Hematopoietic prostaglandin D synthase (HPGDS): a high stability, Val187Ile isoenzyme common among African Americans and its relationship to risk for colorectal cancer

Abstract

Intestinal tumors in Apc(Min/+) mice are suppressed by over-production of HPGDS, which is a glutathione transferase that forms prostaglandin D(2) (PGD(2)). We characterized naturally occurring HPGDS isoenzymes, to see if HPGDS variation is associated with human colorectal cancer risk. We used DNA heteroduplex analysis and sequencing to identify HPGDS variants among healthy individuals. HPGDS isoenzymes were produced in bacteria, and their catalytic activities were tested. To determine in vivo effects, we conducted pooled case-control analyses to assess whether there is an association of the isoenzyme with colorectal cancer. Roughly 8% of African Americans and 2% of Caucasians had a highly stable Val187lle isoenzyme (with isoleucine instead of valine at position 187). At 37°C, the wild-type enzyme lost 15% of its activity in 1h, whereas the Val187Ile form remained >95% active. At 50°C, the half life of native HPGDS was 9min, compared to 42 min for Val187Ile. The odds ratio for colorectal cancer among African Americans with Val187Ile was 1.10 (95% CI, 0.75-1.62; 533 cases, 795 controls). Thus, the Val187Ile HPGDS isoenzyme common among African Americans is not associated with colorectal cancer risk. Other approaches will be needed to establish a role for HPGDS in occurrence of human intestinal tumors, as indicated by a mouse model.

Fig. 1

(A, B, C) Thermal stabilities of HPGDS isoenzymes. Enzymes were pre-incubated for the indicated times, followed by activity assays. Initial activities were set to 100%. Symbols: ●, wild-type; ■, Ile91Val; ▲, Met128Thr; ▼, Val187Ile. (A) Glutathione transferase (GST) activities at 50 °C with 1-chloro-2,4-dinitrobenzene (3 independent assays). Half-lives (minutes) were: wild-type, 8.6 ± 0.1; Ile91Val, 4.3 ± 1.9; Met128Thr, 2.0 ± 0.5; Val187Ile, 41.7 ± 9.3 (P < 0.05, ANOVA with Dunnett’s method). (B) PGD₂ synthesis activities at 50 °C with radiolabeled PGH₂. (C) PGD₂ synthesis activities at 37 °C, as in B. (D, E) Structure of the Val187Ile isoenzyme, based on the crystal structure of native, Mg²⁺-bound HPGDS at 1.8 Å resolution [PDB entries 1IYI and IIYH]. (D) Ribbon drawing of dimeric HPGDS. The Ile187 side chain is a red stick figure in helix-9. (E) Stereo views of the region near Ile187. Added van der Waals interactions occur between the C_δ atom of Ile 187 and the O (Tyr 20, 3.73 Å), C_β (Tyr 24, 3.73 Å) and C_ς (Phe 151, 3.69 Å) atoms of nearby side chains (red dotted lines). Interaction between Ile 187 and Phe 151 is predicted to give extra stability to helix-9.