Sibling cases of congenital hypogonadotropic hypogonadism harboring compound heterozygous KISS1R variants

Highlights

● Compound heterozygous KISS1R variants were identified in two siblings with congenital hypogonadotropic hypogonadism.

● The p.Pro147Leu variant has been reported as pathogenic.

● Familial assessment reclassifies the p.Met136Arg variant as likely pathogenic.

Introduction

Congenital hypogonadotropic hypogonadism (CHH) is a rare disorder characterized by deficient secretion or action of gonadotropin-releasing hormone (GnRH), resulting in incomplete or absent pubertal development and infertility. CHH is clinically and genetically heterogeneous, with over 60 causative genes identified to date, including GNRHR, KISS1R, FGFR1, and others (1). Among these, variants in KISS1R, which encodes the receptor for kisspeptin, have been implicated in both familial and sporadic cases of CHH and account for approximately 2% of genetically confirmed cases. Variable degrees of hypogonadism have been reported in individuals with pathogenic variants in KISS1R, typically showing an autosomal recessive inheritance pattern. However, autosomal dominant and oligogenic contributions have also been described (2). Since reports of KISS1R variants in CHH have been extremely limited, particularly in Japan (3, 4), underscoring the need for continued accumulation of variant data to clarify genotype–phenotype correlations in KISS1R-related CHH.

Here, we report two Japanese male siblings with normosmic CHH harboring compound heterozygous variants in KISS1R. Both patients presented with micropenis in early infancy and failed to undergo spontaneous pubertal onset during adolescence.

Case Reports

Ethical consideration

The written informed consent for genetic analyses and publication of cases was obtained from the patients and their parents.

Patient 1

The proband is a 20-yr-old male born at term to healthy, non-consanguineous Japanese parents. His father experienced a pubertal growth spurt at 16 yr of age, and his mother had menarche at 14 yr, suggesting relatively delayed pubertal timing in both parents. At 4 mo of age, the patient presented with micropenis, with a stretched penile length (SPL) of 2.4 cm (–2.9 SD score), and bilateral undescended testes. Hormonal evaluation during minipuberty revealed low LH (< 0.2 mIU/mL), FSH (< 1.0 mIU/mL), and testosterone (< 0.07 ng/mL) levels; testosterone responded well to hCG stimulation (Table 1). Testosterone enanthate was administered intramuscularly, resulting in an SPL increase to 4.0 cm (+0.72 SD score). The position of the testes did not improve, and bilateral orchiopexy was performed at 9 mo of age. Testicular biopsy demonstrated age-appropriate seminiferous tubules containing Sertoli and spermatogonial cells, arranged in a compact back-to-back fashion without interstitial expansion. At 15 yr of age, he was referred to our department due to the absence of spontaneous pubertal development. His height was 155.0 cm (–2.4 SD) and body weight was 50.7 kg (–0.9 SD). Testicular volume was 3 mL bilaterally. Tanner stage was 1 for genitalia, with no pubic hair. Serum LH (< 0.1 mIU/mL), FSH (1.2 mIU/mL), and testosterone (< 0.07 ng/mL) levels were prepubertal. Both LHRH and hCG stimulation tests demonstrated poor responses (Table 1). Brain MRI revealed normal olfactory bulbs and a normal pituitary gland. Testosterone enanthate was administered monthly at a low dose, and the dosage was gradually escalated to an adult replacement level over 2 yr. After the completion of secondary sexual development at 18 yr of age, testosterone enanthate treatment was discontinued; however, gonadotropin levels remained low, and the diagnosis of CHH was clinically confirmed. At 19 yr of age, combined hCG and recombinant FSH therapy was initiated, which resulted in testicular development.

Table1. Results of stimulation tests.

Patient 2

This 14-yr-old boy was the younger brother of patient 1. He was referred at 7 mo of age for micropenis without cryptorchidism. LHRH and hCG stimulation tests at 7 mo were shown in Table 1. Following the treatment with testosterone enanthate, his SPL increased from 2.3 cm (–3.6 SD) to 3.8 cm (–0.47 SD). At 13 yr of age, his height was 148.7 cm (–1.7 SD) and body weight was 50.6 kg (± 0 SD). Testicular volume was 2 mL bilaterally, with no evidence of pubertal development. In view of the family history of CHH, further endocrine evaluation was performed. Hormonal testing revealed low basal LH and FSH levels, a weak gonadotropin response to LHRH stimulation, and a poor testosterone response to hCG stimulation (Table 1). Brain MRI showed no abnormalities.

Genetic Analysis

Genetic analyses were performed after obtaining written informed consent from the patients and their parents. Genomic DNA was isolated from peripheral blood lymphocytes. Genetic testing was performed using whole-exome sequencing with the Agilent SureSelect Target Enrichment protocol and the NovaSeq X Plus platform (Illumina, San Diego, USA) in Patient 1. Targeted analysis of 25 known CHH-related genes was conducted, including CHD7, FGF8, FGF17, FGFR1, HS6ST1, NSMF, NELP, PROK2, PROKR2, WSR11, ANOS1, KAL1, CCDC141, FEZF1, IL17RD, SEMA3A, SEMA3E, SOX10, GNRH1, GNRHR, KISS1, KISS1R, GPR54, TAC3, and TACR3. This analysis identified two heterozygous variants in KISS1R [NM_032551.5: c.407T>G (p.Met136Arg), rs765739273 and [NM_032551.5: c.440C>T (p.Pro147Leu), rs545635250]. These variants were confirmed by Sanger sequencing. The same variants were also detected in Patient 2 by Sanger sequencing. Parental analysis using Sanger sequencing confirmed that the two variants were located in trans. The Sanger sequencing data are shown in Figure 1.

Fig. 1.

Sanger sequencing for the KISS1R variants identified in the family members. Patient 1 (proband) and Patient 2 both harbor compound heterozygous KISS1R variants: c.407T>G (p.Met136Arg), inherited from the father, and c.440C>T (p.Pro147Leu), inherited from the mother.

Discussion

We identified compound heterozygous variants in the KISS1R gene in two siblings with CHH. The KISS1R gene is located on chromosome 19q13.3 and plays a crucial role in the regulation of puberty and reproductive function. It encodes a G protein-coupled receptor that binds kisspeptin, a neuropeptide essential for stimulating the release of GnRH from hypothalamic neurons. The pathogenic variants in this signaling pathway are associated with CHH (OMIM#614837).

The KISS1R p.Pro147Leu variant was previously reported in CHH (3, 4). It lies in the second intracellular loop (ICL2), a region known to be critical for receptor conformational stability and G protein coupling, and experimentally shown to be pathogenic (3). Functional study of the p.Pro147Leu has shown that this variant causes an expansion of the ligand-binding pocket, thereby impairing ligand binding and destabilizing the receptor-ligand interaction (4). These alterations result in loss of function and are consistent with the clinical presentation of these cases.

The KISS1R p.Met136Arg variant is listed in ClinVar and categorized as being of uncertain significance; however, this variant has not been reported in individuals with CHH as of September 2025. The gnomAD database showed allele frequencies of 0.00005397 overall and 0.00004706 in East Asians. The ToMMo 61KJPN database reported a frequency of 0.000042. In silico analyses (PolyPhen-2 and MutationTaster) predict this variant to be deleterious, and it received a CADD score of 33. Parental analysis confirmed in-trans transmission of this variant, and the same variant in combination with the pathogenic p.Pro147Leu variant was also identified in his sibling with CHH. Thus, familial assessment reclassified the p.Met136Arg variant as likely pathogenic (PM2, PM3, PP1, PP3) according to ACMG guidelines. Mechanistically, since this variant is located at the cytoplasmic end of TM3 near ICL2 and immediately downstream of the DRY motif, a key element for transmitting activation to the G protein (5), this variant may be associated with reduced Gq/11-dependent coupling and downstream signaling. Additional molecular analysis is needed to further clarify the pathogenicity of this variant.

In conclusion, we identified compound heterozygous, likely pathogenic variants in KISS1R in two siblings with CHH. Although KISS1R is an established causative gene for CHH, it remains a relatively rare etiology. Given the limited number of reports, particularly from Japan, our findings contribute to the genetic background of KISS1R-related CHH in Japan and help further clarify its relationship with the phenotypic spectrum.

Conflict of interests

The authors declare no conflict of interest.