Table I. Hit compounds and their validation.
Hits were validated by immunoblotting in 293MSR-GT cells (WB), flow cytometry in BHK cells (Flow), and short circuit current (Isc) analysis in Ussing chambers on epithelial MDCK cells (MDCK) or on primary Human Bronchial Epithelial (HBE) cells harvested from lungs of ΔF508/ΔF508 homozygote patients undergoing lung transplant. 293MSR-GT, BHK, and MDCK cells were stably expressing ΔF508-CFTR. For MDCK, (+) indicates rescue, (−) no observed rescue and (*) indicates increased toxicity in MDCK cells. For HBE, (+) indicates rescue observed in a sample from a patient, with (/) separating between samples from different patients, (−) indicates no observed rescue. For the flow experiments, (+) indicates >10% rescue, (±) 5–10% rescue, (−) indicates no rescue, (*) indicates increased cell toxicity and (#) indicates morphological changes observed in the treated BHK cells. For the immunoblotting experiments, (+) indicates strong rescue of ΔF508-CFTR (manifested as increase in amount of band C in comparison to vehicle-alone control), (±) poor rescue, (−) no rescue, and (*) indicates increased toxicity in 293MSR-GT cells. For dose response experiments (EC50), MDCK cells were treated with increasing concentrations (1 nM to 10 μm range) of select compounds prior to Isc analysis in Ussing chambers, (§) indicates compounds that rescue ΔF508-CFTR function at 10 μm only.
| Compound | Pathway | Target | CAS number | PubChem ID | Clinical Trials | MDCK | EC50 (MDCK) [nM] | HBE | Flow (BHK) | WB (293GT) |
|---|---|---|---|---|---|---|---|---|---|---|
| (5Z)-7-oxozeaenol | Ras/Raf/MEK/ERK or Tak1/p38 | ERK1/2, TAK1 MAP3K | 66018-38-0 | CID 9799061 | Phase I, II (E6201) | + | 60 | + +/+ + +/− +/+/+ | + | + |
| SU6668 (Orantinib) | Ras/Raf/MEK/ERK | PDGFRβ, VEGFR2, FGFR1, EGFR | 252916-29-3 | CID 5329099 | Phase I | + | 1047 | + +/+ | + | * |
| SU5402 | Ras/Raf/MEK/ERK | VEGFR2, FGFR1, PDGFRβ | 215543-92-3 | CID 5289418 | + | 12.9 | +/+/+/− /+ + | + | + | |
| EKI-785, CL-387,785 | Ras/Raf/MEK/ERK | EGFR | 194423-06-8 | CID 2776 | + | 124.6 | −/+ | ± | + | |
| FPA 124 | PI3K/Akt/mTOR | Akt/PKB | 902779-59-3 | CID 16034833 | + | § | +/+ | − | − | |
| CID 9566171 | ||||||||||
| Gsk-3β Inhibitor II | Wnt/GSK-3β | GSK-3β | 478482-75-6 | CID 6539732 | + | 127.8 | + +/+ | + | − | |
| AZD0530, Saracatinib, NSC-735464 | Src-Bcr-Abl activates: Ras/Raf/MEK/ERK (proliferation); JAK/STAT (proliferation); PI3K/Akt (mitochondrion) pathways | Src/Abl | 379231-04-6 | CID 10302451 | Phase II | + | 165.7 | + + | − | ±* |
| 7-Cyclopentyl-5-(4-phenoxyphenyl)-7H-pyrrolo[2,3-d]pyrimidin-4-ylamine (Lopac-C-8863) | T-cell receptor signaling; phosphorylates PKC and PI3K; activates Ras/Raf/MEK/ERK pathway | Lck | 213743-31-8 | CID 6603792 | + | 147.4 | +/+ + | − | ±* | |
| PD173074 | Ras/Raf/MEK/ERK | FGFR1 | 219580-11-7 | CID 1401 | + | § | +/+ | − | − | |
| PD0325901 | Ras/Raf/MEK/ERK | MEK1/2 | 391210-10-9 | CID 9826528 | Phase I | + | 6.1 | + +/+ − | + | − |
| PI-103 | PI3K/Akt/mTOR | p110 PI3Ks, mTORC1/2, DNA-PK | 371935-74-9 | CID 9884685 | + | 16.4 | −/+ | + | − | |
| RDEA-119, AR-119, BAY869766 | Ras/Raf/MEK/ERK | MEK1/2 | 923032-37-5 | CID 44182295 | Phase I, II | + | 41 | +/− | + | − |
| SKF-86002 | Tak1/p38 | p38 MAP kinase | 72873-74-6 | CID 5228 | + | 140.5 | +/− | ± | − | |
| GW5074 | Ras/Raf/MEK/ERK | Raf1 | 220904-83-6 | CID 5924208 | − | − | +/+ | + | − | |
| Kenpaullone | Wnt/GSK-3β | GSK-3β, cdks, Lck | 142273-20-9 | CID 3820 | −* | −* | + + + +/+ | − | + | |
| Alsterpaullone | Wnt/GSK-3β | GSK-3β, cdks, Lck | 237430-03-4 | CID 5005498 | −* | −* | −/+ | +# | * | |
| Ki8751 | VEGFR2 activates: Ras/Raf/MEK/ERK pathway via PKC, Akt/PKB pathway via PI3K | VEGFR2, PDGFRα, FGFR2 | 228559-41-9 | CID 11317348 | − | − | − +/+/+ | +*# | + | |
| 10-DEBC (Akt specific inhibitor X) | PI3K/Akt/mTOR | Akt/PKB | 201788-90-1 | CID 16760284 | − | − | +/− | − | + |