DNA Repair, p53 and Apoptosis Phenotypes in Lung Cancer
Condition: Lung Neoplasms
MeSH terms: Lung Neoplasms
Study type: Observational

Summary: 
      The Laboratory of Human Carcinogenesis and the Pharmocogenetics Section of the Genetic
      Epidemiology Branch will conduct a lung cancer case-control study in Baltimore, Maryland. The
      primary hypothesis of the study is to determine if mutagen sensitivity, p53 induction and
      apoptosis in cultured lymphocytes will be predictive of lung cancer risk. While there are
      some studies that examine mutagen sensitivity, none of these assays has been well-studied in
      an epidemiological setting. Because of methodological issues described herein, and the
      proposed development of new assays, this study will be viewed as a pilot and therefore
      hypotheses generating. The design of this molecular epidemiology study has been specifically
      developed to test the reliability and validity of the mutagen sensitivity assay, where a
      case-control study is needed to assess the possibility of case bias (i.e., results vary due
      to the concurrent presence of lung cancer rather than risk). Importantly, this protocol will
      establish a resource that will allow for the validation of these assays and also for the
      study of other biomarkers and gene-environment interactions, especially those related to DNA
      repair. Th secondary goals of this study are to 1) demonstrate gene-neuro-behavioral
      interactions for smoking addiction in controls and 2) assess the relationship of sex-steroid
      metabolism an and estrogen exposure to lung cancer risk. Cases will have histologically
      confirmed lung cancer and reside in Baltimore an and surrounding areas. They will be
      identified through six hospitals in Baltimore. Cases will be recently diagnosed and blood
      will be collected prior to chemotherapy or radiation therapy. Because of this requirement to
      obtain samples before treatment (or for surgical cases at least two months after surgery), we
      recognize that case ascertainment will be reduced, but critical data to assess differences
      between eligible and ineligible subjects will be collected through tumor registries. Two
      control groups will be used, the first will be hospital-based (frequency matched by age,
      gender, race, smoking and hospital) and the second will be population-based (frequency
      matched by age-, gender and race). The first control group will allow us to examine risk
      factors for lung cancer independent of smoking (odds ration for smoking = 1.0), and the
      second will allow the results to be extrapolated to the general population and also will be
      used to validate the phenotyping assays. The strategy for recruitment will allow us to
      over-sample for women and African Americans, so that after examination of data for the entire
      study group, we can assess differences by these subgroups. Cases and controls will receive a
      structured, in person interview assessing prior medical and cancer history, tobacco use,
      alcohol use, current medications, occupational history, family medical history, menstrual
      history and estrogen use, recent nutritional supplements and caffeine intake, and
      socioeconomic status. The questionnaire also will include the Fagerstrom index for nicotine
      dependence (FTND), Center for Epidemiologic Studies Depression (CES-D) scale, and a modified
      version of the Horn-Waingrow Reasons for Smoking (RFS) Scale. The phenotypic markers to be
      studied will assess DNA repair with cellular response by using lymphocyte cultures exposed in
      vitro to radiation, bleomycin, benzo(a)pyrene-diol-expoxide and N-methyl-nitrosurea and then
      measuring induction of chromosomal aberrations, p53 induction and apoptosis. DNA from cases
      and controls also will be used for genetic polymorphism analysis of carcinogen metabolism,
      and those relating to the dopaminergic system and nicotinic receptors. Tumors from cases will
      be evaluated for estrogen and progesterone receptors. The target accrual number of total
      subjects will be 1,200 where there will be 100 cases for each combination of gender and race
      (Caucasian- and African Americans), matched to 100 each of the hospital-based and
      population-based controls.
    

Detailed description: 
      Background:

      The Laboratory of Human Carcinogenesis is conducting an observational non-small cell lung
      cancer (NSCLC) case-control study in Baltimore, MD. This molecular epidemiology study was
      developed to test the reliability and validity of the mutagen sensitivity assay, where a
      case-control study is needed to assess the possibility of case bias. Importantly, this
      protocol establishes a resource that allows for the study of additional biomarkers and
      gene-environment interactions. Upon recruitment, cases and controls receive a structured, in
      person interview assessing prior medical and cancer history, use of tobacco and electronic
      cigarettes, alcohol use, current medications, occupational history, family medical history,
      menstrual history and estrogen use, recent nutritional supplements and caffeine intake, and
      socioeconomic status. Specimen collection consists of a one-time blood sample and/or
      mouthwash to collect cheek cells (oral cells) and a one-time urine sample. In addition,
      cancer and surrounding non-cancer tissue that was surgically removed and not needed for
      diagnosis may be obtained for cases, as well as current medical information from medical
      records. Primary cell cultures may be established from available fresh tumor tissue. The
      phenotypic markers to be studied will assess proficiency of DNA repair in lymphocyte cultures
      exposed in vitro to radiation, bleomycin, benzo(a)pyrene-diol-epoxide by measuring induction
      of chromosomal aberrations, p53 and apoptosis. DNA from buffy coats or cheek cells will be
      used for analysis of germline variation in the form of Single Nucleotide Polymorphisms (SNPs)
      in genes involved in DNA repair, innate immunity, cell cycle control, angiogenesis,
      apoptosis, cytokines, nicotine addiction, inflammation, hormone metabolism and microRNA.
      Additionally, IRB approval was received in 2010 to include this study in a multi-institution
      genome-wide association study (GWAS) of lung cancer in African Americans. Tumors from cases
      will be evaluated for estrogen and progesterone receptors, somatic mutations, and gene
      expression. Urine, plasma, serum and tissue sample metabolomics will be analyzed by
      untargeted approach.

      Objectives:

        1. To determine if mutagen sensitivity, p53 induction, and apoptosis in cultured
           lymphocytes are predictive of lung cancer risk.

        2. To determine the relationship between sex-steroid metabolism, estrogen exposure, and
           lung cancer risk.

        3. To investigate and develop phenotypic or predictive markers of lung cancer risk and
           survival, based on mutagen sensitivity, polymorphic markers, gene expression, and
           metabolomics.

        4. To investigate racial disparities associated with lung cancer risk and survival.

        5. To examine the relationship between circulating cytokines with risk and survival of lung
           cancer and to establish the most robust method of cytokine detection.

        6. To generate a more accurate measure of ancestry using ancestry informative marker
           analysis and to integrate this variable into our studies of health disparities

        7. To conduct studies of metabolomics on serum, plasma and urine for the purposes of
           discovering novel markers of risk, diagnosis and prognosis. We will use ultraperformance
           liquid chromatography coupled to mass spectrometry (UPLS-MS) to search for small
           molecular weight endogenous metabolites that can classify cancer and predict outcome.

           This is a novel approach for biomarker discovery that also leverages the non-invasive
           process of biospecimen collection. Tumor and corresponding non tumor tissues from
           corresponding patients will also be tested using the same methods to extend the
           discovery of novel tumor metabolites. Further, metabolites of vitamin D will be examined
           on serum samples from lung cancer cases and controls to assess the relationship between
           circulating

           levels of Vitamin D metabolites with cancer risk and survival. This analysis will be
           coupled with testing of Vitamin D pathway SNPs in corresponding patients to determine if
           certain SNPs are also associated with levels of vitamin D.

        8. To evaluate biomarkers of cancer diagnosis and prognosis in circulating tumor DNA

        9. To evaluate the microbiome (microbes) present in lung tissue using in situ hybridization
           of fixed tissues to be completed at Mayo Clinic by collaborators. The collaborators will
           receive no information on the samples other than their tissue of origin.

       10. To collect data and biospecimens on patients that received low dose CT screening as part
           of their lung cancer diagnosis. This is for the purposes of investigating non-invasive
           biomarkers of lung cancer diagnosis and prognosis. Low-dose CT screening is now offered
           at UMMS and the VA for the purposes of early lung cancer detection. These screening
           guidelines are per CMS guidelines, i.e., age 55-77, >=30 pack-years of smoking and have
           quit smoking for 15 years or less. Such high-risk patients will be offered annual
           screening.

       11. To culture lung cancer specific microbiome-bacteria from human lung cancers. This work
           will, in part, be conducted with our collaborator Dr Paul Owrin, who is an expert in the
           culture of microbial species from human tissues. These samples, approximately 10 per
           year, will be de-identified of all patient information.

       12. To perform RNA analysis of tumor and normal tissues in collaboration with Dr Isidore
           Rigoustos with the aim of discovering novel signaling pathways in lung cancer.

       13. To contribute our data to larger consortia so that we can conduct studies with a higher
           power and to make valuable, reproducible observations regarding the epidemiology of lung
           cancer. We will establish this data transfer via a DTA.

       14. To conduct next generation DNA sequencing of tumors (and corresponding normal) of
           patients enrolled in this protocol.

       15. To collaborate with intramural colleagues, specifically Dr Jung Byun at NIMHD on the
           genomic analysis (next generation DNA sequencing) of samples within this contract. We
           anticipate sending up to 400 samples. No personal identifiable information will be sent
           or included with these samples.

       16. To collaborate with extramural colleagues, specifically Dr John Simmons at Personal
           Genome Diagnostics, and Dr Peter Campbell at the Welcome Sanger Institute, England, on
           the genomic analysis (next generation DNA sequencing) of samples within this contract.
           We anticipate sending up to 400 samples. No personal identifiable information will be
           sent or included with these samples.

       17. To complete MTAs with colleagues outlined in point 16 above.

       18. To capture exposures related to reproductive history in both men and women, among both
           cases and controls and related these exposures to cancer risk and health disparity.

       19. To use data collected through the questionnaire to geocode participants.

      Eligibility:

        -  Histologically confirmed NSCLC diagnosed within the past 2 years (case).

        -  Frequency matched to cases according to age (5-year intervals), gender, and race
           (population-based control).

        -  Born in the United States, resident of the state of Maryland

        -  Subject Characteristics:

             -  Speaks English well enough to be interviewed

             -  Physically and mentally capable of performing the interview (i.e., must be able to
                hear the interviewer, mentally comprehend the interviewers questions, and verbally
                respond)

             -  Has never been interviewed as a control for this study

             -  Does not currently reside in an institution such as a prison, nursing home, or
                shelter

             -  No history of cancer other than non-melanoma skin cancer or carcinoma in situ of
                the cervix (population-based control)

             -  Has a residential working phone within the home (population-based control)

      Design:

        -  Case/Control; Observational

        -  Planned statistical analysis: Risk associations between the genotypes and
           cancer/survival will be assessed using unconditional logistic regression model, with
           covariate adjustment, as appropriate.

        -  Number of subjects to be enrolled: Target accrual is 6000 subjects, consisting of 450
           cases/each gender in African Americans and 800 cases/each gender in Caucasians. An equal
           number of controls will be selected for each category of cases based on the combination
           of gender and race.

        -  For patients that undergo low dose CT screening at the participating hospitals in this
           protocol, we will enroll patients that have had a "positive" scan and are attending
           either UMMS or the VA hospital for further follow up.
    

Observational model: Case-Control
Time perspective: Cross-Sectional
Sampling method: Non-Probability Sample
Study population: 
        The population controls are collected from DMV records in the Baltimore region of MD. The
        cases are patients at the University of Maryland Medical System, including the associated
        Veterans' Association Hospital.@@@
      
Keywords: Molecular Epidemiology of Lung Cancer; Health Disparities