TCF12 haploinsufficiency causes autosomal dominant Kallmann syndrome and reveals network-level interactions between causal loci.

Dysfunction of the gonadotropin-releasing hormone (GnRH) axis causes a range of reproductive phenotypes resulting from defects in the specification, migration and/or function of GnRH neurons. To identify additional molecular components of this system, we initiated a systematic genetic interrogation of families with isolated GnRH deficiency (IGD). Here, we report 13 families (12 autosomal dominant and one autosomal recessive) with an anosmic form of IGD (Kallmann syndrome) with loss-of-function mutations in TCF12, a locus also known to cause syndromic and non-syndromic craniosynostosis. We show that loss of tcf12 in zebrafish larvae perturbs GnRH neuronal patterning with concomitant attenuation of the orthologous expression of tcf3a/b, encoding a binding partner of TCF12, and stub1, a gene that is both mutated in other syndromic forms of IGD and maps to a TCF12 affinity network. Finally, we report that restored STUB1 mRNA rescues loss of tcf12 in vivo. Our data extend the mutational landscape of IGD, highlight the genetic links between craniofacial patterning and GnRH dysfunction and begin to assemble the functional network that regulates the development of the GnRH axis.

The p190 RhoGAPs, ARHGAP35, and ARHGAP5 are implicated in GnRH neuronal development: Evidence from patients with idiopathic hypogonadotropic hypogonadism, zebrafish, and in vitro GAP activity assay.

PURPOSE: The study aimed to identify novel genes for idiopathic hypogonadotropic hypogonadism (IHH). METHODS: A cohort of 1387 probands with IHH underwent exome sequencing and de novo, familial, and cohort-wide investigations. Functional studies were performed on 2 p190 Rho GTPase-activating proteins (p190 RhoGAP), ARHGAP35 and ARHGAP5, which involved in vivo modeling in larval zebrafish and an in vitro p190A-GAP activity assay. RESULTS: Rare protein-truncating variants (PTVs; n = 5) and missense variants in the RhoGAP domain (n = 7) in ARHGAP35 were identified in IHH cases (rare variant enrichment: PTV [unadjusted P = 3.1E-06] and missense [adjusted P = 4.9E-03] vs controls). Zebrafish modeling using gnrh3:egfp phenotype assessment showed that mutant larvae with deficient arhgap35a, the predominant ARHGAP35 paralog in the zebrafish brain, display decreased GnRH3-GFP+ neuronal area, a readout for IHH. In vitro GAP activity studies showed that 1 rare missense variant [ARHGAP35 p.(Arg1284Trp)] had decreased GAP activity. Rare PTVs (n = 2) also were discovered in ARHGAP5, a paralog of ARHGAP35; however, arhgap5 zebrafish mutants did not display significant GnRH3-GFP+ abnormalities. CONCLUSION: This study identified ARHGAP35 as a new autosomal dominant genetic driver for IHH and ARHGAP5 as a candidate gene for IHH. These observations suggest a novel role for the p190 RhoGAP proteins in GnRH neuronal development and integrity.

Phenotypic continuum between Waardenburg syndrome and idiopathic hypogonadotropic hypogonadism in humans with SOX10 variants.

PURPOSE: SOX10 variants previously implicated in Waardenburg syndrome (WS) have now been linked to Kallmann syndrome (KS), the anosmic form of idiopathic hypogonadotropic hypogonadism (IHH). We investigated whether SOX10-associated WS and IHH represent elements of a phenotypic continuum within a unifying disorder or if they represent phenotypically distinct allelic disorders. METHODS: Exome sequencing from 1,309 IHH subjects (KS: 632; normosmic idiopathic hypogonadotropic hypogonadism [nIIHH]: 677) were reviewed for SOX10 rare sequence variants (RSVs). The genotypic and phenotypic spectrum of SOX10-related IHH (this study and literature) and SOX10-related WS cases (literature) were reviewed and compared with SOX10-RSV spectrum in gnomAD population. RESULTS: Thirty-seven SOX10-associated IHH cases were identified as follows: current study: 16 KS; 4 nIHH; literature: 16 KS; 1 nIHH. Twenty-three IHH cases (62%; all KS), had ≥1 known WS-associated feature(s). Moreover, five previously reported SOX10-associated WS cases showed IHH-related features. Four SOX10 missense RSVs showed allelic overlap between IHH-ascertained and WS-ascertained cases. The SOX10-HMG domain showed an enrichment of RSVs in disease states versus gnomAD. CONCLUSION: SOX10 variants contribute to both anosmic (KS) and normosmic (nIHH) forms of IHH. IHH and WS represent SOX10-associated developmental defects that lie along a unifying phenotypic continuum. The SOX10-HMG domain is critical for the pathogenesis of SOX10-related human disorders.

Modeling mutant/wild-type interactions to ascertain pathogenicity of PROKR2 missense variants in patients with isolated GnRH deficiency.

A major challenge in human genetics is the validation of pathogenicity of heterozygous missense variants. This problem is well-illustrated by PROKR2 variants associated with Isolated GnRH Deficiency (IGD). Homozygous, loss of function variants in PROKR2 was initially implicated in autosomal recessive IGD; however, most IGD-associated PROKR2 variants are heterozygous. Moreover, while IGD patient cohorts are enriched for PROKR2 missense variants similar rare variants are also found in normal individuals. To elucidate the pathogenic mechanisms distinguishing IGD-associated PROKR2 variants from rare variants in controls, we assessed 59 variants using three approaches: (i) in silico prediction, (ii) traditional in vitro functional assays across three signaling pathways with mutant-alone transfections, and (iii) modified in vitro assays with mutant and wild-type expression constructs co-transfected to model in vivo heterozygosity. We found that neither in silico analyses nor traditional in vitro assessments of mutants transfected alone could distinguish IGD variants from control variants. However, in vitro co-transfections revealed that 15/34 IGD variants caused loss-of-function (LoF), including 3 novel dominant-negatives, while only 4/25 control variants caused LoF. Surprisingly, 19 IGD-associated variants were benign or exhibited LoF that could be rescued by WT co-transfection. Overall, variants that were LoF in ≥ 2 signaling assays under co-transfection conditions were more likely to be disease-associated than benign or 'rescuable' variants. Our findings suggest that in vitro modeling of WT/Mutant interactions increases the resolution for identifying causal variants, uncovers novel dominant negative mutations, and provides new insights into the pathogenic mechanisms underlying heterozygous PROKR2 variants.

DLG2 variants in patients with pubertal disorders.

PURPOSE: Impaired function of gonadotropin-releasing hormone (GnRH) neurons can cause a phenotypic spectrum ranging from delayed puberty to isolated hypogonadotropic hypogonadism (IHH). We sought to identify a new genetic etiology for these conditions. METHODS: Exome sequencing was performed in an extended family with autosomal dominant, markedly delayed puberty. The effects of the variant were studied in a GnRH neuronal cell line. Variants in the same gene were sought in a large cohort of individuals with IHH. RESULTS: We identified a rare missense variant (F900V) in DLG2 (which encodes PSD-93) that cosegregated with the delayed puberty. The variant decreased GnRH expression in vitro. PSD-93 is an anchoring protein of NMDA receptors, a type of glutamate receptor that has been implicated in the control of puberty in laboratory animals. The F900V variant impaired the interaction between PSD-93 and a known binding partner, Fyn, which phosphorylates NMDA receptors. Variants in DLG2 that also decreased GnRH expression were identified in three unrelated families with IHH. CONCLUSION: The findings indicate that variants in DLG2/PSD-93 cause autosomal dominant delayed puberty and may also contribute to IHH. The findings also suggest that the pathogenesis involves impaired NMDA receptor signaling and consequently decreased GnRH secretion.

Reproductive endocrine phenotypes relating to CHD7 mutations in humans.

Mutations in the gene CHD7 cause CHARGE syndrome, a rare multi-organ syndromic disorder. Gonadal defects are common in individuals with CHARGE syndrome (seen in ∼60-80% of cases) and represent the letter "G" in the CHARGE syndrome acronym. The gonadal defect in CHARGE syndrome results from congenital deficiency of the hypothalamic hormone Gonadotropin-releasing hormone (GnRH), which manifests clinically as pubertal failure and infertility, and biochemically as hypogonadotropic hypogonadism (low sex steroid hormone levels with inappropriately normal or low gonadotropin levels). In addition to the gonadal endocrine abnormalities, in a small minority of individuals with CHARGE, additional endocrine defects including growth hormone deficiency, multiple pituitary hormone deficits and primary hypothyroidism may also be seen. CHD7 mutations disrupt the targeting of olfactory axons and the migration of GnRH-synthesizing neurons during embryonic development, resulting in congenital idiopathic hypogonadotropic hypogonadism (IHH) and anosmia (or hyposmia), two features that define human Kallmann syndrome. Since Kallmann syndrome is one of the constituent phenotypes within CHARGE, recent studies have investigated the role of CHD7 mutations in individuals with IHH and established that deleterious missense mutations in CHD7 are associated with Kallmann syndrome as well as normosmic form of IHH. These missense mutations affect the ATPase and nucleosome remodeling activities of the CHD7 protein. These observations suggest that CHD7 protein function is critical for the ontogeny of GnRH neurons and neuroendocrine regulation of GnRH secretion.

Functionally compromised CHD7 alleles in patients with isolated GnRH deficiency.

Inactivating mutations in chromodomain helicase DNA binding protein 7 (CHD7) cause CHARGE syndrome, a severe multiorgan system disorder of which Isolated gonadotropin-releasing hormone (GnRH) deficiency (IGD) is a minor feature. Recent reports have described predominantly missense CHD7 alleles in IGD patients, but it is unclear if these alleles are relevant to causality or overall genetic burden of Kallmann syndrome (KS) and normosmic form of IGD. To address this question, we sequenced CHD7 in 783 well-phenotyped IGD patients lacking full CHARGE features; we identified nonsynonymous rare sequence variants in 5.2% of the IGD cohort (73% missense and 27% splice variants). Functional analyses in zebrafish using a surrogate otolith assay of a representative set of these CHD7 alleles showed that rare sequence variants observed in controls showed no altered function. In contrast, 75% of the IGD-associated alleles were deleterious and resulted in both KS and normosmic IGD. In two families, pathogenic mutations in CHD7 coexisted with mutations in other known IGD genes. Taken together, our data suggest that rare deleterious CHD7 alleles contribute to the mutational burden of patients with both KS and normosmic forms of IGD in the absence of full CHARGE syndrome. These findings (i) implicate a unique role or preferential sensitivity for CHD7 in the ontogeny of GnRH neurons, (ii) reiterate the emerging genetic complexity of this family of IGD disorders, and (iii) demonstrate how the coordinated use of well-phenotyped cohorts, families, and functional studies can inform genetic architecture and provide insights into the developmental biology of cellular systems.

Mutations in FGF17, IL17RD, DUSP6, SPRY4, and FLRT3 are identified in individuals with congenital hypogonadotropic hypogonadism.

Congenital hypogonadotropic hypogonadism (CHH) and its anosmia-associated form (Kallmann syndrome [KS]) are genetically heterogeneous. Among the >15 genes implicated in these conditions, mutations in FGF8 and FGFR1 account for ~12% of cases; notably, KAL1 and HS6ST1 are also involved in FGFR1 signaling and can be mutated in CHH. We therefore hypothesized that mutations in genes encoding a broader range of modulators of the FGFR1 pathway might contribute to the genetics of CHH as causal or modifier mutations. Thus, we aimed to (1) investigate whether CHH individuals harbor mutations in members of the so-called "FGF8 synexpression" group and (2) validate the ability of a bioinformatics algorithm on the basis of protein-protein interactome data (interactome-based affiliation scoring [IBAS]) to identify high-quality candidate genes. On the basis of sequence homology, expression, and structural and functional data, seven genes were selected and sequenced in 386 unrelated CHH individuals and 155 controls. Except for FGF18 and SPRY2, all other genes were found to be mutated in CHH individuals: FGF17 (n = 3 individuals), IL17RD (n = 8), DUSP6 (n = 5), SPRY4 (n = 14), and FLRT3 (n = 3). Independently, IBAS predicted FGF17 and IL17RD as the two top candidates in the entire proteome on the basis of a statistical test of their protein-protein interaction patterns to proteins known to be altered in CHH. Most of the FGF17 and IL17RD mutations altered protein function in vitro. IL17RD mutations were found only in KS individuals and were strongly linked to hearing loss (6/8 individuals). Mutations in genes encoding components of the FGF pathway are associated with complex modes of CHH inheritance and act primarily as contributors to an oligogenic genetic architecture underlying CHH.

An ancient founder mutation in PROKR2 impairs human reproduction.

Congenital gonadotropin-releasing hormone (GnRH) deficiency manifests as absent or incomplete sexual maturation and infertility. Although the disease exhibits marked locus and allelic heterogeneity, with the causal mutations being both rare and private, one causal mutation in the prokineticin receptor, PROKR2 L173R, appears unusually prevalent among GnRH-deficient patients of diverse geographic and ethnic origins. To track the genetic ancestry of PROKR2 L173R, haplotype mapping was performed in 22 unrelated patients with GnRH deficiency carrying L173R and their 30 first-degree relatives. The mutation's age was estimated using a haplotype-decay model. Thirteen subjects were informative and in all of them the mutation was present on the same ~123 kb haplotype whose population frequency is ≤10%. Thus, PROKR2 L173R represents a founder mutation whose age is estimated at approximately 9000 years. Inheritance of PROKR2 L173R-associated GnRH deficiency was complex with highly variable penetrance among carriers, influenced by additional mutations in the other PROKR2 allele (recessive inheritance) or another gene (digenicity). The paradoxical identification of an ancient founder mutation that impairs reproduction has intriguing implications for the inheritance mechanisms of PROKR2 L173R-associated GnRH deficiency and for the relevant processes of evolutionary selection, including potential selective advantages of mutation carriers in genes affecting reproduction.

A genetic basis for functional hypothalamic amenorrhea.

BACKGROUND: Functional hypothalamic amenorrhea is a reversible form of gonadotropin-releasing hormone (GnRH) deficiency commonly triggered by stressors such as excessive exercise, nutritional deficits, or psychological distress. Women vary in their susceptibility to inhibition of the reproductive axis by such stressors, but it is unknown whether this variability reflects a genetic predisposition to hypothalamic amenorrhea. We hypothesized that mutations in genes involved in idiopathic hypogonadotropic hypogonadism, a congenital form of GnRH deficiency, are associated with hypothalamic amenorrhea. METHODS: We analyzed the coding sequence of genes associated with idiopathic hypogonadotropic hypogonadism in 55 women with hypothalamic amenorrhea and performed in vitro studies of the identified mutations. RESULTS: Six heterozygous mutations were identified in 7 of the 55 patients with hypothalamic amenorrhea: two variants in the fibroblast growth factor receptor 1 gene FGFR1 (G260E and R756H), two in the prokineticin receptor 2 gene PROKR2 (R85H and L173R), one in the GnRH receptor gene GNRHR (R262Q), and one in the Kallmann syndrome 1 sequence gene KAL1 (V371I). No mutations were found in a cohort of 422 controls with normal menstrual cycles. In vitro studies showed that FGFR1 G260E, FGFR1 R756H, and PROKR2 R85H are loss-of-function mutations, as has been previously shown for PROKR2 L173R and GNRHR R262Q. CONCLUSIONS: Rare variants in genes associated with idiopathic hypogonadotropic hypogonadism are found in women with hypothalamic amenorrhea, suggesting that these mutations may contribute to the variable susceptibility of women to the functional changes in GnRH secretion that characterize hypothalamic amenorrhea. Our observations provide evidence for the role of rare variants in common multifactorial disease. (Funded by the Eunice Kennedy Shriver National Institute of Child Health and Human Development and others; ClinicalTrials.gov number, NCT00494169.).

A novel syndrome caused by the E410K amino acid substitution in the neuronal ß-tubulin isotype 3.

Missense mutations in TUBB3, the gene that encodes the neuronal-specific protein ß-tubulin isotype 3, can cause isolated or syndromic congenital fibrosis of the extraocular muscles, a form of complex congenital strabismus characterized by cranial nerve misguidance. One of the eight TUBB3 mutations reported to cause congenital fibrosis of the extraocular muscles, c.1228G>A results in a TUBB3 E410K amino acid substitution that directly alters a kinesin motor protein binding site. We report the detailed phenotypes of eight unrelated individuals who harbour this de novo mutation, and thus define the 'TUBB3 E410K syndrome'. Individuals harbouring this mutation were previously reported to have congenital fibrosis of the extraocular muscles, facial weakness, developmental delay and possible peripheral neuropathy. We now confirm by electrophysiology that a progressive sensorimotor polyneuropathy does indeed segregate with the mutation, and expand the TUBB3 E410K phenotype to include Kallmann syndrome (hypogonadotropic hypogonadism and anosmia), stereotyped midface hypoplasia, intellectual disabilities and, in some cases, vocal cord paralysis, tracheomalacia and cyclic vomiting. Neuroimaging reveals a thin corpus callosum and anterior commissure, and hypoplastic to absent olfactory sulci, olfactory bulbs and oculomotor and facial nerves, which support underlying abnormalities in axon guidance and maintenance. Thus, the E410K substitution defines a new genetic aetiology for Moebius syndrome, Kallmann syndrome and cyclic vomiting. Moreover, the c.1228G>A mutation was absent in DNA from ∼600 individuals who had either Kallmann syndrome or isolated or syndromic ocular and/or facial dysmotility disorders, but who did not have the combined features of the TUBB3 E410K syndrome, highlighting the specificity of this phenotype-genotype correlation. The definition of the TUBB3 E410K syndrome will allow clinicians to identify affected individuals and predict the mutation based on clinical features alone.

Heparan sulfate 6-O-sulfotransferase 1, a gene involved in extracellular sugar modifications, is mutated in patients with idiopathic hypogonadotrophic hypogonadism.

Neuronal development is the result of a multitude of neural migrations, which require extensive cell-cell communication. These processes are modulated by extracellular matrix components, such as heparan sulfate (HS) polysaccharides. HS is molecularly complex as a result of nonrandom modifications of the sugar moieties, including sulfations in specific positions. We report here mutations in HS 6-O-sulfotransferase 1 (HS6ST1) in families with idiopathic hypogonadotropic hypogonadism (IHH). IHH manifests as incomplete or absent puberty and infertility as a result of defects in gonadotropin-releasing hormone neuron development or function. IHH-associated HS6ST1 mutations display reduced activity in vitro and in vivo, suggesting that HS6ST1 and the complex modifications of extracellular sugars are critical for normal development in humans. Genetic experiments in Caenorhabditis elegans reveal that HS cell-specifically regulates neural branching in vivo in concert with other IHH-associated genes, including kal-1, the FGF receptor, and FGF. These findings are consistent with a model in which KAL1 can act as a modulatory coligand with FGF to activate the FGF receptor in an HS-dependent manner.

Oligogenic basis of isolated gonadotropin-releasing hormone deficiency.

Between the genetic extremes of rare monogenic and common polygenic diseases lie diverse oligogenic disorders involving mutations in more than one locus in each affected individual. Elucidating the principles of oligogenic inheritance and mechanisms of genetic interactions could help unravel the newly appreciated role of rare sequence variants in polygenic disorders. With few exceptions, however, the precise genetic architecture of oligogenic diseases remains unknown. Isolated gonadotropin-releasing hormone (GnRH) deficiency caused by defective secretion or action of hypothalamic GnRH is a rare genetic disease that manifests as sexual immaturity and infertility. Recent reports of patients who harbor pathogenic rare variants in more than one gene have challenged the long-held view that the disorder is strictly monogenic, yet the frequency and extent of oligogenicity in isolated GnRH deficiency have not been investigated. By systematically defining genetic variants in large cohorts of well-phenotyped patients (n = 397), family members, and unaffected subjects (n = 179) for the majority of known disease genes, this study suggests a significant role of oligogenicity in this disease. Remarkably, oligogenicity in isolated GnRH deficiency was as frequent as homozygosity/compound heterozygosity at a single locus (2.5%). Among the 22% of patients with detectable rare protein-altering variants, the likelihood of oligogenicity was 11.3%. No oligogenicity was detected among controls (P < 0.05), even though deleterious variants were present. Viewing isolated GnRH deficiency as an oligogenic condition has implications for understanding the pathogenesis of its reproductive and nonreproductive phenotypes; deciphering the etiology of common GnRH-related disorders; and modeling the genetic architecture of other oligogenic and multifactorial diseases.

Heterozygous mutations in SOX2 may cause idiopathic hypogonadotropic hypogonadism via dominant-negative mechanisms.

Pathogenic SRY-box transcription factor 2 (SOX2) variants typically cause severe ocular defects within a SOX2 disorder spectrum that includes hypogonadotropic hypogonadism. We examined exome-sequencing data from a large, well-phenotyped cohort of patients with idiopathic hypogonadotropic hypogonadism (IHH) for pathogenic SOX2 variants to investigate the underlying pathogenic SOX2 spectrum and its associated phenotypes. We identified 8 IHH individuals harboring heterozygous pathogenic SOX2 variants with variable ocular phenotypes. These variant proteins were tested in vitro to determine whether a causal relationship between IHH and SOX2 exists. We found that Sox2 was highly expressed in the hypothalamus of adult mice and colocalized with kisspeptin 1 (KISS1) expression in the anteroventral periventricular nucleus of adult female mice. In vitro, shRNA suppression of mouse SOX2 protein in Kiss-expressing cell lines increased the levels of human kisspeptin luciferase (hKiss-luc) transcription, while SOX2 overexpression repressed hKiss-luc transcription. Further, 4 of the identified SOX2 variants prevented this SOX2-mediated repression of hKiss-luc. Together, these data suggest that pathogenic SOX2 variants contribute to both anosmic and normosmic forms of IHH, attesting to hypothalamic defects in the SOX2 disorder spectrum. Our study describes potentially novel mechanisms contributing to SOX2-related disease and highlights the necessity of SOX2 screening in IHH genetic evaluation irrespective of associated ocular defects.

GNRH1 mutations in patients with idiopathic hypogonadotropic hypogonadism.

Idiopathic hypogonadotropic hypogonadism (IHH) is a condition characterized by failure to undergo puberty in the setting of low sex steroids and low gonadotropins. IHH is due to abnormal secretion or action of the master reproductive hormone gonadotropin-releasing hormone (GnRH). Several genes have been found to be mutated in patients with IHH, yet to date no mutations have been identified in the most obvious candidate gene, GNRH1 itself, which encodes the preprohormone that is ultimately processed to produce GnRH. We screened DNA from 310 patients with normosmic IHH (nIHH) and 192 healthy control subjects for sequence changes in GNRH1. In 1 patient with severe congenital nIHH (with micropenis, bilateral cryptorchidism, and absent puberty), a homozygous frameshift mutation that is predicted to disrupt the 3 C-terminal amino acids of the GnRH decapeptide and to produce a premature stop codon was identified. Heterozygous variants not seen in controls were identified in 4 patients with nIHH: 1 nonsynonymous missense mutation in the eighth amino acid of the GnRH decapeptide, 1 nonsense mutation that causes premature termination within the GnRH-associated peptide (GAP), which lies C-terminal to the GnRH decapeptide within the GnRH precursor, and 2 sequence variants that cause nonsynonymous amino-acid substitutions in the signal peptide and in GnRH-associated peptide. Our results establish mutations in GNRH1 as a genetic cause of nIHH.

Prevalence and Phenotypic Effects of Copy Number Variants in Isolated Hypogonadotropic Hypogonadism.

CONTEXT: The genetic architecture of isolated hypogonadotropic hypogonadism (IHH) has not been completely defined. OBJECTIVE: To determine the role of copy number variants (CNVs) in IHH pathogenicity and define their phenotypic spectrum. METHODS: Exome sequencing (ES) data in IHH probands (n = 1394) (Kallmann syndrome [IHH with anosmia; KS], n = 706; normosmic IHH [nIHH], n = 688) and family members (n = 1092) at the Reproductive Endocrine Unit and the Center for Genomic Medicine of Massachusetts General Hospital were analyzed for CNVs and single nucleotide variants (SNVs)/indels in 62 known IHH genes. IHH subjects without SNVs/indels in known genes were considered "unsolved." Phenotypes associated with CNVs were evaluated through review of patient medical records. A total of 29 CNVs in 13 genes were detected (overall IHH cohort prevalence: ~2%). Almost all (28/29) CNVs occurred in unsolved IHH cases. While some genes (eg, ANOS1 and FGFR1) frequently harbor both CNVs and SNVs/indels, the mutational spectrum of others (eg, CHD7) was restricted to SNVs/indels. Syndromic phenotypes were seen in 83% and 63% of IHH subjects with multigenic and single gene CNVs, respectively. CONCLUSION: CNVs in known genes contribute to ~2% of IHH pathogenesis. Predictably, multigenic contiguous CNVs resulted in syndromic phenotypes. Syndromic phenotypes resulting from single gene CNVs validate pleiotropy of some IHH genes. Genome sequencing approaches are now needed to identify novel genes and/or other elusive variants (eg, noncoding/complex structural variants) that may explain the remaining missing etiology of IHH.

Reversal of idiopathic hypogonadotropic hypogonadism.

BACKGROUND: Idiopathic hypogonadotropic hypogonadism, which may be associated with anosmia (the Kallmann syndrome) or with a normal sense of smell, is a treatable form of male infertility caused by a congenital defect in the secretion or action of gonadotropin-releasing hormone (GnRH). Patients have absent or incomplete sexual maturation by the age of 18. Idiopathic hypogonadotropic hypogonadism was previously thought to require lifelong therapy. We describe 15 men in whom reversal of idiopathic hypogonadotropic hypogonadism was sustained after discontinuation of hormonal therapy. METHODS: We defined the sustained reversal of idiopathic hypogonadotropic hypogonadism as the presence of normal adult testosterone levels after hormonal therapy was discontinued. RESULTS: Ten sustained reversals were identified retrospectively. Five sustained reversals were identified prospectively among 50 men with idiopathic hypogonadotropic hypogonadism after a mean (+/-SD) duration of treatment interruption of 6+/-3 weeks. Of the 15 men who had a sustained reversal, 4 had anosmia. At initial evaluation, 6 men had absent puberty, 9 had partial puberty, and all had abnormal secretion of GnRH-induced luteinizing hormone. All 15 men had received previous hormonal therapy to induce virilization, fertility, or both. Among those whose hypogonadism was reversed, the mean serum level of endogenous testosterone increased from 55+/-29 ng per deciliter (1.9+/-1.0 nmol per liter) to 386+/-91 ng per deciliter (13.4+/-3.2 nmol per liter, P<0.001), the luteinizing hormone level increased from 2.7+/-2.0 to 8.5+/-4.6 IU per liter (P<0.001), the level of follicle-stimulating hormone increased from 2.5+/-1.7 to 9.5+/-12.2 IU per liter (P<0.01), and testicular volume increased from 8+/-5 to 16+/-7 ml (P<0.001). Pulsatile luteinizing hormone secretion and spermatogenesis were documented. CONCLUSIONS: Sustained reversal of normosmic idiopathic hypogonadotropic hypogonadism and the Kallmann syndrome was noted after discontinuation of treatment in about 10% of patients with either absent or partial puberty. Therefore, brief discontinuation of hormonal therapy to assess reversibility of hypogonadotropic hypogonadism is reasonable. (ClinicalTrials.gov number, NCT00392756 [ClinicalTrials.gov].).

Human GnRH deficiency: a unique disease model to unravel the ontogeny of GnRH neurons.

Evolutionary survival of a species is largely a function of its reproductive fitness. In mammals, a sparsely populated and widely dispersed network of hypothalamic neurons, the gonadotropin-releasing hormone (GnRH) neurons, serve as the pilot light of reproduction via coordinated secretion of GnRH. Since it first description, human GnRH deficiency has been recognized both clinically and genetically as a heterogeneous disease. A spectrum of different reproductive phenotypes comprised of congenital GnRH deficiency with anosmia (Kallmann syndrome), congenital GnRH deficiency with normal olfaction (normosmic idiopathic hypogonadotropic hypogonadism), and adult-onset hypogonadotropic hypogonadism has been described. In the last two decades, several genes and pathways which govern GnRH ontogeny have been discovered by studying humans with GnRH deficiency. More importantly, detailed study of these patients has highlighted the emerging theme of oligogenicity and genotypic synergism, and also expanded the phenotypic diversity with the documentation of reversal of GnRH deficiency later in adulthood in some patients. The underlying genetic defect has also helped understand the associated nonreproductive phenotypes seen in some of these patients. These insights now provide practicing clinicians with targeted genetic diagnostic strategies and also impact on clinical management.

Hypogonadotropic hypogonadism due to variants in RAB3GAP2: expanding the phenotypic and genotypic spectrum of Martsolf syndrome.

Biallelic pathogenic variants in RAB3GAP2 cause Warburg Micro syndrome (WARBM) and Martsolf syndrome (MS), two rare, phenotypically overlapping disorders characterized by congenital cataracts, intellectual disability, and hypogonadism. Although the initial report documented hypergonadotropic hypogonadism (implying a gonadal defect), an adolescent girl with WARBM/MS was subsequently reported to have hypogonadotropic hypogonadism (implying a central defect in either the hypothalamus or anterior pituitary). However, in adult MS, hypogonadotropism has not been convincingly demonstrated. Additionally, the correlation between the pathogenic severity of variants in RAB3GAP2 and the phenotypic severity also remains unclear. Here we present a clinical report of a woman with congenital cataracts, apparent intellectual disability, and pubertal failure who underwent exome sequencing (ES) to determine a precise molecular diagnosis. Reproductive phenotypes reported previously in individuals with MS and the genotypic spectrum of previous RAB3GAP2 variants were also reviewed. The ES identified pathogenic compound heterozygous RAB3GAP2 variants (c.387-2A > G; p.(Arg428Glu)) combined with her phenotypic features, which enabled a unifying molecular diagnosis of MS. Reproductive evaluation confirmed a normosmic idiopathic hypogonadotropic hypogonadism. Review of the RAB3GAP2 allelic spectrum in WARBM/MS suggests that although variants resulting in complete abrogation of RAB3GAP2 protein function cause severe WARBM, variants associated with partially preserved RAB3GAP2 function cause milder MS. This report expands the genotypic and phenotypic spectrum of MS and demonstrates hypogonadotropic hypogonadism as a key pathophysiologic abnormality in MS. Genotype-phenotype associations of previously reported RAB3GAP2 variants indicate that variants that fully abolish RAB3GAP2 function result in WARBM, whereas MS is associated with variants of lesser severity with residual RAB3GAP2 function.