Table 1. IC50s (in nM) for 6-substituted pyrrolo[2,3-d]pyrimidine thienoyl antifolates 1–4, and classical antifolates in hRFC-, hPCFT-, and FR-expressing cell lines.
Growth inhibition assays were performed for CHO sublines engineered to express hRFC (PC43-10), FRs (RT16, D4), or hPCFT (R2/hPCFT4), for comparison with transporter-null [R2, R2(VC)] CHO cells, and for the KB and IGROV1 tumor sublines (express hRFC, FRα, and hPCFT), as described in the Experimental Section. For the FR experiments, growth inhibition assays were performed in the presence and the absence of 200 nM folic acid (FA). The data shown are mean values from 2–10 experiments (plus/minus SEM in parentheses). Results are presented as IC50 values (in units of nM), corresponding to the concentrations that inhibit growth by 50% relative to cells incubated without drug. Data for 4, MTX, PMX, RTX, LMTX, and 9 were previously published12–15. ND: Not determined
| Antifolate | hRFC | FRα | FRβ | hPCFT | hRFC/FRα/hPCFT | hRFC/FRα/hPCFT | ||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| PC43-10 | R2 | RT16 | RT16 (+FA) |
D4 | D4 (+FA) |
R2/hPCFT4 | R2(VC) | KB | KB (+FA) |
IGROV1 | IGROV1 (+FA) |
|
| 1 | >1000 | >1000 | 122 (86) | >1000 | ND | ND | 291 (71) | >1000 | 41.9 (24) | >1000 | >1000 | >1000 |
| 2 | >1000 | >1000 | 38.9 (10.2) | >1000 | 6.58 (0.95) | >1000 | 135 (18) | >1000 | 3.22 (0.50) | >1000 | 122.7 (32.2) | >1000 |
| 3 | 101.0 (16.6) | 273.5 (49.1) | 0.31 (0.14) | >1000 | 0.17 (0.03) | >1000 | 3.34 (0.26) | 288 (12) | 0.26 (0.03) | >1000 | 0.55 (0.10) | >1000 |
| 4 | >1000 | >1000 | 1.82(0.28) | >1000 | 0.57(0.09) | >1000 | 43.4(4.1) | >1000 | 0.55(0.10) | >1000 | 0.97(0.12) | >1000 |
| MTX | 12(1.1) | 216(8.7) | 114(31) | 461(62) | 106(11) | 211(43) | 121 (17) | >1000 | 6.0(0.6) | 20(2.4) | 21(3.4) | 22(2.1) |
| PMX | 138(13) | 894(93) | 42(9) | 388(68) | 60(8) | 254(78) | 13.2(2.4) | 974 (18) | 68(12) | 327(103) | 102(25) | 200(18) |
| RTX | 6.3(1.3) | >1000 | 15(5) | >1000 | 22(10) | 746(138) | 99.5(11.4) | >1000 | 5.9(2.2) | 22(5) | 12.6(3.3) | 20(4.3) |
| LMTX | 12(2.3) | >1000 | 12(8) | >188(39) | 2.6(1.0) | 275(101) | 38.0(5.3) | >1000 | 1.2(0.6) | 31(7) | 3.1(0.9) | 16(6) |
| 9 | 1.28(0.18) | >1000 | 409(51) | 864(39) | ND | ND | >1000 | >1000 | 5.26(1.07) | 2.90(0.16) | 3.47(0.48) | 2.47(0.38) |