Biomarker or biomarker testing

Immune checkpoint blockade (ICB) has demonstrated profound effects on the anti-tumor immune response and has revolutionized the treatment landscape of non-small cell lung cancer (NSCLC). While chemotherapy has traditionally been viewed as detrimental to the immune system, recent clinical trials have demonstrated an impressive survival benefit when combining checkpoint blockade with chemotherapy (ICB+chemo) relative to chemotherapy alone. Despite this, many NSCLC patients do not achieve clinical benefit from ICB, even when combined with chemotherapy. The role of tumor mutational burden, and specifically T cells targeting neoantigens derived from these mutations, has been demonstrated in several ICB studies, however these factors have yet to be fully explored in chemotherapy-containing ICB treatment regimens. There is thus an urgent need to understand the effects of ICB+chemo on the anti-tumor immune response and to specifically evaluate how chemotherapy perturbs the phenotype and function of anti-tumor T cells. It is of paramount importance that we understand how PD-1 blockade and chemotherapy synergize to promote anti-tumor immunity in order to better stratify patients most likely to benefit from these treatments. We propose herein to perform the first comprehensive evaluation of the phenotype and function of neoantigen-specific T cells and their dynamics before and after treatment with ICB+chemo relative to ICB alone. The findings from this study will provide a detailed atlas of the immune system dynamics following treatment and will result in the development of an immunometabolic signature that will be correlated with clinical outcome. Importantly, the results of the proposed work will deepen our understanding of the immune response to ICB+chemo and will unravel the mechanisms underlying clinical response in NSCLC patients.

Background: Osimertinib and other “third-generation” EGFR TKIs effectively overcome T790M-mediated resistance in EGFR-mutant lung cancer, but our understanding of acquired resistance in this setting remains limited. Importantly, the intrinsic heterogeneity of resistant cancers leads to clinical challenges. We hypothesize that in-depth study of 3rd-generation TKI resistance and the impact of genetic heterogeneity on treatment response will lead to improved therapeutic options and clinical outcomes. Study Design: We will analyze tumor biopsies, ctDNA and autopsy specimens using a variety of techniques such as next-generation sequencing, digital droplet PCR and whole exome sequencing to characterize heterogeneity in resistant cancers. In addition, we will run a prospective clinical trial to test a novel therapeutic combination designed to minimize heterogeneity and delay progression. On-treatment biopsies and ctDNA collection will be used to study how resistance develops. Impact: This project will leverage our broad array of clinical specimens, well-established translational research program and preexisting industry and academic collaborations to characterize the heterogeneity of resistance in EGFR-mutant lung cancer. We hope to illuminate clinical and biologic implications of heterogeneity, with the ultimate goal of identifying more effective therapies for patients.

Immune-targeted agents have generated antitumor responses that are transforming cancer therapeutics in various tumors types including lung cancer. Despite the impressive responses observed, the majority of patients eventually experience therapeutic resistance after an initial response. Considering the increased cost to patients and health systems both financially and in terms of time spent in management of immune-related adverse effects it has become clear that success of immuno-oncology approaches depends on choosing patient populations most likely to benefit. Our proposed research addresses the urgent unmet clinical need to understand the underlying biology of response and develop molecular assays of response and resistance to immune targeted agents. We propose comprehensive genome-wide analyses of tumors and cell-free circulating DNA to examine how cancer genomes change during immune checkpoint blockade. Our approach is focused on discovery of mechanisms of resistance using a multidimensional strategy that combines comprehensive genomic analyses with functional assays of autologous T-cell reactivity. The underlying premise of this proposal is that through our comprehensive molecular analyses, we will identify changes in the genomic landscape of tumors during immune checkpoint blockade that mediate emergence of primary and acquired resistance to these therapies. Unravelling the genomic mechanisms through which cancer adapts to evade anti-tumor immune responses will be critical for future development of tailored cancer immunotherapy strategies. We envision that the results of the proposed research will be used to develop patient-specific immunotherapy approaches in lung cancer and will launch investigator-initiated clinical trials at Hopkins and other institutions.

One of the challenges for early detection and prevention of lung squamous cell carcinoma is the lack of understanding of the molecular and cellular changes that cause initiation and progression of this disease. Airway premalignant lesions are the precursors of lung squamous cell carcinoma and can be detected and monitored by autofluorescence bronchoscopy. However, many of these lesions will regress without clinical intervention, and only a subset will progress to invasive carcinoma. The goal of this study is to characterize the landscape of somatic mutations in airway premalignant lesions and to develop an early detection biomarker using targeted DNA sequencing that can predict risk of lesion progression. Ultra-deep whole-exome sequencing will be performed on a previously collected cohort of airway premalignant lesions from high-risk smokers that have been sampled over time, characterized histologically, and classified into those that are stable, regress, or progress. Significantly mutated genes, recurrent copy number changes, and mutational signatures will be identified, and a determination will be made whether they are associated with lesion histology or progression or regression status. Finally, the findings from this study will be combined with the driver genes from The Cancer Genome Atlas (TCGA) to build a targeted DNA sequencing panel. This panel will serve as a rapid and relatively inexpensive method for monitoring premalignant lesions from patients over time and can be used in a clinical trial to assess the ability of autofluorescence bronchoscopy in combination with molecular profiling to stratify patients and serve as a companion diagnostic in chemoprevention trials.

An estimated 160,000 patients die each year from non-small cell lung cancer (NSCLC), primarily due to the development of metastatic and treatment-refractory disease. Dr. Byers, Dr. Gibbons, and their group have previously demonstrated that the epithelial-mesenchymal transition (EMT) is a unifying mechanism that drives tumor progression, metastasis, resistance to standard therapies, and immune suppression. Furthermore, their group and others have shown that the receptor tyrosine kinase Axl and PD-L1 are overexpressed on tumor cells in the setting of EMT in NSCLC and that targeting either Axl or PD-L1 results in preclinical efficacy in lung cancer models.

Based on these results, they have initiated the first clinical trials of BGB324 (a first-in-class, selective Axl inhibitor) for lung cancer patients in combination with either erlotinib or chemotherapy. They hypothesize that Axl is a driver of EMT and EMT-associated immune escape. Therefore, in this project they will use pre-clinical cell line and animal models to determine the role of Axl in driving EMT, identify predictive markers of response to Axl inhibition, and investigate the efficacy of combination therapy with an Axl inhibitor and an immune checkpoint inhibitor, anti-PD-L1. The biomarkers identified in their preclinical models will be directly tested in fresh tumor biopsies from their Phase 1/2 clinical trial that combines erlotinib with BGB324.

Although NSCLC is a molecularly heterogeneous disease, EMT is a potential universal mechanism of progression and drug resistance across many molecular subtypes. It is their goal to leverage this finding to develop single agent or combination treatment strategies that can be used across the molecular spectrum.

Circulating tumor cells (CTCs) are cancer cells that break away from either the primary tumor or metastatic sites. Increasingly, CTCs have been used as a readily accessible source of tumor cells (“liquid biopsy”). However, shortcomings in current CTC technologies limit downstream analyses to assessment of a selected set of nucleic acid markers. Genome and transcriptome-wide analyses of CTCs have been hampered by the low purity of CTC isolates. The complex and protracted nature of CTC capture protocols further impedes analysis.

Dr. Kulkarni and his group have developed a CTC capture technology known as Vortex Chip, which allows for rapid isolation of highly purified CTCs. In addition to standard immunofluorescence, they have generated proof-of-principle evidence that their capture technology allows for analysis of genetic material. Moreover, because the Vortex Chip isolates CTCs based on their size, their chip does not require antibody-based capture that can potentially bias isolation of CTC subpopulations. Serial CTC analyses performed in the context of treatment changes can characterize the mutational landscape of tumors as they evolve in response to therapy and thereby identify molecular predictors and drivers of treatment resistance and sensitivity to novel anti-tumor therapies.

Their project has two goals: 1) validate use of CTCs as biomarker in lung cancer and 2) explore use of CTC analysis to obtain genetic information about the tumor. They hypothesize that the unique microfluidic characteristics of lung CTCs will drive rapid isolation of pure CTC populations for analysis to characterize heterogeneity and profile therapeutic responsiveness.