Side-chain cleavage enzyme deficiency: Systematic review and case series.

OBJECTIVE: P450 side-chain cleavage deficiency (SCCD) patients present with primary adrenal insufficiency (PAI) with or without undervirilized external genitalia. The distinction between classic and nonclassic steroidogenic acute regulatory protein deficiency has been described, whereas in SCCD is unclear. The data on gonadal function and its correlation with SCCD genotype has not been studied. We describe our experience and perform a systematic review of genetically proven SCCD patients to determine the distinct phenotypic and genotypic characteristics of 46,XY SCCD patients with typical male external genitalia (SCCD-TMG) and atypical (SCCD-AG) external genitalia. DESIGN, PATIENTS AND MEASUREMENTS: Retrospective review of three genetically proven SCCD patients from our centre and per-patient data analysis from a systematic review of 52 probands was performed. SCCD-TMG (n = 19) was defined as external genitalia of Sinnecker score 1 with 46,XY  karyotype; the rest (Sinnecker 2-5) were classified as SCCD-AG (n = 15). RESULTS: We report two new Indian cases of SCCD with three novel likely pathogenic variants and pubertal follow-up of a previously reported patient. In systematic review, age at diagnosis of PAI and elevated renin were not different between 46,XY  SCCD-TMG (n = 19) and SCCD-AG (n = 15), whereas spontaneous puberty (9/9 vs. 0/3, p = .0045), normal prepubertal (5/5 vs. 6/6, p = .002), pubertal gonadotropins (2/9 vs. 0/3, p = 1) and normal pubertal testosterone (9/11 vs. 0/3, p = .027) were more common in SCCD-TMG. Testicular adrenal rest tumours were exclusive to SCCD-TMG (n = 4). SCCD-TMG was associated with four particular genotypes [monoallelic p.Glu314Lys with another deleterious variant on the second allele (p.Glu314Lys/X-CHS: X-compound heterozygous state), biallelic p.Arg451Trp, p.Phe215Ser/p.Arg232Ter and monoallelic p.Val79Ile]. 46,XX SCCD  patients with p.Glu314Lys/X-CHS also had normal gonadotropins with spontaneous puberty. CONCLUSION: SCCD-TMG is associated with four specific genotypes and distinct gonadal characteristics from SCCD-AG with overlapping features of PAI.

Computational study of interactions of anti-cancer drug eribulin with human tubulin isotypes.

Microtubules (MTs) are widely targeted for the treatment of various types of cancer due to their essential role in cell division. MTs are polymers made of αß-tubulin heterodimers. These α- and ß-tubulins have 8 and 10 different isotypes, respectively. It is known that a few tubulin isotypes have anti-cancer drug resistance properties, especially ßIII, which shows poor sensitivity to many potent anti-cancer drugs such as eribulin. However, the molecular-level understanding of drug-resistance due to tubulin isotype variation is poorly understood. This paper presents the study of differential binding affinities of different tubulin isotypes with the potent anti-cancer drug eribulin. Eribulin (MT destabilizer) binds at the inter-dimer interface of MTs near the vinca site and induces a lattice deformation, which results in catastrophic events in MT dynamics. In this study, sequence analysis has been done throughway and the binding sites and based on that 2α-tubulin isotypes (αI and αVIII) and 7ß tubulin isotypes (ßI, ßIIa, ßIII, ßIVa, ßVI, ßVII and ßVIII) were selected. In total, 14 combinations were prepared after building homology models of these selected isotypes. Molecular docking and molecular dynamics simulations were performed to deeply understand the binding mode of eribulin at different MT compositions. RMSD, RMSF, radius of gyration, SASA, ligand-protein interactions, and calculations of binding free energy were performed to investigate the eribulin binding variations to tubulin isotypes and it was found that αIßII showed the maximum binding affinity among all 14 systems to eribulin. The ßIII-tubulin isotype, which shows low sensitivity to eribulin in experimental results, had the least binding affinity in the system αVIIIßIII complex and the average binding affinity in the system αIßIII among all 14 systems. Additionally, we performed steered MD simulations and DynDom analysis of the systems with the lowest binding energy (αIßII) and the highest binding energy (αVIIIßIII) and extracted force, displacement, and H-bonding profiles during the pulling simulations to get a better insight.

Expanding genetic spectrum and discriminatory role of steroid profiling by LC-MS/MS in 11ß-hydroxylase deficiency.

OBJECTIVE: To report clinical, hormonal and structural effects of CYP11B1 pathogenic variations in Indian patients with 11ß-hydroxylase deficiency (11ßOHD) and find hormonal criteria that accurately distinguish 11ßOHD from 21α-hydroxylase deficiency (21OHD). DESIGN: Retrospective record review of genetically diagnosed patients with 11ßOHD. PATIENTS AND MEASUREMENTS: Clinical features, hormonal parameters at diagnosis (by immunoassay) and recent follow-up of 13 genetically proven 11ßOHD patients managed at our centre were retrospectively reviewed. ACTH-stimulated serum adrenal steroids (measured by LC-MS/MS) of 11ßOHD were compared with those of simple virilizing and non-classic 21OHD. Structural analysis of the observed pathogenic variations was performed by computational modelling. RESULTS: Nine (four females) and four (all females) patients had classic and non-classic disease, respectively. All 11ßOHD patients had elevated ACTH-stimulated serum 11-deoxycortisol (26.5-342.7 nmol/L) whereas none had elevated serum 17-hydroxyprogesterone (4.2-21.2 nmol/L); both hormonal parameters distinguished 11ßOHD from 21OHD with 100% accuracy. ACTH-stimulated serum cortisol, but not 11-deoxycortisol, clearly distinguished classic (<70 nmol/L) from non-classic (>160 nmol/L) disease. Thirteen (eight novel, two recurrent) pathogenic variants were observed. Only missense mutations were observed among patients with non-classic disease. Computational modelling predicted the possible affection of enzyme structure and function for all the observed missense mutations. CONCLUSIONS: This first Indian study describes 13 11ßOHD patients, including four with the rarer non-classic variant. A total of eight novel pathogenic variants were identified in our study, highlighting regional genetic heterogeneity. Measurement of ACTH-stimulated adrenal steroids by LC-MS/MS will help avoid the misdiagnosis of 11ßOHD as 21OHD and has potential to distinguish classic from non-classic 11ßOHD.

17α-Hydroxylase/17,20-Lyase Deficiency in 46,XY: Our Experience and Review of Literature.

CONTEXT: There are more than 100 pathogenic variants in CYP17A1 that have been identified in patients with 17α-hydroxylase/17,20-lyase deficiency (17OHD). OBJECTIVE: We aimed to describe 46,XY patients with 17OHD from our center and review the literature. METHODS: We retrospectively analyzed genetically proven index cases of 17OHD from our 46,XY disorders of sex development cohort and reviewed similar cases from the literature (n = 150). Based on the phenotype, 17OHD probands were classified into combined severe deficiency (n = 128) and combined partial deficiency (n = 16). Additionally, patients with the apparent isolated 17,20-lyase deficiency (n = 7, from 6 families) were noted. Residual enzyme activities with the observed mutant enzymes were divided in 2 categories as < 1% and ≥ 1%, each for hydroxylase and lyase. RESULTS: We present 4 index cases of 46,XY 17OHD with a complete spectrum of undervirilization and 2 novel variants in CYP17A1. In the review, the combined severe deficiency was the most common form, with more frequent female sex of rearing, hypertension, hypokalemia, suppressed renin, higher plasma corticotropin, lower serum cortisol, and androgens. Immunoassay-measured serum aldosterone was frequently (68.2%) unsuppressed (>5 ng/dL). Elevated serum progesterone had high sensitivity for diagnosis of combined 17OHD, even in combined partial deficiency (83.3%). Among patients with clinical phenotype of combined severe deficiency, 11.5% had partial 17α-hydroxylase and complete 17,20-lyase deficiency (>1%/<1%) and had significantly higher serum cortisol than those with < 1%/<1% activity. CONCLUSION: We report the first monocentric case series of Asian Indian 46,XY patients with 17OHD. We propose that a phenotype of severe undervirilization with milder cortisol deficiency may represent a distinct subtype of combined severe 17OHD with residual 17α-hydroxylase activity but severe 17,20-lyase deficiency (>1%/<1%), which needs further validation.

Homozygous p.Val89Leu plays an important pathogenic role in 5α-reductase type 2 deficiency patients with homozygous p.Arg246Gln in SRD5A2.

OBJECTIVE: To evaluate the pathogenic role of a few benign variants and hypomorphic pathogenic variants in SRD5A2. DESIGN AND METHODS: We retrospectively analyzed phenotypes and genotypes in 23 Indian patients with genetically proven steroid 5α-reductase 2 (SRD5A2) deficiency. The interactions of the SRD5A2 enzymes resulting due to the most common benign variant (p.Val89Leu), the most common (hypomorphic) pathogenic variant (p.Arg246Gln) and the double variants (p.Val89Leu and p.Arg246Gln) in SRD5A2 were compared with that of the wild type (WT) enzyme by molecular dynamics (MD) simulation. RESULTS: The majority (n = 19, 82.61%) of patients presented for atypical genitalia and had male gender identity (16/20). Including the two novel ones (p.Leu83Pro, p.Ala28Leufs*103), a total of nine different pathogenic variants were observed. p.Arg246Gln was the most common pathogenic variant (n = 12). Homozygous p.Arg246Gln (n = 9) variant was associated with milder undervirilization (Sinnecker score of ≤3a: 8/9 vs 6/14, P = 0.04) and had concurrent homozygous p.Val89Leu in all patients. Interestingly, asymptomatic fathers of two index patients were homozygous for p.Arg246Gln which questioned the pathogenicity of the variation as a sole factor. Unlike all symptomatic homozygous p.Arg246Gln patients who were also homozygous for p.Val89Leu, asymptomatic homozygous p.Arg246Gln fathers were heterozygous for p.Val89Leu. On MD simulation SRD5A2 p.Val89Leu-Testeosterone (T) and SRD5A2 p.Arg246Gln-T complexes, but not SRD5A2 p.Val89Leu and p.Arg246Gln-T complex, demonstrated close interaction between NADPH and T as that of SRD5A2 WT-T. CONCLUSIONS: p.Arg246Gln may not be pathogenic as a sole variation even in the homozygous state; additional contribution of homozygous p.Val89Leu variant may be essential for the pathogenicity of p.Arg246Gln in SRD5A2.