Date of Graduation

5-2014

Document Type

Dissertation (PhD)

Program Affiliation

Biomedical Sciences

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

Dr. Marsha L. Frazier

Committee Member

Dr. Christopher Amos

Committee Member

Dr. Jeffrey Lee

Committee Member

Dr. Elizabeth Grimm

Committee Member

Dr. Jeffrey Morris

Abstract

Cutaneous malignant melanoma (CMM) is a potentially lethal malignancy that warrants attention and further research, as it is known to that there is an increasing rate of incidence in theUnited States, and it is also known that exposure to UV light is its most crucial risk factor, and family history of melanoma is also an important risk factor. Melanoma is an aggressive and lethal cancer in humans. There are an estimated new 132,000 melanoma cases annually worldwide, and the trend has doubled in the past 20 years. However, attempts to treat melanoma have encountered considerable resistance and remained ineffective. The 5-year survival rate for metastatic melanoma remains less than 5%. CMM patients may develop an immune response to their tumors, but innate anti-tumor immune responses are insufficient for controlling the development of the tumor. Melanoma is a very immunogenic tumor, sometimes exhibiting spontaneous remissions, making it one of the foremost targets for immunotherapy. Unfortunately, despite the attempts at making tumor-infiltrating lymphocyte treatments, the clinical response has been borderline. The immunosuppressive microenvironment of tumor cells becomes significant and intertwines with the resistance of tumor treatment. Furthermore, IL-1 can hinder the immune response to melanoma. The pathway of MAPK activation leads to the production interleukin(IL)-1α/β. IL-1 conducts immunomodulatory activity through tumor-associated fibroblasts and locks in the turnkey position of the immunosuppression pathway to resist the cytotoxic T lymphocyte function. Hence, IL-1 is a key target of interest in treating melanoma, along with the entire pathway flowing from it. Polymorphisms in genes regulating the immune response could result in increased susceptibility to and/or poorer prognosis in certain individuals. For this study, one of the objectives was to examine if single nucleotide polymorphisms (SNPs) of certain pro- and anti-inflammatory cytokines and growth factors, namely IL-1, IL-6, IL-8, IFN-γ, and TNF-α are associated with melanoma outcome of death, recurrence, or composite, and thus susceptibility. Those genes are the upstream and downstream targets for the greater IL-1 pathway. The greater IL-1 pathway has multiple genes in between those upstream and downstream targets. Those genes are IL-1RI, IL-1RAcP, IL-1RA, MYD88, TOLLIP, IRAKs, MEKK1, MEK3, MEK6, JNK, P38, c-JUN, ECSIT, TRAF6, TAB1, TAK1, RKIP, NIK, IKKα, IKKβ, IκBα, and NF-κB.

The individual SNP analysis proved to be interesting though inconclusive. There were some borderline significant results, such as associations of the SNPs rs16944, rs1143627, rs1071676, and rs3136558 with the endpoint of death. The French validation verified a significant association of rs3136558 with death but none of the others.

The next objective was to perform a full pathway analysis, incorporating all available SNPs for each gene in the overall IL-1 pathway to determine if any components were significant and if so, to attempt further verification at the protein level if the data were available. By using the SKAT program for pathway analysis, several genes in the IL-1 pathway were found to be significant. They were IL-1, c-JUN, and ECSIT, with borderline significance for TAK1.

With the observation of associations of these four genes with melanoma outcomes at the DNA level, the next step was to determine if they were also significantly associated with melanoma outcomes at the expression level. The data for the DNA level studies did not include protein expression studies, so we used data provided by The Cancer Genome Atlas (TCGA). TAK1 data was not available, but RNA expression data were gathered for IL-1, c-JUN, and ECSIT. At the protein level, only c-JUN data was available. At the RNA level, IL1 showed that survival was proportional to IL-1 level, ECSIT showed lower survival for higher level, while c-JUN showed higher survival for higher levels. At the protein level, c-JUN was significant at the protein level, both adjusted and unadjusted via logistic regression. It seems that for developing further therapy against melanoma, c-JUN may be be a crucial target in the IL-1 pathway. IL-1 and ECSIT could also play important roles related to melanoma outcomes but require future studies where protein expression data is available to confirm the DNA- and RNA-based results.

Keywords

melanoma, SNP, gene association analysis, pathway analysis, protein expression, IL-1, cJUN, bioinformatics