Abstract
The molecular basis for ultraviolet (UV) light-induced nonmelanoma and melanoma skin cancers centers on cumulative genomic instability caused by inefficient DNA repair of dipyrimidine photoproducts. Inefficient DNA repair and subsequent translesion replication past these DNA lesions generate distinct molecular signatures of tandem CC to TT and C to T transitions at dipyrimidine sites. Since previous efforts to develop experimental strategies to enhance the repair capacity of basal keratinocytes have been limited, we have engineered the N-terminally truncated form (Δ228) UV endonuclease (UVDE) from Schizosaccharomyces pombe to include a TAT cell-penetrating peptide sequence with or without a nuclear localization signal (NLS): UVDE-TAT and UVDE-NLS-TAT. Further, a NLS was engineered onto a pyrimidine dimer glycosylase from Paramecium bursaria chlorella virus-1 (cv-pdg-NLS). Purified enzymes were encapsulated into liposomes and topically delivered to the dorsal surface of SKH1 hairless mice in a UVB-induced carcinogenesis study. Total tumor burden was significantly reduced in mice receiving either UVDE-TAT or UVDE-NLS-TAT versus control empty liposomes and time to death was significantly reduced with the UVDE-NLS-TAT. These data suggest that efficient delivery of exogenous enzymes for the initiation of repair of UVB-induced DNA damage may protect from UVB induction of squamous and basal cell carcinomas.
Highlights
IntroductionHumans possess only one mechanism, the nucleotide excision repair (NER) pathway, for repairing dipyrimidine DNA lesions
Since mammalian cells exclusively use nucleotide excision repair (NER) to initiate the repair of dipyrimidine photoproducts, the focus of this investigation was to determine if the induction of UVB-induced NMSCs could be significantly reduced relative to controls through the delivery of an enzyme that repairs both cyclobutane pyrimidine dimers (CPDs) and 6–4 photoproducts (6–4 PPs)
The temporal hierarchy of NER of UV-induced dipyrimidine photoproducts in mammalian cells is characterized by rapid recognition and excision of 6–4 PPs, and preferential repair of CPDs in actively transcribed genes[22,23,24]
Summary
Humans possess only one mechanism, the nucleotide excision repair (NER) pathway, for repairing dipyrimidine DNA lesions. The importance of this DNA repair system in limiting NMSC is best demonstrated by the clinical sequela of patients suffering from the autosomal recessive genetic disorder Xeroderma Pigmentosum (XP). These patients have defects in either NER or DNA translesion synthesis (TLS) of CPDs, either of which will significantly increase the risk for development of NMSC. One strategy for enhancing repair of CPDs in human skin cells has been to deliver, or express, the bacteriophage T4 pdg in these cells[12]. Results of clinical trials with XP patients using topically delivered T4-pdg showed that new pre-cancerous lesions (actinic keratosis) were reduced by 68% and new cases of BCC were reduced by 30% compared to patients treated with placebo lotion[17]
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