SPOREs : Organ-Specific SPORE Programs : Lung

Overall Abstract

Principal Investigator: Gerold Bepler

The 5-year survival from lung cancer has kept pace with the improvement in 5-year survival from all cancers over the last 40 years.  Yet, it remains disappointingly low at ~15%.  This improvement in 5-year survival is a result of heightened awareness, better technology for detection, better selection of patients for various therapeutic options, and the selective use of palliative interventions.  However, lung cancer mortality remains extraordinarily high, and it is the benchmark by which future generations will judge our success in effectively combating this disease.  The Moffitt Cancer Center SPORE in Lung Cancer will 1) elucidate mechanisms of action of crucial molecules in lung carcinogenesis and tumor progression and investigate their impact on therapeutic efficacy and 2) prospectively assess the clinical utility of the molecules for therapeutic and prevention interventions.  Our ultimate goal is to change the standard of care for people at risk for and with lung cancer.  The SPORE has four translational research projects, three cores, a developmental research program, and a career development program.

Project 1:
E2Fs Impact on Therapeutic Efficacy

Principal Investigators: Douglas Cress, Jiandong Chen, Gerold Bepler

Background: The majority of drugs currently used for advanced NSCLC induce cell cycle alterations and apoptosis.  Apoptosis induction by these drugs appears dependent upon the E2F1 pathway.  Specifically, we find these drugs induce E2F1 at the protein level and that E2F1 deficiency protects NSCLC cells from death induced by such agents.  However, we have identified SirT1 as a novel negative regulator of E2F1 that restricts E2F1-mediated induction of apoptosis by cytotoxic agents in a negative feedback loop.  Preliminary data suggest that interference of this negative feedback significantly increases the sensitivity of NSCLC lines to drug-induced apoptosis.

Hypothesis: These results suggest the hypothesis that the E2F1/SirT1 pathway is crucial for efficacy of chemotherapeutic agents in NSCLC and provide proof-of-principle that targeting the E2F1/SirT1 pathway will have therapeutic benefit.  Results suggest that E2F1 activates SirT1 and that SirT1, in turn, limits E2F1-induced cell death.  Preliminary data also suggest that the E2F1/E2F4 ratio may play an important role in drug-induced apoptosis.  However, we do not fully understand whether other E2F family members play any significant roles in this process.

Specific Aims: The first specific aim will define the molecular interactions between SirT1 and members of the E2F family in NSCLC and will define the roles of different E2F family members in drug-induced apoptosis.  It will primarily address experimentation in tissue culture using NSCLC lines.  The second specific aim will determine the contribution of the E2F1-SirT1 pathway on clinical outcome and therapeutic efficacy in lung cancer patients in ongoing clinical studies by the Thoracic Oncology Program.  The purpose of this aim will be to understand whether the pathways that we have defined in cell culture are applicable to the patient and whether E2F1, E2F4, or SirT1 will serve as predictive and/or prognostic markers in NSCLC.  The third specific aim will seek to generate and characterize reagents that will disrupt the E2F/SirT1 pathway and define the functional consequences of this disruption in NSCLC chemotherapeutic interventions.  We hypothesize that these agents will result in increased apoptosis in response to chemotherapeutic drugs and could ultimately have significant therapeutic value in NSCLC.

Project 2:
Antitumor Mechanisms of SRC Inhibitors in Lung Cancer

Principal Investigators: Eric Haura, Jin Cheng

Background: SRC proteins can link receptor tyrosine kinases to critical downstream oncogenic pathways such as PI3K/PTEN/Akt, STATs, and Ras/Raf/ERK.  Regulation of these key pathways allows SRC to control cellular growth and proliferation, survival, invasion, and angiogenesis.  Based on the importance of EGFR signaling in lung cancer and the known cooperation between EGFR and SRC proteins, we evaluated the effectiveness of novel SRC inhibitors in lung cancer cell lines with defined EGFR status.  SRC inhibition reduced mutant EGFR lung cancer cell viability through the induction of apoptosis while having no significant apoptotic effect on cell lines with wildtype EGFR.   The induction of apoptosis in EGFR mutant cell lines corresponded to downregulation of activated Akt and Stat3 survival proteins.  In cell lines without EGFR mutation, SRC inhibition reduced cyclin D and increased p27 protein levels with a corresponding G1 cell cycle arrest.  SRC inhibition also inhibited activated FAK and inhibited lung cancer cell invasion.

Hypothesis: Our central hypothesis is that novel SRC inhibitors will be effective therapy for patients with lung cancer, especially those driven by mutant EGFR proteins.  The goal of this project is to further characterize the effect of novel SRC kinase inhibitors in lung cancer cells, tumor xenografts, and patients with advanced NSCLC.

Specific Aims: (1) We will characterize the effect of SRC inhibitors on apoptosis and growth inhibition in lung cancer cells with defined EGFR status.  We will evaluate the effects on key downstream pathways that control apoptosis and cell growth such as PI3K/PTEN/Akt and STATs.  We will evaluate the effect of combined EGFR and SRC tyrosine kinase inhibitors on apoptosis and growth inhibition. SRC activation will be evaluated in human lung cancer specimens. (2) We will evaluate the effect of SRC inhibition on tumor growth in vivo in lung cancer xenograft models with corresponding biomarker analysis.  We plan to test the hypothesis that SRC inhibitor treatment of lung cancer xenografts with EGFR mutation will undergo tumor regression through enhanced apoptosis while treatment of xenografts with wildtype EGFR will result in growth arrest.  Biomarkers of response defined in the above aim will be further validated in these models.  (3) Two investigator-initiated patient-based clinical trial trials are proposed.  (A) A phase I trial of erlotinib and dasatinib in previously treated NSCLC, and (B) a phase II trial of erlotinib &dasatinib in previously treated NSCLC along with biomolecular analysis based on mechanisms defined in Aims 1&2.    

Project 3:
Lung Cancer Chemoprevention with Enzastaurin

Principal Investigators: Gerold Bepler, Mark Alexandrow

Background: Persons that have quit smoking remain at an increased risk for lung cancer for lifetime that is proportional to the amount of cigarettes smoked.  Approximately 25% of the adult U.S. population are current smokers and 40-50% are former smokers.  Nationally, the focus of lung cancer chemoprevention is on inhibitors of inflammatory response pathways and growth factor signaling pathways and the use of intermediate biomarkers as primary trial endpoints.  This development is based on several inter-related theories.  1) Lung cancer arises in epithelial cells of the airways that have a common genetic and environmental exposure profile.  This so-called “field cancerization” theory implies that the entire airway is at risk that may respond to and require systemic intervention.  2) Cancers in the primary and secondary bronchi often have histopathologic precursors, such as metaplasia and dysplasia.  3) Proliferation and evasion of apoptosis are a requirement for carcinogenesis and progression, which is thought to be triggered by growth factor and inflammatory cytokine signaling on a background of genetically altered response capabilities.  4) Since the target population for chemoprevention consists of persons without overt disease, any therapeutic intervention must have minimal toxic effects.

Hypothesis: One novel agent with antiproliferative, proapoptotic, and minimal toxic effects is enzastaurin (LY317615).  It is an oral protein kinase C (PKC) inhibitor currently undergoing clinical efficacy testing in phase II and III trials in patients with a variety of malignancies.  We hypothesize that enzastaurin, administered orally for 6 months to former smokers with bronchial metaplasia or dysplasia, will reduce the proliferative potential of bronchoepithelial cells.  The compound is not FDA approved, however, Eli Lilly & Co has embraced our proposal to investigate its efficacy as a chemopreventive agent.  The proposal has been endorsed by the Division of Cancer Treatment (NCI), and the FDA has given preliminary approval for conduct of the trial under an IND held by Eli Lilly.

Specific Aims: In aim 1, we will test the clinical utility of this agent in lung cancer prevention.  We will conduct a double-blind, placebo-controlled, randomized trial in former smokers.  Participants will be stratified by lung cancer risk based on airway obstruction (present vs. absent), prior history of completely resected stage I non-small cell lung cancer (NSCLC; present vs. absent), and the histopathology of bronchial biopsies (dysplasia present vs. absent).  A surrogate marker of lung cancer risk, the proliferation marker Ki-67, will be used as the primary endpoint.  In aim 2, we will measure inhibition of enzastaurin’s primary target, PKC, and downstream molecules in bronchoepithelial cells and explore the relationship between target inhibition, proliferation, and classes of histopathologically defined airway morphology.  In aim 3, we will determine the ability of enzastaurin, over a 6-month time period, to reduce bronchoepithelial cell proliferation by measuring the expression index of DNA replication origin licensing factors taking chromatin interaction of these factors into account.

Project 4:
p53-Based Vaccine for Small Cell Lung Cancer

Principal Investigators: Dmitry Gabrilovich, Scott Antonia

Background: Small cell lung cancer (SCLC) is the incurable, most aggressive form of lung cancer.  SCLC is frequently associated with somatic mutations in the p53 gene that often results in p53 overexpression.  This overexpression produces a variety of antigenic epitopes that form the basis for tumor specific cellular immunotherapy.  Dependence of tumor cells on abnormal p53 for their survival makes this protein an “ideal” candidate for cancer immunotherapy.  Because of their unique features, dendritic cells (DC) are the best vehicles for delivery of tumor antigens (Ags).  We have developed a new vaccine based on transduction of dendritic cells (DC) with wild-type p53 using an adenoviral construct.  This vaccine demonstrated potency in pre-clinical experiments.  We have completed an initial phase I/II trial that was designed to test the safety and efficacy of the Ad.p53-DC vaccine in patients who have extensive stage SCLC.  Twenty-nine patients have been fully evaluated clinically and immunologically, and another 15 patients are in the process of evaluation.  The vaccine itself was safe, and produced major tumor responses in two patients.  P53-specific T cell responses were induced by the vaccine in half of the treated patients.  These levels of immunological and clinical responses to vaccination were similar to those reported in other clinical trials.  Thus, a substantial proportion of patients did not develop an immunological response to vaccination.  Our data demonstrated that the lack of immune response to vaccination was closely associated with accumulation of immature myeloid cells (ImC), previously shown to be immunosuppressive. However, the main findings from the trial were an unusually high frequency of major objective tumor regressions in patients treated with chemotherapy immediately after the vaccine. These observations were quite unexpected since the existing paradigm suggests that chemotherapy is detrimental to the maintenance of an immune response.  During the last 8 months, four groups including ours nearly simultaneously reported similar observations in different cohorts of patients treated with different vaccines and chemotherapeutics.

Hypothesis: We hypothesize that the combination of immunotherapy and chemotherapy in direct sequence will provide substantial clinical benefits.  All previous trials including ours were not designed to evaluate this paradigm.  We believe that this issue is of a paramount significance for the entire field and deserves definitive testing.  Therefore we propose to test the following hypotheses: (1) the combination of the Ad.p53-DC vaccine and subsequent chemotherapy will result in a substantial improvement in the clinical response, and (2) the addition of all-trans-retinoic acid (ATRA) to the Ad.p53-DC vaccine may substantially improve the p53-specific immune response and hence clinical response in SCLC patients.

Specific Aims: (1) We will determine the clinical response to the Adv-p53 DC vaccine in patients with extensive stage SCLC, whether chemotherapy given after the vaccine is more effective, and whether all-ATRA enhances this response.  Only previously untreated patients or patients during or immediately after first-line chemotherapy (carboplatin and etoposide) for extensive stage SCLC will be eligible.  P53 staining of the tumor tissue will be performed.  All patients enrolled will receive 4 cycles of carboplatin and etoposide as first-line chemotherapy, and will then be restaged.  The expected responses from this are: 10% CR, 58% PR, 20% SD, and 12% PD.  Patients with progressive disease will be taken off protocol. Patients with stable disease or better (88% of the patients enrolled) will be randomized to one of three arms.  Arm A: Observation until evidence of progressive disease (standard of care).  Arm B: Ad.p53-DC vaccines.  Arm C: Ad.p53-DC vaccines in combination with ATRA.  At the time of disease progression, all patients in all 3 arms will be treated with second-line chemotherapy, paclitaxel.  To address the hypothesis that the combination of the Ad.p53-DC vaccine and subsequent chemotherapy will result in a substantial improvement in the clinical response, the clinical response to paclitaxel of patients in Arm B and Arm C will be separately compared to the control arm.  All of the randomized patients will be evaluable for this analysis.  To address the hypothesis that the addition of ATRA to the Ad.p53-DC vaccine may substantially improve p53-specific immune responses and hence clinical responses, the clinical response to vaccine treatment of the patients in Arm B will be compared to those in Arm C.  Because 10% of the patients randomized will have had a CR with first-line chemotherapy, 90% of the patients in Arms B and C will be evaluable for this analysis, i.e. have measurable disease.  (2) We will determine the immune modifying effect of immunization and chemotherapy on p53-specific immunity.  We will evaluate p53-specific immune response using ELISPOT, LYSISPOT and tetramer assays; evaluate association between p53 specific immune response and anti-adenoviral response, and the presence of immature myeloid cells, dendritic cells and regulatory T cells.  These parameters will be correlated with clinical response to vaccination.

Core A:
Tissue Procurement, Pathology, and Bioinformatics

Core Directors: Soner Altiok, David Fenstermacher

Tissue procurement and pathology will provide the scientific and technical expertise in correlative methodologies suitable for validation of molecular discoveries in relevant patient sub-sets.  The core will serve as an integrating mechanism with the biostatistics and clinical trials core, the administrative and patient advocacy core, and shared resources within the Cancer Center.  It will facilitate tissue procurement and distribution in accordance to established institutional guidelines.  Since the Moffitt Cancer Center SPORE in Lung Cancer has a clear translational focus, the core will assist the project leaders in their basic science endeavors using a variety of methodologies available within the shared resources of the Cancer Center.  A significant emphasis will be placed on high throughput technologies like tissue microarray and cancer micro-genomics using laser capture micro dissection.  New methodologies will be validated and incorporated to harvest small quantities of macromolecules (DNA, RNA, and protein) for target molecule analysis.  The primary role of bioinformatics is to provide expertise with database design, computing, programming, web development, and data sharing for the program.

Core B:
Clinical Trials & Biostatistics

Core Directors: Michael Schell, Gerold Bepler

The primary role of the clinical trials and biostatistics core is to provide expertise with all aspects of study design, data management, data analysis, and data reporting for all investigators.  Clinical trials data will be entered into ONCORE, the clinical trials database used across the Cancer Center.  It will provide the logistics necessary for development and support of clinical trials associated with this SPORE.  Specifically, this core will provide analytical and statistical support including the design, analysis, interpretation, and reporting of results for all projects.

Core C:
Administration & Patient Advocacy

Core Directors: Gerold Bepler, Douglas Cress, Gwen Quinn

The administrative core will provide organizational and financial oversight of all projects and cores.  This resource will assure that all administrative requirements for a successful completion of the overall SPORE goals are met.  It will integrate patient advocacy as a vital component for laboratory and clinical lung cancer research.

Developmental Research Program:

Program Directors: Gerold Bepler, Douglas Cress

The developmental research program will provide guidance, advise, evaluation, and funding for up to four pilot projects per year.  The goal for each pilot project is to generate sufficient high-quality preliminary data relevant in the field of lung cancer prevention, treatment, basic research, or applied research to allow for submission of an independent project to the NIH or other federal agency for funding.

Career Development Program:

Program Directors: Eric Haura, Douglas Cress

The career development program will provide guidance, advice, evaluation, and financial support for up to two faculty level persons per year.  The goal is to allow motivated individuals to develop an independent research career in translational science focused on lung cancer.

List of Investigators:

Publications Related to the SPORE Projects

(click on investigators name for listing of publications)

E2Fs impact on therapeutic efficacy: 
Doug Cress
Jiandong Chen
Gerold Bepler

Antitumor mechanism of src inhibitors: 
Eric Haura
Jin Cheng

Chemoprevention with enzastaurin: 
Gerold Bepler
Mark Alexandrow

P53-based vaccine for small cell lung cancer: 
Dmitry Gabrilovich
Scott Antonia

Currently Funded Developmental Projects

(click on investigators name for contact information)

Novel mechanism of tumor escape in cancer
Dmitry Gabrilovich

Role of Stat3 and Id transcription factors in Non-Small Cell Lung Cancer
Srikumar Chellappan 

Targeting the AKT Pathway in Lung Cancer
Jin Cheng

BRCA1 status as a marker of clinical outcome in Lung Cancer
Alvaro Monteiro

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