Tuberous sclerosis complex (TSC) is an autosomal-dominant genetic disease characterized by the development of tumors in the brain, kidney, lung, skin, heart, and retina, as well as neurodevelopmental syndromes known as tuberous sclerosis complex-associated neurologic disorder.1,2 The incidence of TSC is estimated to be 1/6000 to 1/10,000 among live births, and around 1/20,000 in the population.3 Two causative genes, TSC1 (online Mendelian inheritance in man #191100) and TSC2 (online Mendelian inheritance in man #613254) have been discovered. Hence, genetic testing has been recommended as a diagnostic criterion in clinical practice.3,4
A high variability in the phenotype and disease progression have been described among TSC patients, and even within families.5 Multiple cardiac rhabdomyo-mas identified by fetal ultrasound are highly supportive of a prenatal diagnosis of TSC.6,7 However monozygotic twins with TSC are rarely reported prenatally. Here, we report two cases of monochorionic diamniotic twins diagnosed prenatally as having TSC and discordant fetal phenotypes. This is the first description of a phenotypic discrepancy among monozygotic twins. The study protocol was approved by the ethics committee of the Tongji University in Shanghai, China (Research Ethics board registration number: 2018yxy27, December 24, 2018). The parents of both families gave their consent for description and publication of the data of their fetuses.
A healthy 31-year-old woman, who experienced three spontaneous miscarriages in the first trimester previously, received in vitro fertilization with sperm donation and embryo transfer. One embryo was transferred and mono-chorionic diamniotic twins were confirmed by ultrasound at 6 weeks of gestation. Both fetuses had normal nuchal translucency (1.2 mm/1.5 mm) and normal growth parameters according to ultrasound screening at weeks 7,11, and 18 of gestation. An anatomy scan at 21 weeks and 6 days revealed a hyperechogenic mass (9 mm × 8 mm) in the apex of the left ventricle in one fetus, whereas the other fetus was not affected. Therefore, she was referred to a fetal medicine center for a precise diagnosis.
The fetal echocardiogram at 23 weeks and 5 days confirmed that one fetus was affected by multiple rhabdomyomas in the apex of the left ventricle, with the largest lesion being of dimension 11.8 mm × 0.9 mm × 11.8 mm. Nodules were also identified in the fetal brain. Four weeks later, the rhabdomyomas were extended to the left-ventricular lateral wall. The largest cardiac rhabdo-myoma developed to a dimension of 14.5 mm × 11.8 mm × 11.8 mm (Fig. 1A). A nodule was not found in the heart or brain in the counter twin fetus after ultrasound or magnetic resonance imaging throughout the pregnancy.
Amniocentesis was offered for a genetic test. TSC1 and TSC2 were checked by clinical exome sequencing using the SureSelect XT Human All Exon V6 kit (Agilent Technologies, Santa Clara, CA, USA), and HiSeqTM X Ten (Illumina, San Diego, CA, USA) sequencing. The sequencing quality was assessed, and all reads were aligned to the GRCh37/hg19 human reference sequence. All acquired single-nucleotide variants were annotated further and filtered on the Ingenuity® Variant Analysis platform (Ingenuity Systems, Redwood City, CA, USA). Zygosity was tested by 20 short tandem repeat markers spanning 46 chromosomes.
A nonsense mutation TSC2 (NM_000548.3): c.4762C>T(p.Gln1588*) was identified in both fetuses but not in the mother. Clinical phenotypes, such as hypopigmented macules, angiofibromas, ungual fibromas, shagreen patches, dental pits, or intraoral fibromas were not identified in the mother or her parents. The renal ultrasound and echocardiogram for the mother were negative. The sperm donor did not have a history of major disease, but a DNA sample was not available.
The family decided to terminate the pregnancy at 27 weeks and 5 days, and a pathological examination was declined.
The couple was healthy, 34 years of age, and non-consanguineous. The woman had experienced three abortions in the first trimester previously due to personal reasons. The current pregnancy was conceived naturally. Monochorionic diamniotic twins were identified at 12-week gestational age. Nuchal translucencies of both fetuses were within normal range (0.8 mm/1.5 mm). Noninvasive prenatal testing for fetal aneuploidy was low risk. A rhabdomyoma was suspected on the apex of the left ventricle in one fetus at 17 weeks and 4 days of gestation. A level-II ultrasound at 20 weeks and 6 days revealed a growth discrepancy between the two fetuses with an estimated weight of 30%. The rhabdomyoma in the left ventricle increased from 6.6 mm × 4.4 mm × 3.7 mm to 9.5 mm × 4.2 mm × 3.5 mm within 7 days in the larger fetus. At week-23 and 5 days, multiple rhabdo-myomas were confirmed in the anterior papillary muscle of the mitral valve, as well as the apex and lateral wall of the left ventricle of the larger fetus, with the largest dimension being 20.8 mm × 10.6 mm × 15 mm. These multiple rhab-domyomas also affected the right ventricle at 25 weeks and 6 days of gestation (Fig. 1B). Tubers in the brain were also identified by ultrasound and magnetic resonance imaging of the fetus (Fig. 1C). However, the smaller fetus remained clinically asymptomatic until 25 weeks and 6 days, when multiple rhabdomyomas were also identified.
The mother presented with multiple renal hamartomas with normal renal function. Hypopigmented macules, angiofibromas, ungual fibromas, shagreen patches, dental pits, or intraoral fibromas were not identified in the mother or her parents.
Both fetuses carried a heterozygous segmental deletion encompassing the 30th exon of TSC2 (chr16:2,131,578–2,131,817). The deletion was confirmed by electrophoresis of the amplicon covering exon 30. The band of the deleted amplicon was significantly lighter for the mother compared with that of the twins, which indicated a mosaic status (Fig. 1D).
The couple decided to terminate the pregnancy at 28 weeks and 4 days. They declined a postmortem pathology examination even after approval by the ethics committee.
TSC cases show high clinical variabilities among family members and even in monozygotic twins.8 Some postnatal cases have suggested clinical discordance in monozygotic twins suffering from TSC. Martin et al.9 described one pair of monozygotic twins who had similar levels of motor delay and mental retardation. One twin had a shagreen patch and a cardiac rhabdomyoma. The twin brother did not have skin or cardiac manifestations but had renal angiomyolipomas and cystic alteration at 3 years of age.9 A similar case was reported by Kondo and colleagues.10 One twin had an angiofibroma, a shagreen patch, angiomyolipoma, calcifications in the brain, and hypopigmented macules, whereas the other twin had only hypopigmented macules.10 Humphrey11 indicated that cognitive functioning and autistic-evaluation results were significantly different between one pair of clinically and genetically confirmed TSC twins. Here, we reported two families with prenatal TSC. In family 1, one fetus had multiple rhabdomyomas and brain tubers as early as 23 weeks, whereas the other twin fetus remained negative throughout the 27 weeks of pregnancy. In family 2, the smaller fetus had multiple cardiac rhabdomyomas 8-weeks later than the larger fetus. These monozygotic twins suffering from TSC prenatally highlighted discordance in the onset and progression of fetal cardiac rhabdomyomas and tubers in the central nervous system, which suggested that the clinical variability of TSC patients might arise from theintrauterinestage.
The variant c.4762C>Tof TSC2 described in our study resulted in a premature stop codon at the 1588th amino acid, and was located in the RapGap-like domain (1517th to the 1674th amino acid) of tuberin, which catalyzes the hydrolysis of Rheb Guanosine Tri-Phosphate to Gheb Guanosine Diphosphate.12 This variant can be classified as “pathogenic” according to the American College of Medical Genetics and Genomics guideline, which results in deleterious protein function. Consistent with the two-hit hypothesis, c.4762C>Twas identified previously in an angiomyolipoma sample from a non-TSC patient with a 4-Mb copy neural loss of heterozygosity of chromo-some 16 compassing TSC2 on the other allele.13 In the present study, the germline mutation c.4762C>Twas present in both fetuses of family 1. The 16p loss of heterozygosity was excluded by high-density single nucleotide polymorphism arrays and whole-genome sequencing. Mosaic status was not detected upon testing of peripheral blood from the mother. Studies have also shown TSC2 mutations to be more frequent that TSC1 mutations, and cardiac rhabdomyomas to be more frequent in cases with TSC2 mutations,14,15 data which are in accordance with the results from the current study. Family 1 also highlights the controversy of whether the detailed medical assessment of the sperm donor should include screening for genetic diseases.16 If screening for genetic diseases is included in the sperm-donor process, the number and type of diseases should be discussed thoroughly and documented with the ethics committee. The possible explanation for the difference in clinical manifestations in these TSC twins is gene expression.17,18 Gene expression can be affected by environmental factors, epigenetic influences, and different tissue localization. The twins may not be “truly identical” due to differences in the intrauterine environment, such as unequal allocation, placental vascular supply, or possible mitotic crossing-over. Recently, several studies have shown that DNA methylation can influence fetal development.19,20 Animal studies have provided evidence of epigenetic modulation of interleukin-1β signaling in TSC.21 Thus, we speculated the phenotypes for monozygotic twins carrying identical mutations may be modified by discordance of DNA methylation.
The variation in clinical manifestations of TSC can also be affected by mosaicism. The mother of family 2 had a subclinical manifestation only after her fetuses had been diagnosed. She had a mosaic deletion of exon 30 in TSC2. This feature also demonstrated intrafamily variability for TSC due to tissue mosaicism. Other at-risk asymptotic family members would be encouraged to be tested for the TSC2 deletion and genetic counseling.
In conclusion, we reported two cases of monozygotic TSC twins caused by TSC2 defects manifested as discordant intrauterine phenotypes. One mother had subtle symptoms and carried a mosaic TSC2 exon deletion. Our study provides intrauterine examples of clinical variability among TSC patients. A prenatal diagnosis of cardiac rhabdomyomas should consider genetic analyses of TSC2 and TSC2 in the index fetuses and parents to assist the final clinical diagnosis and treatment.
The authors thank the family members who volunteered to be involved in this study.
This work was funded by the National Key R&D Program of China (2018YFC1002900), National Natural Science Foundation of China (82071656), Shanghai Shenkang Hospital Development Center (SHDC2020CR6028–005), and the research program of Shanghai First Maternity and Infant hospital (2019B05).
Conflicts of Interest
Luming Sun is an Associate Editor of Maternal-Fetal Medicine. The article was subject to the journal’s standard procedures, with peer review handled independently of this editor and their research groups.
1. Curatolo P, Moavero R, Roberto D, et al. Genotype/phenotype correlations in tuberous sclerosis complex
. Semin Pediatr Neurol 2015;22(4):259–273. doi:10.1016/j.spen.2015.10.002.
2. de Vries PJ, Whittemore VH, Leclezio L, et al. Tuberous sclerosis associated neuropsychiatric disorders (TAND) and the TAND checklist. Pediatr Neurol 2015;52(1):25–35. doi:10.1016/j.pedia-trneurol.2014.10.004.
3. Northrup H, Krueger DA. International Tuberous Sclerosis Complex
Consensus Group. Tuberous sclerosis complex
diagnostic criteria update: recommendations of the 2012 International Tuberous Sclerosis Complex
Consensus Conference. Pediatr Neurol 2013;49(4):243–254. doi:10.1016/j.pediatrneurol.2013.08.001.
4. Samueli S, Abraham K, Dressler A, et al. Tuberous sclerosis complex
: new criteria for diagnostic work-up and management. Wien Klin Wochenschr 2015;127(15-16):619–630. doi:10.1007/s00508-015-0758-y.
5. Lyczkowski DA, Conant KD, Pulsifer MB, et al. Intrafamilial phenotypic variability in tuberous sclerosis complex
. J Child Neurol 2007;22(12):1348–1355. doi:10.1177/0883073807307093.
6. Shukla AK, Reddy AK, Latha A, et al. Cardiac rhabdomyoma: an antenatal illustration. BMJ Case Rep 2015;2015:bcr2014209256. doi:10.1136/bcr-2014-209256.
7. Zhen L, Yang YD, He Y, et al. Prenatal genetic diagnosis of cardiac rhabdomyoma: a single-center experience. Eur J Obstet Gynecol Reprod Biol 2020;249:7–10. doi:10.1016/j.ejogrb.2020.03.051.
8. Wang F, Xiong S, Wu L, et al. A novel TSC2 missense variant associated with a variable phenotype of tuberous sclerosis complex
: case report of a Chinese family. BMC Med Genet 2018;19(1):90. doi:10.1186/s12881-018-0611-z.
9. Martin N, Zügge K, Brandt R, et al. Discordant clinical manifestations in monozygotic twins with the identical mutation in the TSC2 gene. Clin Genet 2003;63(5):427–430. doi:10.1034/j.1399-0004.2003.00073.x.
10. Kondo S, Yamashina U, Sato N, et al. Discordant expression of tuberous sclerosis in monozygotic twins. J Dermatol 1991;18(3):178–180. doi:10.1111/j.1346-8138.1991.tb03063.x.
11. Humphrey A, Higgins JN, Yates JR, et al. Monozygotic twins with tuberous sclerosis discordant for the severity of developmental deficits. Neurology 2004;62(5):795–798. doi:10.1212/01.wnl.0000113745.58425.ef.
12. Scrima A, Thomas C, Deaconescu D, et al. The Rap-RapGAP complex: GTP hydrolysis without catalytic glutamine and arginine residues. EMBO J 2008;27(7):1145–1153. doi:10.1038/emboj.2008.30.
13. Giannikou K, Malinowska IA, Pugh TJ, et al. Whole exome sequencing identifies TSC1/TSC2 biallelic loss as the primary and sufficient driver event for renal angiomyolipoma development. PLoS Genet 2016;12(8):e1006242. doi:10.1371/journal.pgen.1006242.
14. Józwiak S, Kotulska K. Are all prenatally diagnosed multiple cardiac rhabdomyomas a sign of tuberous sclerosis?Prenat Diagn 2006;26(9):867–869. doi:10.1002/pd.1506.
15. Hung CC, Su YN, Chien SC, et al. Molecular and clinical analyses of 84 patients with tuberous sclerosis complex
. BMC Med Genet 2006;7:72. doi:10.1186/1471-2350-7-72.
16. Pennings G. Expanded carrier screening should not be mandatory for gamete donors. Hum Reprod 2020;35(6):1256–1261. doi:10.1093/humrep/deaa088.
17. Jentarra GM, Rice SG, Olfers S, et al. Evidence for population variation in TSC1 and TSC2 gene expression. BMC Med Genet 2011;12:29. doi:10.1186/1471-2350-12-29.
18. Jonsson H, Magnusdottir E, Eggertsson HP, et al. Differences between germline genomes of monozygotic twins. Nat Genet 2021;53(1):27–34. doi:10.1038/s41588-020-00755-1.
19. DuPriest E, Hebert J, Morita M, et al. Fetal renal DNA methylation and developmental programming of stress-induced hypertension in growth-restricted male mice. Reprod Sci 2020;27(5):1110–1120. doi:10.1007/s43032-019-00121-5.
20. Fujioka K, Nishida K, Ashina M, et al. DNA methylation of the Rtl1 promoter in the placentas with fetal growth restriction. Pediatr Neonatol 2019;60(5):512–516. doi:10.1016/j.pedneo.2019.01.001.
21. Fuso A, Iyer AM, van Scheppingen J, et al. Promoter-specific hypomethylation correlates with IL-1β overexpression in tuberous sclerosis complex
(TSC). J Mol Neurosci 2016;59(4):464–470. doi:10.1007/s12031-016-0750-7.
Edited By Yang Pan
How to cite this article: Xiong S, Wu F, Chen G, Wang J, Yang Y, Xing Y, Sun L. Prenatal Phenotypical Discrepancy in Monozygotic Twins with Tuberous Sclerosis Complex. Maternal Fetal Med 2022;4(4):286–289. doi: 10.1097/FM9.0000000000000109.