Mapping DNA Repair and Error-Prone DNA Synthesis in Geriatric Skin

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PROJECT SUMMARY/ABSTRACT Non-melanoma skin cancers (NMSCs) are a growing problem among our nation’s Veteran population, and current treatments are both costly and potentially disfiguring. The primary risk factors for NMSC development are exposure to ultraviolet B (UVB) wavelengths of sunlight and advanced age. Though the mutagenic effects of UVB-induced bipyrimidine dimers in genomic DNA are well known, the mechanisms by which age impacts UVB- dependent NMSC initiation in skin keratinocytes are less clear. The unique structural and physiological properties of geriatric skin likely influence both the number and distribution of photolesions that form across the genome of basal keratinocytes after UV exposure and the ways in which these cells respond to the damage. Moreover, aged tissues including skin are known to contain a higher number of senescent dermal fibroblasts than younger skin, which may alter cytokine and growth factor production and contribute to a pro-tumorigenic microenvironment. One such factor that is expressed at lower levels in geriatric skin is insulin-like growth factor 1 (IGF-1). Our published and preliminary data show that the loss of IGF-1/IGF-1 receptor (IGF-1R) signaling in epidermal keratinocytes results in UVB-irradiated keratinocytes that are unable to efficiently remove UVB photoproducts from DNA or to properly activate the apical DNA damage response kinase ATR, which functions to suppress error-prone DNA synthesis on damaged DNA templates. The objective of this proposal is to determine how age and specifically IGF-1 receptor activation affect the formation and replication of UVB DNA photoproducts in basal epidermal keratinocytes in human skin. The background of the research team, which includes expertise in DNA repair, replication stress response signaling, genomics, keratinocyte biology, and clinical dermatology, makes this group uniquely qualified to address this important clinical issue. The central hypothesis of this proposal is that the skin of geriatric individuals is prone to carry out a more mutagenic form of DNA synthesis following UVB exposure than the skin of younger individuals. This hypothesis has been derived from significant preliminary and published data with both cultured keratinocytes in vitro and human skin biopsies ex vivo and in vivo. The rationale for this proposed research is that it will define how the unique features of geriatric skin predispose it to UVB mutagenesis and carcinogenesis. Our hypothesis will be tested by carrying out the following three specific aims: Aim 1) Define how age and IGF-1 status influence UVB-induced DNA photoproduct formation and repair across the keratinocyte genome in human skin epidermis; Aim 2) Determine how IGF-1 signaling impacts the utilization of error-prone DNA polymerases in UVB-irradiated human skin explants ex vivo and cultured keratinocytes in vitro; and Aim 3) Define how subject age and IGF-1 status affect the replication of UVB-damaged DNA in human skin epidermis in vivo. Our approach is innovative because it will provide a detailed, quantitative characterization of DNA damage formation and replication in skin epidermis using novel technical approaches that have not previously been applied to human skin. The proposed research is significant to the VA because it will show how the geriatric skin microenvironment contributes to UV mutagenesis and carcinogenesis. Ultimately, we expect that the knowledge that will be gained here will provide further mechanistic rationale for the use of clinical skin rejuvenation interventions aimed at preventing NMSC in specific patient populations. Lastly, because many other cancers are associated with both advanced age and the exposure of epithelial tissues to environmental and dietary DNA damaging agents, our work likely has implications for understanding the pathogenesis of lung, colon, and esophageal cancers in older veterans.