. 2020 Sep 9;63(21):12929–12941. doi: 10.1021/acs.jmedchem.0c01376

Table 3. In Vitro Functional Activities of OP-NT Hybrid Peptides and Reference Peptides to the Human Opioid and NT Receptors.

	in vitro agonist activity^a
	MOR [³⁵S]GTPγS binding		MOR β-arrestin-2 recruitment		DOR [³⁵S]GTPγS binding		KOR [³⁵S]GTPγS binding		NTS1 Gαq activation
compound code/number	EC₅₀ (nM)	E_max (%)	EC₅₀ (nM)	E_max (%)	EC₅₀ (nM)	E_max (%)	EC₅₀ (nM)	E_max (%)	EC₅₀ (nM)	E_max (%)
KGOP01	0.10 ± 0.03	100 ± 7	5.53 ± 0.61	107 ± 2	1.27 ± 0.76	104 ± 14	>10,000	N.D.^c	N/A^b	N/A^b
NT(8-13)	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	0.15 ± 0.03	100 ± 6
PK20	13.3 ± 3.2	91 ± 3	956 ± 100	69 ± 4	479 ± 127	78 ± 2	47.1 ± 10	65 ± 8	19.1 ± 3.9	101 ± 3
1	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	N/A^b	1.70 ± 0.6	97 ± 10
3	12.8 ± 5.0	94 ± 6	1329 ± 365	72 ± 4	301 ± 41	106 ± 3	514 ± 7.0	34 ± 3	0.24 ± 0.2	111 ± 7
4	64.6 ± 15	99 ± 6	2459 ± 219	67 ± 6	1320 ± 67	87 ± 4	1649 ± 144	31 ± 3	0.26 ± 0.1	99 ± 17
5	9.12 ± 1.1	96 ± 2	1142 ± 187	79 ± 6	313 ± 61	91 ± 6	415 ± 56	37 ± 3	1.3 ± 0.5	101 ± 12
6	48.7 ± 14	95 ± 4	1965 ± 528	62 ± 1	898 ± 187	60 ± 5	1574 ± 681	22 ± 2	3.9 ± 1	109 ± 15
7	28.7 ± 8.8	94 ± 2	1066 ± 306	68 ± 3	305 ± 71	99 ± 4	158 ± 82	46 ± 6	0.86 ± 0.6	114 ± 16
8	65.4 ± 6.5	97 ± 2	3044 ± 526	69 ± 6	248 ± 81	97 ± 5	239 ± 62	36 ± 5	1.8 ± 1	112 ± 10
9	4.56 ± 0.25	98 ± 5	934 ± 82	67 ± 5	227 ± 8.0	97 ± 3	399 ± 69	20 ± 6	1900 ± 100	74 ± 11
10	45.6 ± 13	94 ± 3	2163 ± 99	66 ± 2	842 ± 86	91 ± 0.3	1556 ± 852	20 ± 3	N.D.^c	N.D.^c
11	6.49 ± 1.3	95 ± 6	359 ± 70	81 ± 3	177 ± 36	110 ± 2	424 ± 146	28 ± 4	6.7 ± 4	95 ± 6
12	27.4 ± 9.4	87 ± 2	1271 ± 153	66 ± 5	452 ± 118	90 ± 2	401 ± 50	30 ± 5	15 ± 7	90 ± 13

Determined in the [³⁵S]GTPγS binding assays using membranes from CHO cells stably expressing the human opioid receptors, in the PathHunter β-arrestin-2 recruitment assay with U2OS cells coexpressing the hMOR and the enzyme acceptor-tagged β-arrestin-2 fusion protein, or in the bioluminescence resonance energy transfer (BRET)-based assay conducted in HEK293 cells transiently transfected with hNTS1 and BRET biosensors. E_max values are expressed as the percentage relative to the reference agonists DAMGO (MOR), DPDPE (DOR), U69,593 (KOR), and NT(8-13) (NTS1). Data are means ± SEM of three independent experiments performed in duplicate (activity to the opioid receptors) or triplicate (activity to the NTS1 receptor).

N/A, not applicable.

N.D., no detectable signal.