Twins With Pathogenic RNF113A Variant Presenting With Testicular Regression Syndrome

Abstract

RNF113A-related disorders are rare X-linked conditions characterized by neurodevelopmental impairment, endocrine abnormalities, and gonadal dysgenesis. To date, this condition has been documented in only a few individuals. We report 46,XY twin siblings with a hemizygous pathogenic RNF113A variant [c.901C > T, p.(Gln301*)], presenting with a remarkably consistent phenotype that includes microcephaly, corpus callosum dysgenesis, intractable epilepsy, subclinical hypothyroidism, microphallus, and bilateral testicular regression syndrome (TRS). Whole-exome sequencing identified a maternally inherited variant consistent with X-linked recessive inheritance. Despite the absence of testicular tissue, both twins had evidence of prenatal androgen exposure. Thyroid dysfunction evolved during infancy. These cases expand the phenotypic spectrum associated with pathogenic RNF113A variant and provide evidence linking this gene to TRS. They further highlight the importance of whole-exome sequencing in evaluating complex 46,XY disorders of sex development with multisystem involvement. RNF113A should be considered in the genetic evaluation of 46,XY disorder of sex development and TRS, particularly when accompanied with multisystem involvement.

Introduction

Disorders of sex development (DSD) (also referred to as differences of sex development) comprise a heterogeneous group of conditions involving atypical development of chromosomal, gonadal, or anatomical sex. In individuals with a 46,XY karyotype, the complete absence or severe dysgenesis of the testes is rare [1]. Testicular regression syndrome (TRS) occurs in ∼1 in 20 000 newborn males, it accounts for 0.5% to 4.5% of cryptorchidism cases. Bilateral TRS is characterized by undetectable testosterone, inhibin B, and INSL3, elevated gonadotropins, and a 46,XY karyotype.

The etiology of TRS is unclear, though vascular events such as torsion or thrombosis have been proposed. The timing of regression influences the phenotype: loss of testicular function in mid-gestation leads to microphallus in ∼50% of affected males, whereas near-term regression typically results in normal penile length [2].

A genetic basis for TRS has not been well established, although variants in DHX37 have been suggested in rare cases [3]. RNF113A, an X-linked gene involved in RNA splicing and DNA repair, has recently been linked to a syndromic disorder featuring neurodevelopmental delay and endocrine abnormalities, including TRS and microphallus, though only a few patients have been reported [4-6].

We describe twin siblings with a pathogenic RNF113A variant, both presenting with TRS and microphallus further supporting its potential role for RNF113A in prenatal testicular development.

Case Presentation

46,XY twins were born at 36 + 4/7 weeks via cesarean section. Noninvasive prenatal testing reported monozygotic twins. Birth weights were 2.16 kg (6th percentile; z = −1.56) and 2.01 kg (3rd percentile; z = −1.87), with head circumferences of 32.0 cm (18th percentile; z = −0.92) and 34.0 cm (60th percentile; z = 0.25), respectively.

The twins were born to White nonconsanguineous parents. The mother is 157.5 cm (5′2″; 10th percentile, z = −1.3) and the father is 170 cm (5′7″; 15th percentile, z = −1.0) in height. They have 1 unaffected older sister, age 6 years, with no known endocrine, neurologic, or developmental abnormalities.

Both twins were discharged but readmitted to another hospital at 9 days of life for failure to thrive and hypothermia. At that time, weights were 1.81 kg and 1.80 kg (<1st percentile; z = −3), and head circumferences measured 30.0 cm each (<1st percentile; z = −2.7). Physical examination revealed bilateral nonpalpable testes and microphallus.

Diagnostic Assessment

On examination, both twins had microcephaly and atypical genitalia including microphallus, bilateral nonpalpable testes, labioscrotal fusion, and a single perineal opening (Fig. 1A). Twin B had associated chordee. Stretched penile length at 6 weeks were 1.25 cm (<3rd percentile) and 1.7 cm (<3rd percentile), respectively.

Figure 1.

(A) Twin A at 3 months of age, showing microphallus and absent scrotal sac. (B) Twin B brain MRI demonstrating a hypoplastic, dysplastic corpus callosum (white arrow).

Laboratory evaluations indicated vanished testes, with elevated FSH, low testosterone, and undetectable anti-müllerian hormone and inhibin B levels (Table 1). Ultrasounds at 2 weeks of age (outside hospital) suggested right inguinal testis for twin A and the left not visualized. Twin B appeared to have both testes in the inguinal canals. By 1 month of age, ultrasound at our center showed no detectable testicular tissue in Twin A, and left inguinal testis with the right not visualized in Twin B. At 1 year, neither twin had identifiable testicular tissue.

Table 1.

Endocrine laboratory results at first clinic visit (5 weeks old)^a

	Twin A	Twin B	Normal range
FSH	58.0 (58.0) mIU/mL (IU/L)	50.8 (50.8) mIU/mL (IU/L)	0.00-1.52 mIU/mL (0.00-1.52 IU/L)
LH	5.7 (5.7) mIU/mL (IU/L)	3.7 (3.7) mIU/mL (IU/L)	0.00-1.18 mIU/mL (0.00-1.18 IU/L)
Testosterone	<2.5 (<0.09) ng/dL (nmol/L)	2.9 (0.10) ng/dL (nmol/L)	60-400 ng/dL (2.1-13.9 nmol/L)
Inhibin B	<10.0 (<10) pg/mL (ng/L)	<10.0 (<10) pg/mL (ng/L)	42-516 pg/mL (42-516 ng/L)
AMH	0.01 (0.07) ng/mL (pmol/L)	0.01 (0.07) ng/mL (pmol/L)	23.8-124.0 ng/mL (170-884 pmol/L)
TSH	6.6 (6.6) µIU/mL (mIU/L)	7.8 (7.8) µIU/mL (mIU/L)	1.00-4.90 µIU/mL (1.00-4.90 mIU/L)
FT4	1.1 (14.2) ng/dL (pmol/L)	1.2 (15.5) ng/dL (pmol/L)	0.69-1.56 ng/dL (8.9-20.1 pmol/L)

Abbreviations: AMH, anti-müllerian hormone; FT4, free thyroxine.

^a Samples for Inhibin B and AMH were obtained at 3 weeks old.

Although early ultrasound scans reported testes within the inguinal canals, the discrepancy with hormonal studies and later imaging suggests that the reliability of the initial findings may have been limited.

Both twins exhibited complete labioscrotal fusion and a phallic structure with a single perineal opening, consistent with prenatal androgen exposure. These findings support a diagnosis of TRS rather than primary testicular agenesis.

Both twins demonstrated evolving thyroid dysfunction during infancy, prompting close monitoring. The newborn screening (NBS) program is TSH and total thyroxine based. Twin A’s NBS showed TSH 23.6 µIU/mL (23.6 mIU/L) (normal below 25 µIU/mL [25 mIU/L]) and total thyroxine 9.4 µg/dL (121.3 nmol/L) (normal 5.1-30 µg/dL [65.6-386.1 nmol/L]). At 1 month, serum TSH was 10.2 µIU/mL (10.2 mIU/L) (normal range 1-30 days; 1-9.1 µIU/mL [1-9.1 mIU/L]) with a free thyroxine (FT4) 1.26 ng/dL (16.3 pmol/L) (normal range 1-30 days; 0.83-3.09 ng/dL [10.7-39.8 pmol/L]). Monthly thyroid function tests showed a gradual decrease in TSH to 6.8 µIU/mL (6.8 mIU/L) (normal range 1-11 months; 0.8-8.2 µIU/mL [0.8-8.2 mIU/L]), followed by a rise to 9.6 µIU/mL (9.6 mIU/L) at 4 months, with FT4 trending down to 0.89 ng/dL (11.5 pmol/L) (normal range 1-11 months; 0.48-2.32 ng/dL [6.2-29.9 pmol/L]).

Twin B’s NBS showed TSH 27.4 µIU/mL (27.4 mIU/L) and total thyroxine 8.6 µg/dL (111.0 nmol/L). At 1 month, serum TSH was 7.8 µIU/mL (7.8 mIU/L) with a FT4 1.17 ng/dL (15.1 pmol/L). By 6 months of age, Twin B’s FT4 was 0.79 ng/dL (10.2 pmol/L) and TSH 6.48 µIU/mL (6.48 mIU/L).

Both twins developed intractable epilepsy by 4 months. Brain MRI revealed significant anomalies, Twin A: callosal dysgenesis, underdeveloped frontal lobes and horns, partial absence of the septum pellucidum, and a mild Dandy-Walker complex. The pituitary bright spot appeared slightly small on precontrast imaging. Twin B: hypoplastic and dysplastic corpus callosum with absent splenium. No pituitary abnormalities were observed (Fig. 1B).

At 5 months, Twin A developed transient thrombocytopenia and neutropenia during a seizure-related admission. Bone marrow aspiration and flow cytometry ruled out malignancy. Counts normalized, suggesting medication-related or transient viral suppression.

Cosyntropin stimulation (15 mcg/kg IV) confirmed intact adrenal reserve in both twins, with peak cortisol level of 27 and 33 µg/dL (745 and 910 nmol/L) (institutional cutoff ≥15 µg/dL [≥415 nmol/L]), respectively.

Because the biochemical profile supported primary end-organ dysfunction evidenced by elevated TSH with declining FT4, elevated FSH and LH with undetectable anti-müllerian hormone and inhibin B, and adrenal function was intact, we did not obtain further pituitary evaluation of insulin-like growth factor 1 (IGF-1) and insulin-like growth factor binding protein 3 (IGF-BP3) or prolactin in infancy. These assays have limited sensitivity in early infancy; we plan longitudinal auxologic surveillance with growth hormone (GH)-axis testing if growth velocity remains subnormal.

No structural cardiac abnormalities were identified on echocardiograms. Abdominal ultrasound findings were unremarkable, ruling out renal or visceral anomalies.

Prior genetic testing, including karyotype, chromosomal microarray, and targeted gene panels, was nondiagnostic. Twin B underwent an Invitae DSD panel, which identified a heterozygous likely benign WT1 variant, consistent with a known population polymorphism. An Invitae epilepsy panel performed on both twins revealed heterozygous variants of uncertain significance in ADAR (c.1765A > G, p.Thr589Ala) and RELN (c.4437T > A, p.Asp1479Glu). Inheritance of RELN variants can be autosomal dominant so parental testing was performed. The father carried the same variant but was clinically unaffected, supporting a likely benign classification. Trio whole-exome sequencing of both twins and their parents identified a maternally inherited hemizygous pathogenic variant in RNF113A [c.901C > T, p.(Gln301*)], consistent with X-linked inheritance.

Treatment

Both twins received 3 monthly intramuscular testosterone injections (25 mg each) starting at 3 months of age to mimic minipuberty of infancy. This resulted in phallic growth, with stretched penile length increasing from 1.25 cm (<3rd percentile) and 1.7 cm (<3rd percentile) at 6 weeks to 3.0 cm (25th percentile) and 2.5 cm (10th percentile), respectively, by 7 months.

Although the biochemical pattern suggested subclinical hypothyroidism, the decision to initiate therapy at 4 and 6 months, respectively, was influenced by concern for neurodevelopmental vulnerability in the setting of their structural brain anomalies and seizures. For Twin A, levothyroxine 37.5 mcg daily was initiated, leading to improvement in TSH to 2.16 µIU/mL (2.16 mIU/L) and FT4 of 0.99 ng/dL (12.8 pmol/L). For Twin B, levothyroxine 25 mcg daily was started, leading to improvement in TSH to 2.60 µIU/mL (2.60 mIU/L) and FT4 of 1.02 ng/dL (13.2 pmol/L).

Both twins required multiple antiseizure medications.

Outcome and Follow-Up

At the latest follow-up, at 18 months, both twins continue to have moderate global developmental delay and intractable epilepsy, managed with multiple antiepileptic medications. Thyroid function remains stable on levothyroxine at 37.5 mcg daily for Twin A and 25 mcg daily for Twin B. Neither twin has developed adrenal insufficiency to date. Growth velocity from 12 to 18 months was appropriate (6-8 cm), but no catch-up growth has occurred; weight, length, and head circumference all remain <1st percentile.

Discussion

This case of twin siblings with a shared pathogenic RNF113A variant provides compelling evidence for the gene's role in prenatal development of both the central nervous system and endocrine organs, particularly in testicular development. Both twins exhibited an almost identical phenotype: microcephaly, corpus callosum dysgenesis, intractable epilepsy, thyroid dysfunction, microphallus, and TRS. This remarkable consistency strengthens the argument for a causal relationship between RNF113A dysfunction and this constellation of clinical features. The presence of TRS, microphallus, and thyroid dysfunction alongside severe neurodevelopmental anomalies highlights the potential impact of this gene on multiple organ systems during fetal life.

RNF113A encodes a zinc finger protein involved in RNA splicing and DNA repair. Loss-of-function mutations are associated with oxidative stress and NRF2 pathway activation [7]. Studies in mice have identified Rnf113a1 as a regulator of neural stem cell survival, proliferation, and differentiation, supporting its involvement in brain development [8].

Only a few postnatal cases of RNF113A-related disease have been described; their key clinical features are summarized in Table 2 [4-6].

Table 2.

Phenotypic comparison of current twin cases with previously reported individuals with RNF113A variants

Clinical features	Corbett P1 [6]	Corbett P2 [6]	Tessarech fetus 1 [5]	Tessarech fetus 2 [5]	Mendelsohn P1 [4]	Mendelsohn P2 [4]	Present case: twin A	Present case: twin B
Genetic mutation	p.(Gln301*)	p.(Gln301*)	p.(Gln301*)	p.(Gln301*)	p.(Gln302fs*12)	p.(Cys299*)	p.(Gln301*)	p.(Gln301*)
Gestational age	36 WG, SGA	38 WG, SGA	TOP at 34 WG, IUGR	TOP at 22 WG	38 WG	Term	36 + 4 WG, SGA	36 + 4 WG, SGA
Birth parameters (weight and HC)	2.18 kg/31 cm	2.32 kg/29 cm	NA	NA	2.30 kg/NA	1.90 KG/NA	2.16 kg/32 cm	2.01 kg/34 cm
Craniofacial Abnormalities	Microcephaly, high forehead, broad mouth, widely spaced decidual teeth, prominent chin	Microcephaly, high forehead, sparse hair, absent outer eyebrows, macrostomia, widely spaced teeth	Microcephaly, receding forehead, broad nasal root, epicanthus, retrognathia, ear deformity	Microcephaly, receding forehead, broad nasal root, epicanthus, retrognathia, ear deformity	Dolichocephaly, prominent forehead, high anterior hairline, sparse eyebrows, downslanting palpebral fissures, defined philtrum, thin upper lip	Triangular face, broad forehead, wide mouth, absent eyelashes and eyebrows	Microcephaly, low-set ears with ear pits	Microcephaly, bitemporal narrowing, low-set ears with ear pit, right eye ptosis
Genital abnormalities	Microphallus, absent testes	Absent/rudimentary testes	Microphallus, agenesis of right testis, rudimentary left testis	Microphallus, agenesis of right testis, rudimentary left testis	Microphallus, absent testes	Microphallus, absent testes, hypospadias, bifid scrotum	Microphallus, absent testes	Microphallus, absent testes
CNS abnormalities	Seizures Cerebellar hypoplasia, cerebral atrophy, partial agenesis of corpus callosum, Dandy-Walker complex	Seizures Cerebellar hypoplasia, partial agenesis of corpus callosum, Dandy-Walker complex	Partial agenesis of corpus callosum, absent Probst bundles	Partial agenesis of corpus callosum, absent Probst bundles	Seizures Thin corpus callosum	Seizures Absent corpus callosum	Seizures Callosal dysgenesis, underdeveloped frontal lobes and horns, partial absence of septum pellucidum, mild Dandy-Walker complex	Seizures Hypoplastic/dysplastic corpus callosum, reduced fronto-occipital distance, underdeveloped frontal horns
Endocrine abnormalities	↑ TSH → GH	↓ T4, ↑ TSH, ↑ LH, FSH ↓ Testosterone ↓ ADH and cortisol	Adrenal hypoplasia with cortical hypoplasia	Adrenal hypoplasia with cortical hypoplasia	↑ TSH, ↓→T4 ↑ LH, FSH, and prolactin ↓ Testosterone ↓ IGF-1, IGFBP-3, and cortisol	→ TSH, T4, ACTH, and cortisol	↑ TSH, ↓ FT4 ↑ LH, FSH ↓ Testosterone	↑ TSH, ↓ FT4 ↑ LH, FSH ↓ Testosterone
Other findings	Very short esophagus, ureteric obstruction, recurrent infections, cutis marmorata	Recurrent infections, chronic diarrhea, eczema, cutis marmorata	Bilateral sandal gap, 11 ribs, renal cortical cysts	Short first toes, 11 rib pairs	Frequent infections, low IgG1, duodenal strictures	Frequent infections, multicystic dysplastic kidney	Transient cytopenias, normal abdominal ultrasound	Normal abdominal ultrasound

Abbreviations: ADH, antidiuretic hormone; FT4, free thyroxine; HC, head circumference; IGFBP-3, insulin-like growth factor binding protein 3; IUGR, intrauterine growth restriction; SGA, small for gestational age; TOP, termination of pregnancy; US, ultrasound; WG, weeks of gestation.

Our twin cases share many of these previously reported features and add valuable reinforcement to the phenotypic pattern, particularly through the consistent presence of TRS. Our patients have not yet demonstrated previously reported features such as adrenal insufficiency, recurrent infections, or renal anomalies. Transient cytopenias in Twin A may represent an underrecognized feature or medication-related bone marrow suppression.

Despite the widespread impact of RNF113A variants on multiple organ systems, none of the published postnatal or fetal cases, including our own, has demonstrated cardiac anomalies [4-6]. Given its broad developmental role, the consistent absence of cardiac findings across all reports may suggest relative sparing of early cardiac morphogenesis or the presence of compensatory pathways during cardiogenesis. Additional cases are needed to confirm whether this is a true pattern or sampling limitation.

The [c.901C > T, p.(Gln301*)] variant identified results in a premature stop codon, consistent with loss-of-function mechanisms reported previously. The pattern of X-linked recessive inheritance is also consistent with prior observations [4-6].

Interestingly, short maternal stature has been noted in all prior reports of maternally inherited RNF113A variants, without syndromic features: 147 cm (Mendelsohn [4]), 142 and 151 cm (Corbett [6]), and 160 cm (Tessarech [5]). In our report, the mother is asymptomatic with a height of 157.5 cm, again slightly below population average.

Our report is notable for presenting affected twins with genetically confirmed RNF113A-related disorder and offers compelling support for RNF113A as a novel gene implicated in TRS. The consistency of the phenotype between these twins, and across published cases, argues strongly against environmental or isolated vascular causes for TRS in these instances. We propose that RNF113A be considered a candidate gene in the genetic evaluation of 46,XY DSD, particularly in cases presenting with TRS, microphallus, and neurodevelopmental anomalies.

This case provides unique insight into the phenotypic reproducibility of RNF113A-related disease within genetically similar individuals. Longitudinal follow-up will be essential to track cognitive development, endocrine function, and growth, and to better understand the natural history of this emerging syndrome. Further research is needed to elucidate the molecular mechanisms by which RNF113A influences these embryonic developmental pathways.

Learning Points

• RNF113A-related disorders can present with a distinctive combination of endocrine, neurologic, and urogenital anomalies.

• Consistent phenotypes across affected twins strengthen evidence for a causal role of RNF113A in prenatal gonadal development.

• Whole-exome sequencing is essential in evaluating complex 46,XY DSD with multisystem involvement.

Contributors

All authors made individual contributions to authorship. O.R., G.B.: participated in the diagnosis and management of the patients, literature review, and manuscript preparation. O.R.: primary manuscript drafting, table and figure preparation. G.B.: critical revision of the manuscript. All authors reviewed and approved the final draft.

Abbreviations

DSD

disorders of sex development

FSH

follicle-stimulating hormone

FT4

free thyroxine

GH

growth hormone

IGF-1

insulin-like growth factor 1

IGF-BP3

insulin-like growth factor binding protein 3

LH

luteinizing hormone

MRI

magnetic resonance imaging

NBS

newborn screen

SPL

stretched penile lengths

TRS

testicular regression syndrome

TSH

thyroid-stimulating hormone

Funding

No public or commercial funding.

Disclosures

None declared.

Informed Patient Consent For Publication

Signed informed consent obtained directly from the patients' relatives or guardians.

Data Availability Statement

Original data generated and analyzed for this case report are included in this published article.