Synthetic rescue of Xeroderma Pigmentosum C phenotype via PIK3C3 downregulation.

Xeroderma Pigmentosum C is a dermal hereditary disease caused by a mutation in the DNA damage recognition protein XPC that belongs to the Nucleotide excision repair pathway. XPC patients display heightened sensitivity to light and an inability to mend DNA damage caused by UV radiation, resulting in the accumulation of lesions that can transform into mutations and eventually lead to cancer. To address this issue, we conducted a screening of siRNAs targeting human kinases, given their involvement in various DNA repair pathways, aiming to restore normal cellular behavior. We introduced this siRNA library into both normal and XPC patient-derived fibroblasts, followed by UVB exposure to induce DNA damage. We assessed the reversal of the XPC phenotype by measuring reduced photosensitivity and enhanced DNA repair. Among the 1292 kinase-targeting siRNAs screened, twenty-eight showed significant improvement in cellular survival compared to cells transfected with non-targeting siRNA after UV exposure in XPC cells. From these candidates, PIK3C3 and LATS1 were identified as particularly effective, promoting over 20% repair of 6-4 photoproduct (6-4PP) DNA lesions. Specifically targeting the autophagy-related protein PIK3C3 alone demonstrated remarkable photoprotective effects in XPC-affected cells, which were validated in primary XPC patient fibroblasts and CRISPR-Cas9 engineered XPC knockout keratinocytes. PIK3C3 knock down in XP-C cells ameliorated in UVB dose response analysis, decreased apoptosis with no effect on proliferation. More importantly, PIK3C3 knock down was found to induce an increase in UVRAG expression, a previously reported cDNA conveying lower photosensitivity in XP-C cells. Thus, attempts to improve the XPC photosensitive and deficient repair phenotype using PIK3C3 inhibitors could pave a way for new therapeutic approaches delaying or preventing tumor initiation.