In vitro characterization of peptide binding and activation
Potency at the PACAP/VIP receptors (PAC1, VPAC1, and VPAC2) was measured by competitive binding (IC50) of peptide to the PACAP/VIP receptors in CHO cell membranes overexpressing human PACAP/VIP receptors and peptide signaling (EC50) through the PACAP/VIP receptors expressed on the surface of same CHO cells using cAMP SPA assay as described under Materials and Methods. Data are the mean ± S.E. of at least two experiments. N.D., not determined.
Peptide | Receptor Competition (IC50) | cAMP SPA (EC50) | ||||||||
|---|---|---|---|---|---|---|---|---|---|---|
| VPAC2 | VPAC1 | PAC1 | VPAC2 | VPAC1 | PAC1 | |||||
| nM | nM | |||||||||
| BAY 55-9837 | 60 | 8700 | >10,000 | 0.4 | 100 | >1000 | ||||
| AcBAY | 110 ± 3 | >10,000 | >10,000 | 0.4 ± 0.1 | >1000 | >1000 | ||||
| BAY(T2) | 10,000 | >10,000 | >10,000 | 3.6 ± 0.6 | >1000 | >1000 | ||||
| BAY(E3) | >10,000 | >10,000 | >10,000 | 1.3 ± 0.1 | >1000 | >1000 | ||||
| BAY(Q9Q28) | 32 ± 1 | >10,000 | >10,000 | 0.3 ± 0.1 | 233 ± 57 | >1000 | ||||
| BAY(T2E3) | >10,000 | >10,000 | >10,000 | 12.4 ± 0.8 | >1000 | >1000 | ||||
| BAY(T2E3Q9Q28) | >10,000 | >10,000 | >10,000 | 7.8 ± 1.6 | >1000 | >1000 | ||||
| AcBAY(T2Q9Q28) | 133 ± 30 | >10,000 | >10,000 | 0.2 ± 0.1 | 196 ± 66 | >1000 | ||||
| BAY(C32) | 343 ± 86 | >10,000 | >10,000 | 2.0 ± 0.8 | 379 ± 88 | >1000 | ||||
| BAY(C32)PEG22 | N.D. | N.D. | >10,000 | 2.0 ± 0.3 | >1000 | >1000 | ||||
| BAY(C32)PEG43 | N.D. | N.D. | >10,000 | 33 ± 4 | >1000 | >1000 | ||||
| BAY(S32W33C34) | 985 ± 256 | >10,000 | >10,000 | 5.0 ± 1.6 | 883 ± 32 | >1000 | ||||
| BAY(S32W33C34)PEG22 | N.D. | N.D. | >10,000 | 3.1 ± 0.6 | >1000 | >1000 | ||||
| BAY(S32W33C34)PEG43 | N.D. | N.D. | >10,000 | 16 ± 5 | >1000 | >1000 | ||||
| BAY(Q9Q28C32) | 20 ± 3 | >10,000 | >10,000 | 0.38 ± 0.1 | >1000 | >1000 | ||||
| BAY(Q9Q28C32)PEG22 | 126 ± 9 | >10,000 | >10,000 | 1.3 ± 0.1 | >1000 | >1000 | ||||
| BAY(Q9Q28C32)PEG43 | 200 ± 26 | >10,000 | >10,000 | 18 ± 3 | >1000 | >1000 | ||||