Loss of Uracil DNA Glycosylase Selectively Resensitizes p53-Mutant and -Deficient Cells to 5-FdU

Abstract

Thymidylate synthase (TS) inhibitors including fluoropyrimidines [e.g., 5-Fluorouracil (5-FU) and 5-Fluorodeoxyuridine (5-FdU, floxuridine)] and antifolates (e.g., pemetrexed) are widely used against solid tumors. Previously, we reported that shRNA-mediated knockdown (KD) of uracil DNA glycosylase (UDG) sensitized cancer cells to 5-FdU. Because p53 has also been shown as a critical determinant of the sensitivity to TS inhibitors, we further interrogated 5-FdU cytotoxicity after UDG depletion with regard to p53 status. By analyzing a panel of human cancer cells with known p53 status, it was determined that p53-mutated or -deficient cells are highly resistant to 5-FdU. UDG depletion resensitizes 5-FdU in p53-mutant and -deficient cells, whereas p53 wild-type (WT) cells are not affected under similar conditions. Utilizing paired HCT116 p53 WT and p53 knockout (KO) cells, it was shown that loss of p53 improves cell survival after 5-FdU, and UDG depletion only significantly sensitizes p53 KO cells. This sensitization can also be recapitulated by UDG depletion in cells with p53 KD by shRNAs. In addition, sensitization is also observed with pemetrexed in p53 KO cells, but not with 5-FU, most likely due to RNA incorporation. Importantly, in p53 WT cells, the apoptosis pathway induced by 5-FdU is activated independent of UDG status. However, in p53 KO cells, apoptosis is compromised in UDG-expressing cells, but dramatically elevated in UDG-depleted cells. Collectively, these results provide evidence that loss of UDG catalyzes significant cell death signals only in cancer cells mutant or deficient in p53. This study reveals that UDG depletion restores sensitivity to TS inhibitors and has chemotherapeutic potential in the context of mutant or deficient p53. .

HDACi mediate UNG2 depletion, dysregulated genomic uracil and altered expression of oncoproteins and tumor suppressors in B- and T-cell lines

“…Moreover, HDACi-mediated inhibition of certain DNA repair pathways can be exploited to increase efficacy and/or to overcome drug resistance. Increased cytotoxicity of 5-fluorouracil has been reported when combined with HDACi [66,67] and one study has demonstrated that loss of UNG resensitizes cells to 5-fluorodeoxyuridine (5-FdU) [68]. Our findings provide a mechanistic explanation for these observations and warrant further investigation on combinatorial effects of HDACi-and antifolate regimens.…”

A robust photoluminescence screening assay identifies uracil-DNA glycosylase inhibitors against prostate cancer

“…Since UDG depletion is known to sensitize cancer cells to chemotherapy, 93 the ability of the identied UDG inhibitor A8 to synergize with the anticancer drug, 5-FU, was next investigated in prostate cancer cells. Using the Chou-Talalay method, signicant synergism was shown between A8 and 5-FU at inhibiting RM-1 cell proliferation (Fig.…”

Targeting BER enzymes in cancer therapy

“…Instead, 5-FU cytotoxicity under the conditions employed was found to be mainly due to its incorporation into RNA rather than the DNAmediated effects [16]. However, Yan et al [18] pointed out that p53 status together with UNG plays a role in determining the cytotoxicity of 5-Fluoro-2'-deoxyuridine (5-FdU, floxuridine), another fluoropyrimidine that is efficiently incorporated into DNA rather than RNA. In absence of UNG, genomic 5-FU was found to block replication and induce double-strand breaks [125].…”

“…Moreover, many (if not most) classical chemotherapeutic treatments exert their anti-cancer effect through damage to DNA [8,9]. Irradiation [10][11][12][13][14][15], fluoropyrimidines [16][17][18], antifolates [19][20][21], platinum drugs [22][23][24][25][26][27][28], demethylating agents [29], anthracyclines and monofunctional alkylating agents such as temozolomide [30][31][32] have been shown to directly or indirectly cause the DNA damage that is typically repaired by BER. Despite progress in immuno-and targeted therapies, these classical cancer treatment agents are likely to also be extensively used in the future.…”