Kisspeptin receptor agonist has therapeutic potential for female reproductive disorders

Abstract

BACKGROUNDKisspeptin is a key regulator of hypothalamic gonadotropin-releasing hormone (GnRH) neurons and is essential for reproductive health. A specific kisspeptin receptor (KISS1R) agonist could significantly expand the potential clinical utility of therapeutics targeting the kisspeptin pathway. Herein, we investigate the effects of a KISS1R agonist, MVT-602, in healthy women and in women with reproductive disorders.METHODSWe conducted in vivo and in vitro studies to characterize the action of MVT-602 in comparison with native kisspeptin-54 (KP54). We determined the pharmacokinetic and pharmacodynamic properties of MVT-602 (doses 0.01 and 0.03 nmol/kg) versus KP54 (9.6 nmol/kg) in the follicular phase of healthy women (n = 9), and in women with polycystic ovary syndrome (PCOS; n = 6) or hypothalamic amenorrhea (HA; n = 6). Further, we investigated their effects on KISS1R-mediated inositol monophosphate (IP1) and Ca2+ signaling in cell lines and on action potential firing of GnRH neurons in brain slices.RESULTSIn healthy women, the amplitude of luteinizing hormone (LH) rise was similar to that after KP54, but peaked later (21.4 vs. 4.7 hours; P = 0.0002), with correspondingly increased AUC of LH exposure (169.0 vs. 38.5 IU∙h/L; P = 0.0058). LH increases following MVT-602 were similar in PCOS and healthy women, but advanced in HA (P = 0.004). In keeping with the clinical data, MVT-602 induced more potent signaling of KISS1R-mediated IP1 accumulation and a longer duration of GnRH neuron firing than KP54 (115 vs. 55 minutes; P = 0.0012).CONCLUSIONTaken together, these clinical and mechanistic data identify MVT-602 as having considerable therapeutic potential for the treatment of female reproductive disorders.TRIAL REGISTRATIONInternational Standard Randomised Controlled Trial Number (ISRCTN) Registry, ISRCTN21681316.FUNDINGNational Institute for Health Research and NIH.

Keywords: Endocrinology; Fertility; Reproductive Biology; Reproductive biochemistry.

Figure 1. Study protocol.

Study participants were admitted to the Clinical Research Facility at 8 am on the morning of each study visit. An i.v. cannula was inserted into one antecubital fossa, and blood was sampled at T –30 minutes, T –15 minutes, and T = 0 hours before administration of each intervention to determine the basal hormonal values. An s.c. bolus of kisspeptin-54 (KP54) or 0.9% saline (A), or of MVT-602 (B), was administered at T = 0 hours. (A) Study protocol diagram for the KP54 and 0.9% saline visits. After an s.c. bolus of KP54 (9.6 nmol/kg) or 0.9% saline at T = 0 hours, serum hormone levels (LH, FSH, estradiol, and progesterone) were measured every 5–15 minutes for the first 30 minutes, and then every 30 minutes until 10 hours, and additionally at 24 hours. (B) Study protocol diagram for MVT-602 visits. After an s.c. bolus of MVT-602 (0.03 nmol/kg) was administered at T = 0 hours, serum hormone levels (LH, FSH, estradiol, and progesterone) were measured every 5–15 minutes for the first 30 minutes, then every 30 minutes until 14 hours, and then every 60 minutes until 24 hours and additionally at 28, 32, and 48 hours. A further blood test at 72 hours was carried out in studies using estradiol pretreatment.

Figure 2. Clinical studies of MVT-602 and KP54 in healthy women in the follicular phase.

(A) Mean (± SEM) plasma concentration of MVT-602 (left y axis) or KP54 (right y axis) in pmol/L versus time (hours) in healthy women receiving an s.c. bolus of MVT-602 at 0.03 nmol/kg (blue) or 0.01 nmol/kg (black) or of KP54 (red) in the first 6 hours after s.c. administration at time 0 hours of each peptide. (B, D, and E) Mean (± SEM) change from baseline levels in serum LH (IU/L) (B), serum FSH (IU/L) (D), and serum estradiol (pmol/L) (E) in healthy women during the early follicular phase (n = 9) receiving an s.c. bolus of MVT-602 at time 0 hours. MVT-602 doses of 0.01 nmol/kg are presented in blue (over 48 hours), 0.03 nmol/kg MVT-602 in black (over 48 hours), 9.6 nmol/kg KP54 in red (over 24 hours), and 0.9% saline in purple (over 24 hours). (C) Median (IQR) of modeled values of AUC of serum LH (IU∙h/L) after KP54 (9.6 nmol/kg), MVT-602 (0.01 nmol/kg), and MVT-602 (0.03 nmol/kg). Groups were compared by Kruskal-Wallis test with post hoc Dunn’s multiple comparison. The duration of sampling was 24 hours for KP54, as LH had returned to baseline within this time frame, whereas it was 48 hours for MVT-602. *P < 0.05, **P < 0.01.

Figure 3. Effect of KP54 and MVT-602 on KISS1R-mediated Gα_q/11 signaling via IP₁ and Ca²⁺ activation.

(A) Intracellular levels of IP₁ accumulation at varying concentrations of KP54 or MVT-602 following 45 minutes of stimulation in HEK293 cells expressing FLAG‑KISS1R. Data represent mean (± SEM) of n = 4 independent experiments conducted in triplicate wells and are normalized as a percentage of ligand response. (B) Intracellular Ca²⁺ levels measured by Fluo-4AM Ca²⁺ indicator dye and live confocal microscopy. Intensity profile produced following 10 nM stimulation with KP54 (red) or MVT-602 (blue) over 1 hour of chronic stimulation in cells transiently transfected with FLAG‑KISS1R of n = 4 independent experiments conducted in duplicate wells. Data are shown following subtraction of the average background intensity for each cell as described in Methods. (C and D) Maximum intensity (C) and total area under the curve (AUC) (D), calculated from data depicted in B following ligand treatments over 1 hour of chronic stimulation. Data show individual cell analysis (total number of cells: basal, n = 1324; KP54, n = 472; MVT-602, n = 852) overlaid with the mean (± SD) values of n = 4 independent experiments conducted in duplicate wells. Cells attributed to each biological repeat are shown in the corresponding color. There were no significant differences between KP54 and MVT-602 following analysis by 2-tailed, unpaired Student’s t test.

Figure 4. Targeted extracellular recordings of firing rate of GFP-identified GnRH neurons reveal that both KP54 and MVT-602 rapidly increase action potential firing rate, but the response to MVT-602 is more prolonged.

Spontaneous basal activity (control) was recorded for 10 minutes, then either 10 nM KP54 or 10 nM MVT-602 was bath-applied for 5 minutes, followed by a wash period of at least 40 minutes. (A and B) Firing frequency (Hz in 1-minute bins) of individual GnRH neurons versus time (hours) (n = 7) before, during (gray bar), and after exposure to either 10 nM KP54 (A) or 10 nM MVT-602 (B). (C) Mean (± SEM) firing frequency of GnRH neurons (n = 7 per group, 5-minute bins) over time (hours) before, during (gray bar), and after exposure to either 10 nM KP54 (red) or 10 nM MVT-602 (blue). Statistical analysis by mixed-effect model (restricted maximum likelihood) was truncated at 65 minutes because all KP54 cells had returned to baseline (P = 0.003). (D) Median (IQR) of duration of response (hours) of GnRH neurons maintaining a firing frequency greater than 1 Hz (n = 7 per group) after exposure to 10 nM KP54 (red) or 10 nM MVT-602 (blue). Groups were compared by the Mann-Whitney U test (**P = 0.0012). (E) Median (IQR) of the peak firing frequency (Hz) of GnRH neurons (n = 7 per group) at any time point after exposure to 10 nM KP54 (red) or 10 nM MVT-602 (blue). Groups were compared by the Mann-Whitney U test (**P = 0.007).

Figure 5. Clinical studies of MVT-602 in women with oligo/anovulatory disorders.

(A) Mean (± SEM) plasma concentration of MVT-602 (pmol/L) versus time (hours) in women receiving an s.c. bolus of 0.03 nmol/kg of MVT-602. Healthy women in the follicular phase are presented in black (n = 9), women with PCOS in green (n = 6), and women with HA in blue (n = 6). No significant difference was detected by 2-way ANOVA. (B, D, and E) Mean (± SEM) change from baseline levels in serum LH (IU/L) (B), serum FSH (IU/L) (D), and serum estradiol (pmol/L) (E) in healthy women (black; n = 9), women with PCOS (green; n = 6), and women with HA (blue; n = 6) after receiving a single s.c. bolus of 0.03 nmol/kg MVT-602. Groups were compared by 2-way ANOVA. *P = 0.025 for LH, *P = 0.001 for FSH; *P = 0.02 for estradiol. (C) Median (IQR) of modeled values of AUC of serum LH (IU∙h/L) with 0.03 nmol/kg MVT-602 in women in the healthy follicular phase, women with PCOS, and women with HA. No significant difference was detected by Kruskal-Wallis test.

Figure 6. Effect of estradiol supplementation on response to MVT-602 in women during the healthy follicular phase.

(A and B) Mean (± SEM) change from baseline in serum LH (IU/L) (A) and serum FSH (IU/L) (B) in healthy women receiving a single s.c. bolus of 0.03 nmol/kg MVT-602 (blue), and in women with estradiol supplementation (n = 5 per group) via a 200-μg/d transdermal patch applied from 24 hours before MVT-602 administration at time 0 hours and continued until time 48 hours (red). (C and D) Scatterplot of median (IQR) of maximal rise in serum LH (IU/L) (C) and serum FSH (IU/L) (D) in healthy women receiving a single s.c. bolus of 0.03 nmol/kg MVT-602 (blue), and in women with estradiol supplementation (n = 5 per group) via a 200-μg/d transdermal patch applied from 24 hours before MVT-602 administration at time 0 hours and continued until time 48 hours (red). Groups were compared by Mann-Whitney U test (*P = 0.016, **P = 0.0079).