Spatial Mapping and Immunomodulatory Role of the OX40/OX40L Pathway in Human Non-Small Cell Lung Cancer.

PURPOSE: To evaluate the tissue distribution and clinical significance of OX40 and OX40L in human non-small cell lung cancer (NSCLC). EXPERIMENTAL DESIGN: Using multiplexed quantitative immunofluorescence, we conducted simultaneous and localized measurements of OX40 and OX40L proteins, major T-cell subsets, and conventional type 1 dendritic cells (cDC1) in 614 primary NSCLCs from three independent cohorts represented in tissue microarrays. We also measured OX40L protein in samples from a phase I clinical trial of intratumor administration of a lipid nanoparticle encapsulated mRNA encoding OX40L (mRNA-2416) in human solid tumors. Finally, we studied the OX40 pathway in 212 uterine/ovarian serous carcinomas. RESULTS: OX40 protein was expressed in approximately 90% of NSCLCs, and OX40L was detected in approximately 10% of cases. Increased expression of OX40 was associated with higher CD4+ and CD8+ T lymphocytes, as well as cDC1s. Elevated expression of OX40L was consistently associated with increased CD4+ tumor-infiltrating lymphocytes and longer overall survival. No association was found between OX40 or OX40L levels and oncogenic driver mutations in EGFR and KRAS in lung adenocarcinomas. Delivering OX40L mRNA using intratumor mRNA-2416 injection mediated increased local OX40L protein levels that was most prominent in a patient with ovarian serous carcinoma. Detectable OX40L protein levels were observed in 15% of primary uterine/ovarian serous malignancies and associated with longer survival. CONCLUSIONS: The OX40 pathway is expressed in a fraction of NSCLCs and is associated with a favorable immune contexture. Although OX40L is uncommonly expressed in NSCLC and serous malignancies, it is associated with better prognosis and can be introduced using exogenous mRNA.

Differential immunomodulatory effect of PARP inhibition in BRCA1 deficient and competent tumor cells.

Poly-ADP-ribose polymerase (PARP) inhibitors are active against cells and tumors with defects in homology-directed repair as a result of synthetic lethality. PARP inhibitors (PARPi) have been suggested to act by either catalytic inhibition or by PARP localization in chromatin. In this study, we treat BRCA1 mutant cells derived from a patient with triple negative breast cancer and control cells for three weeks with veliparib, a PARPi, to determine if treatment with this drug induces increased levels of mutations and/or an inflammatory response. We show that long-term treatment with PARPi induces an inflammatory response in HCC1937 BRCA1 mutant cells. The levels of chromatin-bound PARP1 in the BRCA1 mutant cells correlate with significant upregulation of inflammatory genes and activation of the cyclic GMP-AMP synthase (cGAS)/signaling effector stimulator of interferon genes (STING pathway). In contrast, an increased mutational load is induced in BRCA1-complemented cells treated with a PARPi. Our results suggest that long-term PARP inhibitor treatment may prime both BRCA1 mutant and wild-type tumors for positive responses to immune checkpoint blockade, but by different underlying mechanisms.

Uncoupling the Senescence-Associated Secretory Phenotype from Cell Cycle Exit via Interleukin-1 Inactivation Unveils Its Protumorigenic Role.

Cellular senescence has emerged as a potent tumor suppressor mechanism in numerous human neoplasias. Senescent cells secrete a distinct set of factors, collectively termed the senescence-associated secretory phenotype (SASP), which has been postulated to carry both pro- and antitumorigenic properties depending on tissue context. However, the in vivo effect of the SASP is poorly understood due to the difficulty of studying the SASP independently of other senescence-associated phenotypes. Here, we report that disruption of the interleukin-1 (IL-1) pathway completely uncouples the SASP from other senescence-associated phenotypes such as cell cycle exit. Transcriptome profiling of IL-1 receptor (IL-1R)-depleted senescent cells indicates that IL-1 controls the late arm of the senescence secretome, which consists of proinflammatory cytokines induced by NF-κB. Our data suggest that both IL-1α and IL-1ß signal through IL-1R to upregulate the SASP in a cooperative manner. Finally, we show that IL-1α inactivation impairs tumor progression and immune cell infiltration without affecting cell cycle arrest in a mouse model of pancreatic cancer, highlighting the protumorigenic property of the IL-1-dependent SASP in this context. These findings provide novel insight into the therapeutic potential of targeting the IL-1 pathway in inflammatory cancers.

Neoadjuvant nivolumab modifies the tumor immune microenvironment in resectable glioblastoma.

Glioblastoma is the most common primary central nervous system malignancy and has a poor prognosis. Standard first-line treatment, which includes surgery followed by adjuvant radio-chemotherapy, produces only modest benefits to survival1,2. Here, to explore the feasibility, safety and immunobiological effects of PD-1 blockade in patients undergoing surgery for glioblastoma, we conducted a single-arm phase II clinical trial (NCT02550249) in which we tested a presurgical dose of nivolumab followed by postsurgical nivolumab until disease progression or unacceptable toxicity in 30 patients (27 salvage surgeries for recurrent cases and 3 cases of primary surgery for newly diagnosed patients). Availability of tumor tissue pre- and post-nivolumab dosing and from additional patients who did not receive nivolumab allowed the evaluation of changes in the tumor immune microenvironment using multiple molecular and cellular analyses. Neoadjuvant nivolumab resulted in enhanced expression of chemokine transcripts, higher immune cell infiltration and augmented TCR clonal diversity among tumor-infiltrating T lymphocytes, supporting a local immunomodulatory effect of treatment. Although no obvious clinical benefit was substantiated following salvage surgery, two of the three patients treated with nivolumab before and after primary surgery remain alive 33 and 28 months later.

Reversible dual inhibitor against G9a and DNMT1 improves human iPSC derivation enhancing MET and facilitating transcription factor engagement to the genome.

The combination of defined factors with small molecules targeting epigenetic factors is a strategy that has been shown to enhance optimal derivation of iPSCs and could be used for disease modelling, high throughput screenings and/or regenerative medicine applications. In this study, we showed that a new first-in-class reversible dual G9a/DNMT1 inhibitor compound (CM272) improves the efficiency of human cell reprogramming and iPSC generation from primary cells of healthy donors and patient samples, using both integrative and non-integrative methods. Moreover, CM272 facilitates the generation of human iPSC with only two factors allowing the removal of the most potent oncogenic factor cMYC. Furthermore, we demonstrated that mechanistically, treatment with CM272 induces heterochromatin relaxation, facilitates the engagement of OCT4 and SOX2 transcription factors to OSKM refractory binding regions that are required for iPSC establishment, and enhances mesenchymal to epithelial transition during the early phase of cell reprogramming. Thus, the use of this new G9a/DNMT reversible dual inhibitor compound may represent an interesting alternative for improving cell reprogramming and human iPSC derivation for many different applications while providing interesting insights into reprogramming mechanisms.

Reversal of hyperglycemia by insulin-secreting rat bone marrow- and blastocyst-derived hypoblast stem cell-like cells.

ß-cell replacement may efficiently cure type 1 diabetic (T1D) patients whose insulin-secreting ß-cells have been selectively destroyed by autoantigen-reactive T cells. To generate insulin-secreting cells we used two cell sources: rat multipotent adult progenitor cells (rMAPC) and the highly similar rat extra-embryonic endoderm precursor (rXEN-P) cells isolated under rMAPC conditions from blastocysts (rHypoSC). rMAPC/rHypoSC were sequentially committed to definitive endoderm, pancreatic endoderm, and ß-cell like cells. On day 21, 20% of rMAPC/rHypoSC progeny expressed Pdx1 and C-peptide. rMAPCr/HypoSC progeny secreted C-peptide under the stimulus of insulin agonist carbachol, and was inhibited by the L-type voltage-dependent calcium channel blocker nifedipine. When rMAPC or rHypoSC differentiated d21 progeny were grafted under the kidney capsule of streptozotocin-induced diabetic nude mice, hyperglycemia reversed after 4 weeks in 6/10 rMAPC- and 5/10 rHypoSC-transplanted mice. Hyperglycemia recurred within 24 hours of graft removal and the histological analysis of the retrieved grafts revealed presence of Pdx1-, Nkx6.1- and C-peptide-positive cells. The ability of both rMAPC and HypoSC to differentiate to functional ß-cell like cells may serve to gain insight into signals that govern ß-cell differentiation and aid in developing culture systems to commit other (pluripotent) stem cells to clinically useful ß-cells for cell therapy of T1D.