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Background: Structural congenital anomalies like orofacial clefts (OFCs) occur prenatally. OFCs mostly present as cleft lip (CL), cleft palate (CP) or both, with incidence varying from 1/500 to 1/2500 births depending on human ancestral populations. Affected individuals suffer from severe feeding, speech, middle ear infections, and dental challenges. Genetics and environmental risk factors have been implicated in the aetiology of the disease. The current study deciphered the genetic architecture of twins concordant for CL and prenatal teeth.
Methods: DNA from affected twins and mother underwent whole-exome sequencing (WES) on the Illumina HiSeq platform at 100X. Bioinformatics analysis of WES data utilised the Sentione workflow. Normalised VCF files were annotated with Ensembl VEP. Variants were classified and prioritised using American College of Medical Genetics and Genomics (ACMG) guidelines on variant classifications. Suspected pathogenic variants for CL-prenatal teeth underwent further segregation analysis in all 13 family members using Sanger sequencing. The zygosity of the twins was computed using the Pandas package in Python 3. Clinically actionable secondary findings (SF) were ascertained using the ACMG guidelines on SFs.
Results: The two affected male twins presented with CL and prenatal teeth, though there was no family history of this. The zygosity of the twins, computed based on variant concordance rate, was 99.37%, suggesting monozygotic twins. Potentially causative variants that segregated with the disease were observed in genes such as AMER2 (c.604C>G, p.Arg202Gly), SMOC2 (c.454C>T, p.Arg152Trp), and NOTCH4 (c.1577C>T, p.Ala526Val). Variants observed in ACMG SF genes included MSH6 (c.3911G>A, p.Arg1304Lys), SCN5A (c.5549C>T, p.Ser1850Leu), TP53 (c.935C>G, p.Thr312Ser), DSG2 (c.2368C>T, p.His790Tyr), RBM20 (c.1816G>A, p.Val606Met) and RYR1 (c.13550C>T, p.Pro4517Leu).
Conclusion: Monozygotic twins concordant for rare conditions enable identification of aetiologic risk variants. This approach helps to identify causative variants for OFCs and related phenotypes, alongside clinically actionable SFs. These observations significantly impact pathophysiology, genetic counselling and personalised medicine.