Funded Projects

Side effects of immune checkpoint therapies, termed immune-related adverse effects (irAEs), are unique and generally secondary to inflammatory tissue damage. The onset and duration of irAEs are unpredictable and predisposing factors for individuals to develop irAEs remains unclear. Some irAEs that emerge with immune checkpoint blockade share clinical features of autoimmune conditions. Preexisting auto-reactive antibodies and their contribution to immune side effects with immunotherapy and radiation have not been defined. This maybe particularly relevant to immune related adverse effects, since recent studies suggest almost 8-9% of the US population carries autoimmune diseases and 26% of healthy individuals have strong IgG humoral responses to variety of self-antigens. For the proposed study, we will use a set of longitudinally collected patient blood samples from a clinical trial which evaluates the clinical activity of combination immune checkpoint inhibitor therapy, nivolumab and ipilimumab with or without local consolidative therapy (stereotactic body irradiation) 

We plan to follow preexisting auto-reactive antibodies, baseline T and B cell receptor clonality and their temporal dynamics for combination immune checkpoint therapy with or without radiation therapy and correlate these findings with irAEs and response to therapy.    

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.

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.

Statement of the Problem: Cancer develops in a sequenced manner. Patches of lung cells gain the ability to multiply faster than their neighboring normal cells by acquiring mutations. These patches of cells are called “premalignant lesions" (PMLs). Some of these lesions may eventually become cancer. Knowledge about how some PMLs may develop into lung cancer is central to developing tools for Cancer Interception—interrupting the cancer development process and blocking progression to advanced metastatic disease. However, we lack effective lung cancer interception approaches due to our incomplete understanding of the earliest molecular events associated with the development of lung cancer, as well as the challenge in developing personalized tools for early detection and prevention.

Proposed Solution: The Dream Team will use a holistic approach to understand the genetics, immunology, radiological (imaging) profile, and treatment response of patients who show evidence of abnormal lung tissue that puts them at high risk to develop lung cancer. The Team has access to unique groups of patient populations (adenocarcinoma—ADC and squamous cell carcinoma—SCC) with signs of lung cancer risk, and has assembled a collection of blood and tissue specimens from these individuals. The main objective of the Team is to understand how lung cancer develops and test methods that will block the development of cancer. Their three-pronged approach to solve this problem will include:

• Creation of a molecular pre-cancer genome atlas (PCGA) of the lung with the goal of identifying which types of precancerous lung tissue require aggressive treatment and which treatments will block the development of these abnormal lesions to invasive lung cancer.
• Development of two diagnostic tools that can be directly applied in the clinic for simple, yet accurate, detection of early lung cancer: 1) the use of nasal swabs, blood tests, and imaging to confirm whether lung abnormalities found on chest imaging are lung cancer or benign lung disease. 2) blood tests that can identify patients at the earliest stages of lung cancer recurrence, thus enabling their timely and effective intervention with therapies such as those that boost the immune system.
• Development of tests to identify which individuals are most likely to benefit from a number of treatment strategies to intercept lung cancer, including emerging immunotherapies.

Statement of the Problem: Currently, low dose CT (LDCT) screening for lung cancer is recommended for high-risk individuals, defined as current or former smokers (quit within the past 15 years) with a 30-or-more pack-year smoking history, 55 to 74 years of age, and with no evidence of lung cancer. However, uptake of LDCT screening has been impeded by high rates of false-positive results, which in turn add significant physical and emotional stress to patients as well as additional healthcare costs. Since LDCT is now commonly available, lung nodules detected in LDCT scans are plentiful, but imprecise guidelines for care make it a challenge to ensure quality follow-up and expedited diagnoses for those with lung cancer. Furthermore, screening is not available for those who don’t meet the screening criteria (younger age or low smoking history) when they are the exact population among whom lung cancer rates are rising.

Proposed Solution: Leveraging expertise of academic research centers (Massachusetts General Hospital, Broad Institute, and Stanford University) and a non-profit managed-care consortium (Kaiser Permanente), the Dream Team will develop a test called the Lung Cancer Interception Assay (LCIA) to complement LDCT screening.

The proposal is novel in that it will develop complex computational algorithms to combine high-sensitivity/high-accuracy blood-based molecular biomarkers with image-based biomarkers to create the  LCIA assay. At the end of the award period, a streamlined product, the LCIA, will be ready to use in population-level definitive trials for the early detection of lung cancer.

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.

Inadequate biopsy yield is a significant limitation to early lung cancer diagnosis and is a two-pronged problem: (1) Early-stage lung cancers are not adequately targeted during biopsy due to their small size and (2) even if a nodule is adequately targeted, often there is insufficient tumor in the biopsy to make a definitive diagnosis and/or accomplish all diagnostic testing for patient care. Optical coherence tomography (OCT) is a high-resolution imaging modality that provides virtual visualization of tissue volumes orders of magnitude larger than biopsy without tissue removal. Dr. Hariri’s group has developed OCT catheters compatible with standard bronchoscopes and demonstrated that OCT can identify lung nodules and assess pathologic features of lung cancer with high sensitivity/specificity. This project’s aims are to dramatically improve diagnostic capability in lung cancer using large volume OCT to provide (1) virtual intra-procedural evaluation in real-time (VIPER) of biopsy site location and 2) large volume virtual optical biopsy for subsequent pathology diagnosis as a complement to tissue biopsy. In Aim 1, they will assess if in vivo OCT VIPER biopsy assessment increases tumor yield on bronchoscopic biopsy in a powered, blinded clinical study. In Aim 2, they will use the data obtained in Aim 1 to determine the diagnostic accuracy of large volume OCT optical biopsy with tissue biopsy as compared to tissue biopsy alone. This project will result in clinical translation of a powerful, robust new optical bronchoscopy tool that could reduce unnecessary diagnostic procedures, eliminate delayed diagnoses, provide earlier intervention, and improve patient morbidity/mortality.

Small cell lung cancer (SCLC) is among the most aggressive and deadly human cancers. It represents approximately 15% of all lung cancers, and it is responsible for over 30,000 deaths in the US and 200,000 deaths worldwide every year. SCLC responds to treatment but recurs frequently and aggressively, with the majority of patients dying within 12 months of diagnosis. No targeted therapies are yet available for SCLC. New strategies are needed to improve outcomes for patients with SCLC. This project will expand upon preliminary data identifying at least two distinct subpopulations of cells in SCLC cell lines and patient-derived xenografts comprising a neuroendocrine (NE) and mesenchymal-like (ML) set of cell line populations. Dr. Lehman and his team will use an innovative approach using mass cytometry, using rare earth antibody conjugates to antibody label more than 30 proteins simultaneously, including phospho and signaling proteins. This technique will establish at a single-cell resolution tumor heterogeneity and signaling in small cell lung carcinoma in multiple primary human samples as well as in patient-derived xenograft samples before and after chemotherapy treatment. This proposal aims to (1) characterize the signaling of subpopulations in human primary SCLCs at a single-cell resolution, (2) identify changes in tumor heterogeneity and in these subpopulations after chemotherapy treatment and relapse, and (3) disrupt and manipulate developmental signaling/ associated subpopulations (prototypically the HH [Hedgehog] and Notch/DLL3 pathways) to reduce tumor burden and lead to targeted rational combination therapy for SCLC.

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.

KRAS mutations activate signaling from downstream effector proteins and are though to be required for tumor formation. Exactly how effector signaling is activated, however, is not fully understood. In addition, whether tumors harboring KRAS mutations depend on this oncogene for growth is unclear.

Dr. Lito and his group will dissect the phenotype of KRAS-mutant lung cancer by using a novel allosteric inhibitor that selectively targets KRAS G12C. They will first utilize biochemical assays in order to determine the mechanism of action of this inhibitor. An emphasis will be placed on evaluating how feedback pathways modulate this process, since this is likely to identify factors that may be used as biomarkers of response to treatment.

They will then determine which effector pathways drive the phenotype of KRAS G12C mutant lung cancer, and whether these exhibit adaptation (or reactivation) over time. If so, they will next explore combination-based interventions in order to establish the most effective antiproliferative strategy.

Finally, they will utilize novel patient-derived xenograft and cell line models, established from patients with lung cancer treated at MSKCC, in order to determine if KRAS G12C mutant lung cancers depend on this oncoprotein for growth. These models will also be used to test the most effective combination strategies determined above, as well as to test the potential benefit of intermittent administration schemes designed to best target tumor adaptation to this novel class of inhibitors.

The grand objective of this project is to establish a mutant KRAS-directed therapy for patients with lung cancer. 

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-associated deaths worldwide. Given the limited effectiveness of current treatments, there is a significant need to identify new driver genes that participate in lung cancer pathogenesis. Dr. O’Donnell and her group identified PROTOCADHERIN 7 (PCDH7) in a functional screen for genes that promote transformation of human bronchial epithelial cells (HBECs). This is of interest because PCDH7 encodes a poorly characterized cell surface receptor that is frequently overexpressed in NSCLC and high expression strongly associates with poor survival of NSCLC patients. Moreover, the presence of PCDH7 on the cell surface highlights the potential of this molecule as a novel target for future therapeutics.

They demonstrated that 1) enforced expression of PCDH7 is sufficient to promote transformation of HBECs, 2) PCDH7 cooperates with oncogenic KRAS to promote tumorigenesis, and 3) PCDH7 potently enhances KRAS-mediated activation of the ERK1/2 pathway. They will elucidate the role of this lung cancer driver by testing the following central hypothesis: Overexpression of PCDH7 initiates a signal transduction cascade that potentiates KRAS-induced MAPK-ERK signaling to promote lung tumorigenesis. In Aim 1, they will correlate PCDH7 immunoexpression with clinical features of resected NSCLCs. In Aim 2, they will elucidate the mechanisms through which PCDH7 potentiates MAPK signaling and tumorigenesis. In Aim 3, they will develop and test the therapeutic efficacy of PCDH7 blocking antibodies in patient-derived xenografts.

These experiments will yield insights into the mechanisms of PCDH7-mediated transformation and reveal new opportunities to pursue this cell-surface receptor as a therapeutic target in NSCLC.

The overarching goal of the project is to develop a new technology for accurate and cost-effective lung cancer screening in current/former smokers. Lung cancer is curable if detected early. However, there are no robust screening techniques with options such as CT scans fraught with cost, harms, and a high false positive rate. Dr. Backman and his group hope to transform the clinical practice of lung cancer screening by exploiting field carcinogenesis, the concept that the genetic/environmental milieu that results in lung tumor impacts upon the entire aerodigestive mucosa including the buccal mucosa, which was referred to as a “molecular mirror” of lung carcinogenesis.

Their data show that the alteration of nanoscale architecture of buccal cells is exquisitely sensitive to lung field carcinogenesis and may serve as a robust biomarker for lung cancer. These nanoarchitectural changes can be detected in a practical and highly accurate fashion via a new technology, partial wave spectroscopic (PWS) microscopy (‘nanocytology’).

In the proposed study, they will conduct a blinded validation case-control trial to determine the sensitivity and specificity of buccal nanocytology for lung cancer. They will evaluate the ability of the test to identify patients who have lung cancer and how many patients with lung cancer would be missed using this technique. They will perform a subgroup analysis for Stage I lung cancers. They will assess the potential confounding effects of smoking frequency and time of cessation and other aerodigestive malignancies.

This project will provide the requisite data for the future definitive multicenter validation trial followed by FDA submission.

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.

Background & Significance: Phase I studies with the anti-programmed death-1 antibody (anti-PD-1), nivolumab, in advanced non-small-cell lung cancer (NSCLC) have demonstrated remarkably durable responses in heavily pretreated patients that lasted a median 17 months. However, in these studies very few patients have had tumors available for immune analysis, and basic data on immune measures pre- and post-treatment are not available. This study will move the treatment paradigm forward by assessing the safety and feasibility of preoperative anti-PD-1 in NSCLC and providing detailed information on the effect of PD-1 modulation on the lung tumor immune microenvironment.

Methods: Patients with newly diagnosed stage IB-IIIA NSCLC will receive two doses of nivolumab 3mg/kg IV on day-28 and day-14 prior to planned surgery on day 0. Serial blood samples for immune markers will be obtained. The primary endpoint of this study is the safety and feasibility of preoperative nivolumab in resectable NSCLC. For the exploratory immune analyses, initial tumor biopsy, post-treatment resection specimen, and serial peripheral blood samples will be evaluated for expression of immune markers with the primary comparison being intra-patient, pre- and post-treatment.

Experimental Approach & Statistical Analysis Plan: As this is the first time that nivolumab has been administered pre-operatively in NSCLC, the study will commence with a run-in phase where six patients will be monitored continuously for adverse events through eight weeks following surgery. Overall, the study will continue without pause to a 12-patient expansion phase if none or one of the six patients experiences DLT in the run-in. Detailed statistical plans for the safety and feasibility endpoints and exploratory immune endpoints are described in the main body of this application.

The ALK tyrosine kinase inhibitor (TKI) crizotinib has demonstrated significant activity in patients with ALK+ lung cancer, but its efficacy is limited by the development of acquired resistance. One potential resistance mechanism involves activation of the EGFR and HER2 kinases. This activation occurs in the absence of EGFR/HER2 mutation or amplification, suggesting an alternative mechanism for up-regulation of these kinases. We discovered a novel association between ALK and the EGFR/HER2 negative regulator, MIG-6. MIG-6 is known to inhibit EGFR/HER2 kinase activity through direct binding. However, to date, there is no known link between ALK and MIG-6. In our preliminary data, we demonstrate that MIG-6 protein levels decrease after ALK TKI treatment, a process abrogated by addition of a proteasome inhibitor. Furthermore, MIG-6 transcript and total protein levels are decreased in ALK+/TKI resistant compared to isogenic ALK+/TKI sensitive cells. The decline in MIG-6 correlates with an increase in EGFR and HER2 protein levels in the resistant cells. Based upon these data, we hypothesize that ALK serves as an upstream regulator of MIG-6, and that MIG-6 suppression drives EGFR/HER2 activation in the ALK TKI resistant state. To test this hypothesis, we propose to: 1) elucidate the molecular mechanisms whereby ALK regulates MIG-6 expression, and 2) determine the mechanisms whereby MIG-6 contributes to EGFR/HER2 pathway activation in TKI-resistant ALK+ lung cancer cells. The proposed studies may lead to strategies that augment or restore expression of MIG-6 and therefore will represent a novel therapeutic approach for patients with acquired resistance to ALK inhibitor therapy.