe defects in T follicular helper development and T_H2 polarization, as seen in a house dust mite exposure model. In addition, enhanced T_H1 and T_H17 immune responses are seen in experimental autoimmune encephalomyelitis. Blockade of ICOSL rescues T cell ICOS surface expression and rescues, at least in part, T follicular helper numbers and the abnormal Ab production previously reported in these mice. Overall, we propose a novel regulation of the ICOS/ICOSL axis, with ADAM10 playing a direct role in regulating ICOSL, as well as indirectly regulating ICOS, thus controlling ICOS/ICOSL-dependent responses.

Effective antitumor immunity ultimately depends on the priming and activation of tumor-specific cytotoxic CD8+ T cells; however, the role of intratumoral cell–cell immune interactions remains incompletely understood. Recent work has revealed that the temporospatial co-localization of dendritic cells (DC), T helper (Th) cells, and cytotoxic T lymphocytes (CTL) within the tumor immune microenvironment following immune checkpoint blockade correlates with clinical response. In this study, we report the integration of more than 1 million spatially resolved single-cell profiles across six spatial proteomic and transcriptomic assays, which demonstrated that DC:Th:CTL three-cell-type clusters were common even in immunotherapy-naïve and highly desmoplastic tumors, such as fibrolamellar carcinoma and pancreatic ductal adenocarcinoma. We found that these immune triads were enriched for functionally important type 1 conventional DC, mature DCs enriched in immunoregulatory molecules, CXCL13+ Th, and GZMK+ effector CTL phenotypes. Subsequent multiplex immunofluorescence imaging of more than 450 primary pancreatic ductal adenocarcinoma tumors showed that the density of antigen-presenting cell:Th:CTL three-cell-type clusters was correlated with intratumoral T-cell clonal expansion and improved overall survival. These findings suggest that DC:Th:CTL triads are conserved across solid tumors and highlight the importance of intratumoral spatial niches in mediating endogenous antitumor immunity.

Background: Stromal hyaluronic acid (HA) poses a physical barrier and protects tumor cells from immune surveillance. Stroma targeting with pegylated human recombinant PH20 hyaluronidase (PEGPH20) demonstrated improved infiltration of cytotoxic T-lymphocytes and delivery of chemotherapy and PD1/PD-L1 antibodies in tumor models. This multicenter phase II study of PEGPH20 plus pembrolizumab evaluated the efficacy, safety and immune and stromal biomarkers in patients with HA-high refractory metastatic pancreatic ductal adenocarcinoma (mPDA). Patients and Methods: Patients were treated with PEGPH20 3 µg/kg IV weekly and pembrolizumab 200 mg IV in 3-week cycles. Tumor and blood samples were collected at baseline and on-study for biomarker analyses. Results: Between May and November 2019, 38 patients were screened and 8 treated, with median age 68 years (range 60–73) and median two (range 1–4) prior therapies. The study was closed to accrual early by pharmaceutical sponsor. Treatment was well tolerated, with expected grade 1/2 musculoskeletal toxicities. Best response was stable disease in 2 of 7 evaluable patients (29%). Median overall and progression-free survival were 7.2 months (95% CI 1.2–11.8) and 1.5 months (95% CI 0.9–4.4), respectively. Prolonged survival (range 10.2–27.6 months) occurred in patients treated with subsequent chemotherapy. Higher baseline tumor T cell receptor (TCR) clonality correlated with longer survival. Conclusions: Pembrolizumab with PEGPH20 was safe but did not have significant efficacy in refractory HA-high metastatic PDA.

Bone morphogenetic protein (BMP) gene delivery to Lewis rat lumbar intervertebral discs (IVDs) drives bone formation anterior and external to the IVD, suggesting the IVD is inhospitable to osteogenesis. This study was designed to determine if IVD destruction with a proteoglycanase, and/or generating an IVD blood supply by gene delivery of an angiogenic growth factor, could render the IVD permissive to intra-discal BMP-driven osteogenesis and fusion. Surgical intra-discal delivery of naïve or gene-programmed cells (BMP2/BMP7 co-expressing or VEGF₁₆₅ expressing) +/- purified chondroitinase-ABC (chABC) in all permutations was performed between lumbar 4/5 and L5/6 vertebrae, and radiographic, histology, and biomechanics endpoints were collected. Follow-up anti-sFlt Western blotting was performed. BMP and VEGF/BMP treatments had the highest stiffness, bone production and fusion. Bone was induced anterior to the IVD, and was not intra-discal from any treatment. chABC impaired BMP-driven osteogenesis, decreased histological staining for IVD proteoglycans, and made the IVD permissive to angiogenesis. A soluble fragment of VEGF Receptor-1 (sFlt) was liberated from the IVD matrix by incubation with chABC, suggesting dysregulation of the sFlt matrix attachment is a possible mechanism for the chABC-mediated IVD angiogenesis we observed. Based on these results, the IVD can be manipulated to foster vascular invasion, and by extension, possibly osteogenesis.

Supplemental Table S3. Characteristics for PDAC patients undergoing upfront resection included in the Xenium Prime dataset

Supplemental Figure S2. KPC tumor cell types.