Abstracts

Rationale: Tuberous sclerosis complex (TSC) is a genetic disorder associated with neurological, renal, dermatological and other anomalies. Over 85% of reported patients have germline mutations in TSC1 or TSC2, with a higher yield of pathogenic variants in cohorts with more severe, earlier onset disease. Some patients appear to have “de novo” variants on routine testing of parental blood-derived DNA, yet one parent may have clinical features of TSC or multiple affected children carrying the same germline mutation, indicating the likelihood of low-level parental mosaicism. We aimed to identify parental mosaicism in 7 families with TSC and features suggesting underlying parental mosaicism.

Methods: Seven families had known pathogenic germline mutations previously identified in the probands on clinical genetic testing. All parents were negative for their child’s variant on segregation analysis via clinical Sanger sequencing. Blood, saliva, buccal and urine DNA were obtained from all parents. Droplet digital PCR (ddPCR) or deep targeted amplicon sequencing (5,000x) were used to detect and quantitate the variants in parental tissues.

Results: We found parental mosaicism in blood-derived DNA in five of 7 families at low frequency ranging from 0.1-8.8%, all well below the threshold of detection by Sanger sequencing. In 3 parents with mosaic variants, we determined variant allele frequency (VAF) in different tissues. One parent had mosaicism at a similar level across four different tissues (7.26-9.11% VAF); while in the other two parents, mosaicism was variable (0.9-3.12% VAF across three tissues and 0.39-2.2% VAF across four tissues, respectively). Of the two families without mosaicism detected, one had two affected children carrying the same germline mutation so one parent must have gonadal mosaicism that we could not detect in peripheral tissues. In the second negative family, the mother has unilateral renal hamartoma without any other clinical features of TSC; so, her hamartoma may not be due to TSC.

Conclusions: Our findings confirm that low-level parental mosaicism missed on routine clinical testing can be detected with high levels of coverage. This finding has critical implications for reproductive counseling.

Funding: Please list any funding that was received in support of this abstract.: This study was supported by a Sanming Project of Medicine in Shenzhen, China (SZSM201812005) to J.L. and I.E.S., a Shenzhen Key Medical Discipline Construction Fund (SZXK033) to J.L., a Shenzhen Fund for Guangdong Provincial High-level Clinical Key Specialties (SZGSP012) to F.W., an Australian National Health and Medical Research Council Program Grant (1091593) to I.E.S. and S.F.B., a Project Grant (1129054) to S.F.B., a Project Grant (1079058) to M.S.H., a Practitioner Fellowship (1006110) to I.E.S., and a R.D Wright Career Development Fellowship (1063799) to M.S.H.