Sepiapterin Ameliorates Chemically Induced Murine Colitis and Azoxymethane-Induced Colon Cancer

Abstract

The effects of modulating tetrahydrobiopterin (BH4) levels with a metabolic precursor, sepiapterin (SP), on dextran sodium sulfate (DSS)-induced colitis and azoxymethane (AOM)-induced colorectal cancer were studied. SP in the drinking water blocks DSS-induced colitis measured as decreased disease activity index (DAI), morphologic criteria, and recovery of Ca 21 -induced contractility responses lost as a consequence of DSS treatment. SP reduces inflammatory responses measured as the decreased number of infiltrating inflammatory macrophages and neutrophils and decreased expression of proinflammatory cytokines interleukin 1b (IL-1b), IL-6, and IL-17A. Highperformance liquid chromatography analyses of colonic BH4 and its oxidized derivative 7,8-dihydrobiopterin (BH2) are inconclusive although there was a trend for lower BH4:BH2 with DSS treatment that was reversed with SP. Reduction of colonic cGMP levels by DSS was reversed with SP by a mechanism sensitive to 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), a specific inhibitor of the NO-sensitive soluble guanylate cyclase (sGC). ODQ abrogates the protective effects of SP on colitis. This plus the finding that SP reduces DSS-enhanced protein Tyr nitration are consistent with DSS-induced uncoupling of NOS. The results agree with previous studies that demonstrated inactivation of sGC in DSS-treated animals as being important in recruitment of inflammatory cells and in altered cholinergic signaling and colon motility. SP also reduces the number of colon tumors in AOM/DSS-treated mice from 7 to 1 per unit colon length. Thus, pharmacologic modulation of BH4 with currently available drugs may provide a mechanism for alleviating some forms of colitis and potentially minimizing the potential for colorectal cancer in patients with colitis.

The Role of Nitric Oxide Synthase Uncoupling in Tumor Progression

“…We tested whether the cytotoxic activity of SP could be inhibited by the NOS inhibitor, L-NNA, previously shown to inhibit NOS in MCF-7 but without cytotoxic activity in vitro by itself (19, 35). The present results confirmed these results but also demonstrated that L-NNA completely inhibited SP induced cell killing (Figure 4C).…”

“…Breeding pairs of MMTVneu mice were from Jackson Laboratories. Drinking water was doped with 40 µg/ml SP or 0.64 mg/kg/day (19) significantly less than used in studies on vascular function (21). All animal experiments conform to protocols approved by the Institutional Animal Care and Use Committee of Virginia Commonwealth University.…”

“…Clonogenic assays and analytical methods for biopterins and cGMP have been described (3, 19). OCT blocks of human colon tissue were dissolved in PBS and the tissue collected by centrifugation prior to extracting in 0.1 N HCl for HPLC analysis.…”

“…We showed that sepiapterin (SP), a salvage pathway precursor for BH4, prophylactically blocked dextran sodium sulfate (DSS) induced colitis in mice and significantly reduced the incidence of colorectal tumors in an azoxymethane/DSS mouse model for colorectal cancer (19). HPLC analyses of total colon biopterins were inconclusive although there was a trend for lower BH4:BH2 in colons from mice treated with DSS that was reversed with SP.…”

Sepiapterin Enhances Tumor Radio- and Chemosensitivities by Promoting Vascular Normalization

“…SP also targets a single pathway, but the end product, NO, has multiple effects. As discussed above, it promotes perivascular recruitment but also regulates VEGF activity (Dulak et al, 2000), reduces infiltration of inflammatory macrophages, and reduces inflammation as measured by interleukin-6 and interleukin-1b (Cardnell et al, 2013). Tumor-associated macrophages are known to be important for tumor progression as well, and SP may influence the function of these macrophages in normalizing vasculature.…”