Effects of checkpoint kinase 1 inhibition by prexasertib on the tumor immune microenvironment of head and neck squamous cell carcinoma.

Prognosis for patients with recurrent and/or metastatic head and neck squamous cell carcinoma (HNSCC) remains poor. Development of more effective and less toxic targeted therapies is necessary for HNSCC patients. Checkpoint kinase 1 (CHK1) plays a vital role in cell cycle regulation and is a promising therapeutic target in HNSCC. Prexasertib, a CHK1 inhibitor, induces DNA damage and cell death, however, its effect on the tumor immune microenvironment (TIME) is largely unknown. Therefore, we evaluated a short-term and long-term effects of prexasertib in HNSCC and its TIME. Prexasertib caused increased DNA damage and cell death in vitro and significant tumor regression and improved survival in vivo. The gene expression and multiplex immunohistochemistry (mIHC) analyses of the in vivo tumors demonstrated increased expression of genes that are related to T-cell activation and increased immune cell trafficking, and decreased expression of genes that related to immunosuppression. However, increased expression of genes related to immunosuppression emerged over time suggesting evasion of immune surveillances. These findings in gene expression analyses were confirmed using mIHC which showed differential modulation of TIME in the tumor margins and as well as cores over time. These results suggest that evasion of immune surveillance, at least in part, may contribute to the acquired resistance to prexasertib in HNSCC.

Treatment with a VEGFR-2 antibody results in intra-tumor immune modulation and enhances anti-tumor efficacy of PD-L1 blockade in syngeneic murine tumor models.

Prolonged activation of vascular endothelial growth factor receptor-2 (VEGFR-2) due to mis-regulation of the VEGF pathway induces aberrant blood vessel expansion, which supports growth and survival of solid tumors. Therapeutic interventions that inhibit the VEGFR-2 pathway have therefore become a mainstay of cancer treatment. Non-clinical studies have recently revealed that blockade of angiogenesis can modulate the tumor microenvironment and enhance the efficacy of concurrent immune therapies. Ramucirumab is an FDA-approved anti-angiogenic antibody that inhibits VEGFR-2 and is currently being evaluated in clinical studies in combination with anti-programmed cell death (PD-1) axis checkpoint inhibitors (pembrolizumab, durvalumab, or sintilimab) across several cancer types. The purpose of this study is to establish a mechanistic basis for the enhanced activity observed in the combined blockade of VEGFR-2 and PD-1-axis pathways. Pre-clinical studies were conducted in murine tumor models known to be responsive to anti-PD-1 axis therapy, using monoclonal antibodies that block mouse VEGFR-2 and programmed death-ligand 1 (PD-L1). Combination therapy resulted in enhanced anti-tumor activity compared to anti-PD-L1 monotherapy. VEGFR-2 blockade at early timepoints post-anti-PD-L1 therapy resulted in a dose-dependent and transient enhanced infiltration of T cells, and establishment of immunological memory. VEGFR-2 blockade at later timepoints resulted in enhancement of anti-PD-L1-driven immune cell infiltration. VEGFR-2 and PD-L1 monotherapies induced both unique and overlapping patterns of immune gene expression, and combination therapy resulted in an enhanced immune activation signature. Collectively, these results provide new and actionable insights into the mechanisms by which concurrent VEGFR-2 and PD-L1 antibody therapy leads to enhanced anti-tumor efficacy.

The Folate Pathway Inhibitor Pemetrexed Pleiotropically Enhances Effects of Cancer Immunotherapy.

PURPOSE: Combination strategies leveraging chemotherapeutic agents and immunotherapy have held the promise as a method to improve benefit for patients with cancer. However, most chemotherapies have detrimental effects on immune homeostasis and differ in their ability to induce immunogenic cell death (ICD). The approval of pemetrexed and carboplatin with anti-PD-1 (pembrolizumab) for treatment of non-small cell lung cancer represents the first approved chemotherapy and immunotherapy combination. Although the clinical data suggest a positive interaction between pemetrexed-based chemotherapy and immunotherapy, the underlying mechanism remains unknown. EXPERIMENTAL DESIGN: Mouse tumor models (MC38, Colon26) and high-content biomarker studies (flow cytometry, Quantigene Plex, and nCounter gene expression analysis) were deployed to obtain insights into the mechanistic rationale behind the efficacy observed with pemetrexed/anti-PD-L1 combination. ICD in tumor cell lines was assessed by calreticulin and HMGB-1 immunoassays, and metabolic function of primary T cells was evaluated by Seahorse analysis. RESULTS: Pemetrexed treatment alone increased T-cell activation in mouse tumors in vivo, robustly induced ICD in mouse tumor cells and exerted T-cell-intrinsic effects exemplified by augmented mitochondrial function and enhanced T-cell activation in vitro. Increased antitumor efficacy and pronounced inflamed/immune activation were observed when pemetrexed was combined with anti-PD-L1. CONCLUSIONS: Pemetrexed augments systemic intratumor immune responses through tumor intrinsic mechanisms including immunogenic cell death, T-cell-intrinsic mechanisms enhancing mitochondrial biogenesis leading to increased T-cell infiltration/activation along with modulation of innate immune pathways, which are significantly enhanced in combination with PD-1 pathway blockade.See related commentary by Buque et al., p. 6890.

The CDK4/6 Inhibitor Abemaciclib Induces a T Cell Inflamed Tumor Microenvironment and Enhances the Efficacy of PD-L1 Checkpoint Blockade.

Abemaciclib, an inhibitor of cyclin dependent kinases 4 and 6 (CDK4/6), has recently been approved for the treatment of hormone receptor-positive breast cancer. In this study, we use murine syngeneic tumor models and in vitro assays to investigate the impact of abemaciclib on T cells, the tumor immune microenvironment and the ability to combine with anti-PD-L1 blockade. Abemaciclib monotherapy resulted in tumor growth delay that was associated with an increased T cell inflammatory signature in tumors. Combination with anti-PD-L1 therapy led to complete tumor regressions and immunological memory, accompanied by enhanced antigen presentation, a T cell inflamed phenotype, and enhanced cell cycle control. In vitro, treatment with abemaciclib resulted in increased activation of human T cells and upregulated expression of antigen presentation genes in MCF-7 breast cancer cells. These data collectively support the clinical investigation of the combination of abemaciclib with agents such as anti-PD-L1 that modulate T cell anti-tumor immunity.

Unequal access of chromosomal regions to each other in Salmonella: probing chromosome structure with phage lambda integrase-mediated long-range rearrangements.

We have investigated the fluidity of the Salmonella chromosome architecture using the phage lambda site-specific recombination system as a probe. We determined how chromosome position affects the extent of integrase-mediated recombination between pairs of inversely oriented att sites at various loci. We also investigated the accessibility of each chromosomal att site to an extrachromosomal partner carried on a low-copy plasmid. Recombination events were assayed by semi-quantitative polymerase chain reaction of the attP product. The extent of recombination between the chromosome and the plasmid was generally higher than intrachromosomal recombination except for two loci, araA::attL and galT::attL, which gave no detectable recombination with any other locus. Based on 20 intervals, we found that chromosomal locations are not equally accessible to each other. Although multiple factors probably affect accessibility, the most important is the specific combination of the end-points used. Neither the size of the intervals nor the accessibility of individual end-points to extrachromosomal sequences is as important. These results suggest that the chromosome is not completely fluid but rather organized in some way, with barriers that limit the movement of DNA within the cell. The nature of the barriers involved in chromosomal organization remains to be determined.