The incidence of melanoma has been on the rise over the last 30 years with the 5 year survival rate approaching 2% for patients with metastatic melanoma; veterans are up to 4.5 times more likely to develop melanoma than the general population. These dismal statistics are partly due to the fact that melanoma is notoriously resistant to cancer therapies. We reported that melanoma cell lines and patient samples express elevated levels of the IL-1 receptor-associated kinase-4 (IRAK-4) and IRAK-1. IRAK-4 plays a central role in Toll-like receptor, IL-1, IL-18 and IL-33 signaling. Attenuating IRAK-1,-4 signaling in melanoma cells with pharmacological inhibitors or siRNA diminished the production of various chemokines/cytokines known to promote angiogenesis and the induction of tumor-associated fibroblasts. IRAK inhibition also enhanced the effects of certain chemotherapies in vitro and in tumor-bearing mice. In contrast, the activation of IRAK-4 signaling in melanoma cells increased the production of these cytokines/chemokines and promoted chemotherapeutic resistance. Supernatant from tumor cells engineered to overexpress the IRAK-4 gene augmented endothelial cell proliferation and activated human fibroblasts, as demonstrated by increased expression of Tenascin-C and -SMA. Interestingly, while IRAK-4 is known to localize in the cytosol, we observed that in melanoma IRAK-4 translocates to the nucleus and this is increased upon chemotherapy exposure. The overarching hypothesis is that IRAK-4 nuclear mobilization promotes chemoresistance and that IRAK signaling orchestrates changes in the tumor microenvironment that promotes tumor growth and metastases. The proposed studies are focused on gaining greater mechanistic insights (molecular and cellular) on how IRAK-4 signaling in tumor cells: (1) influences angiogenesis, chemoresistance and metastases, (2) changes the tumor microenvironment, and (3) can be used as a therapeutic target to enhance the efficacy of chemotherapies.
Aim 1 is to gain a mechanistic understanding regarding the role that nuclear IRAK-4 plays in chemotherapy resistance and gene expression. Determining the function of nuclear IRAK-4 is novel and independent of known mechanisms of IRAK-4-mediated activation. This will be investigated using melanoma lines engineered to express IRAK-4 variants including wild type, shRNA-IRAKs, kinase dead IRAKs, and IRAK- 4 SUMO, in which SUMOylation sites have been mutated to increase cytosolic levels but prevent nuclear mobilization.
Aim 2 is to determine the effect that inhibiting IRAK-4 in melanoma has on metastasis and angiogenesis. We will use an inducible model of melanoma in mice (BRAFV600PTEN mice) as well as human and mouse melanoma cells engineered to overexpress or knockdown IRAK-4 to monitor tumor neovascularization and metastases using in vivo and ex vivo models.
Aim 3 is to discover the role that IRAK-4 signaling in melanoma plays in altering the tumor microenvironment to promote tumor growth. Building on preliminary data showing that IRAK-4 signaling in melanoma induces the expression of factors that support tumor associated fibroblasts, this aim will explore the cellular mechanism and investigate the previously unrecognized concept that IRAK signaling within tumor cells is critical for orchestrating the generation of a microenvironment favoring tumor growth and metastasis.
Aim 4 seeks to determine the therapeutic efficacy of targeted delivery of IRAK inhibitor to tumors when combined with chemotherapies. We will use anti-PD-L1 antibodies to deliver FDA-approved chemotherapies plus IRAK-1,-4 inhibitor (currently in Phase II/III clinical trials). The results from these studies will provide a greater molecular and cellular understanding of how IRAK-4 signaling in melanoma enhances chemoresistance and metastasis and will identify new therapeutic targets to enhance chemotherapeutic responses.

Public Health Relevance

US veterans between ages 55-59, 50-54, and 45-49 are 450%, 17%, and 6% more likely to develop melanoma than the general US population. The survival rate of patients with advanced melanoma at 5 years is less than 5%. Melanoma is notoriously resistant to all existing forms of cancer therapies. We have discovered a protein named IL-1 receptor associated kinase-4 (IRAK-4) that is highly expressed in melanoma that contributes to the production of factors that promote tumor growth by increasing the flow of nutrients to the cancer cell and helps melanoma resist chemotherapies. We will test the idea that blocking IRAK-4 in melanoma cells will block tumor growth, prevent metastases and enhance the effects of certain chemotherapies in tumor-bearing mice and in tissue culture experiments.

Agency
National Institute of Health (NIH)
Institute
Veterans Affairs (VA)
Type
Non-HHS Research Projects (I01)
Project #
1I01BX002142-01A1
Application #
8735412
Study Section
Oncology A (ONCA)
Project Start
2015-01-01
Project End
2018-12-31
Budget Start
2015-01-01
Budget End
2015-12-31
Support Year
1
Fiscal Year
2015
Total Cost
Indirect Cost
Name
Baltimore VA Medical Center
Department
Type
DUNS #
796532609
City
Baltimore
State
MD
Country
United States
Zip Code
21201
Kaczanowska, Sabina; Joseph, Ann Mary; Guo, Jitao et al. (2017) A Synthetic CD8?:MyD88 Coreceptor Enhances CD8+ T-cell Responses to Weakly Immunogenic and Lowly Expressed Tumor Antigens. Cancer Res 77:7049-7058
Tsai, Alexander K; Khan, Asra Y; Worgo, Christina E et al. (2017) A Multikinase and DNA-PK Inhibitor Combination Immunomodulates Melanomas, Suppresses Tumor Progression, and Enhances Immunotherapies. Cancer Immunol Res 5:790-803