Expression of the bifunctional suicide gene CDUPRT increases radiosensitization and bystander effect of 5-FC in prostate cancer cells

Abstract

Purpose To test a hypothesis that, with 5-fluorocytosine (5-FC) treatment, the co-expression of cytosine deaminase (CD) and uracil phosphoribosyltransferase (UPRT) can lead to greater radiosensitization and bystander effect than CD-expression alone. Methods and Materials R3327-AT cell lines stably expressing CD or CDUPRT were generated. The 5-FC and 5-FU cytotoxicity, and the radiosensitivity with/without 5-FC treatment, of these cells were evaluated under both aerobic and hypoxic conditions. The bystander effect was assessed by apoptosis staining and clonogenic survival. The pharmacokinetics of 5-FU and 5-FC metabolism was followed in mice bearing CD- or CDUPRT-expressing tumors using 19F MR spectroscopy (MRS). Results CDUPRT-expressing cells were more sensitive to 5-FC and 5-FU than CD-expressing cells. CDUPRT-expression more effectively enhanced the radiosensitizing effect of 5-FC than CD-expression. A 25-fold-lower dose of 5-FC resulted in the same magnitude of radiosensitization in CDUPRT-expressing cells, relative to that in CD-expressing cells. The 5-FC cytotoxicity in co-cultures of parental cells mixed with 10–20% CDUPRT cells was similar as that in 100% CDUPRT cells. 19F MRS measurements showed that expression of CDUPRT leads to enhanced accumulation of fluorine nucleotide (FNuc) in vivo. Conclusion Our study indicates that CDUPRT/5-FC strategy may be more effective than CD/5-FC when used as an adjuvant to radiotherapy.

Targeted Chemotherapy for Head and Neck Cancer with a Chimeric Oncolytic Adenovirus Coding for Bifunctional Suicide Protein FCU1

“…Instead, efficacy was better with the combination, which might increase the therapeutic window of the approach because enhanced efficacy together with less virus replication might mean less replication-associated side effects. Lastly, we observed an increase in apoptosis in tumors treated with Ad5/3-Δ24FCU1 + 5-FC comparing to Ad5/3-Δ24FCU1 or 5-FC alone, confirming the mechanism of action of 5-FU and 5-FUMP (40). Because the data on apoptosis and oncolytic adenovirus replication are somewhat discordant (41)(42)(43)(44), perhaps in part due to the antiapoptotic activities of E1B and E4 (44,45), we assume that the increase in apoptosis seen here may be chiefly due to the 5-FU-mediated bystander effect.…”

A triple suicide gene strategy that improves therapeutic effects and incorporates multimodality molecular imaging for monitoring gene functions

“…CD and HSV1-TK, or CD and UPRT), resulting in improved tumoricidal effect for different tumors. 12–15,23 Both the in vitro and in vivo data in the present study (Fig. 3–5 and supplementary Fig.…”

“…25 Previously, we have demonstrated that CDUPRT/5-FC approach has greater radiosensitization effect than the CD/5-FC system. 15 Therefore, it is not surprising that significant tumor control effects were found as triple suicide gene approach combined with radiation.…”

“…12,13 Simultaneous expression of CD and UPRT genes could lead to synergistic effects resulting in increased sensitivity to 5-FC. 14,15 Among double suicide gene therapy strategies, the combination of HSV1-TK/GCV and CD/5-FC is most widely studied and has been evaluated in clinical trials 16,17 . Subsequently, a reporter gene system (the human sodium iodide symporter, hNIS) was incorporated into a double suicide gene (TK/CD) adenovirus, and Na 99m TcO 4 SPECT images demonstrated gene expression after virus injection into human prostate cancer.…”

Regulation of gene expression in prostate cancer cells with an artificially constructed promoter responsive to radiation

“…It has been pointed out that combination of radiation therapy and gene therapy, called 'radio-genetic therapy' , is an attractive prospect, [11][12][13] as the combination may potentially overcome these defects of either therapy. It may use a gene whose product is capable of improving the anticancer effects of ionizing radiation, and/or controlling the expression of therapeutic genes by irradiation.…”