The role of molecular genetic methods in the diagnosis of McCune—Albright syndrome

McCune-Albright syndrome (MAS) is a rare genetic disorder which is caused by somatic mutations in the GNAS gene. Clinical symptoms of MAS include café-au-lait skin pigmentation, fibrous dysplasia, and autonomous endocrine hyperfunction. Somatic character of the gene defects determines wide variety of syndrome manifestations, from mild forms with minimum presentation to severe conditions with aggressive course. Potential multicomponent form of the MAS syndrome necessitates the dynamic monitoring, including regular screening for possible components of the disease. Therefore, additional methods specifying the diagnosis of MAS syndrome, especially of its suppressed forms, should facilitate selection of patient management strategy and monitoring rate and/or complete exclusion of the diagnosis. Molecular genetic verification of the diagnosis may be one of these methods.

Objective — the study was aimed at evaluating massive parallel sequencing (next generation sequencing, NGS) and real-time polymerase chain reaction using the TaqMan technique for detection of somatic mutations (competitive allele-specific TaqMan PCR, CAST-PCR) in the diagnosis of somatic mutations R201C and R201H in the GNAS gene based on DNA obtained from the peripheral blood.

Material and methods. The study included patients diagnosed with and suspected for MAS syndrome. Molecular genetic testing of R201C and R201H mutations in the GNAS gene based on DNA extracted from peripheral blood leukocytes was carried out by Next generation sequencing (NGS) and real-time polymerase chain reaction methods using the TaqMan technique for detection of somatic mutations (competitive allele-specific TaqMan PCR, CAST-PCR). Based on clinical data, patients were divided into groups depending on the severity of the disease and the number of MAS manifestations. The results were evaluated by comparing the rate of detected molecular genetic defects in the formed groups of patients.

Results. Molecular genetic study included 39 children with MAS syndrome and 6 children with suspected MAS. R201C and R201H mutations in GNAS gene were detected in 16 patients with severe to moderate MAS 16 (41%) 39. No mutations were detected in other MAS patients and patients with suspected MAS.

Conclusion. NGS and CAST-PCR methods can detect the presence of mutant alleles R201C and R201H of GNAS gene in DNA samples obtained from the blood in the case of severe to moderate MAS syndrome, but they cannot be recommended for MAS diagnosis based on the peripheral blood samples in children with mild signs of the syndrome or suspected diagnosis.

McCune—Albright syndrome, somatic mutations, massive parallel sequencing, real-time polymerase chain reaction

1. Albright F, Butler AM, Hampton AO, Smith P. Syndrome characterized by osteitis fibrosa disseminata, areas of pigmentation and endocrine dysfunction, with precocious puberty in females. N Engl J Med. 1937;216(17):727-746. doi: 10.1056/nejm193704292161701

2. McСune DJ, Bruch H. Osteodystrophia fibrosa: report of a case in which the condition was combined with precocious puberty, pathologic pigmentation of the skin and hyperthyroidism, with a review of the literature. Am J Dis Child. 1937;54(4):806-848. doi: 10.1001/archpedi.1937.01980040110009

3. Брайцев Б.Р. Osteodystrophia fibrosa localisata. Вестник хирургии и пограничных областей. — 1928. — №1. — С. 301-315. [Braytsev BR. Osteodystrophia fibrosa localisata. Vestnik khirurgii i pogranicnyh oblastej. 1928;(1):301-315. (In Russ.)].

4. Happle R. The McCune—Albright syndrome: a lethal gene surviving by mosaicism. Clin Genet. 2008;29(4):321-324. doi: 10.1111/j.1399-0004.1986.tb01261.x

5. Angelousi A, Fencl F, Faucz FR, et al. McCune Albright syndrome and bilateral adrenal hyperplasia: the GNAS mutation may only be present in adrenal tissue. Hormones (Athens). 2015;14(3):447-450. doi: 10.14310/horm.2002.1578

6. Meyerson M, Gabriel S, Getz G. Advances in understanding cancer genomes through second-generation sequencing. Nat Rev Genet. 2010;11(10):685-696. doi: 10.1038/nrg2841

7. Bohers E, Viailly PJ, Dubois S, et al. Somatic mutations of cell-free circulating DNA detected by next-generation sequencing reflect the genetic changes in both germinal center B-cell-like and activated B-cell-like diffuse large B-cell lymphomas at the time of diagnosis. Haematologica. 2015;100(7):e280-e284. doi: 10.3324/haematol.2015.123612

8. Roma C, Esposito C, Rachiglio AM, et al. Detection of EGFR mutations by TaqMan mutation detection assays powered by competitive allele-specific TaqMan PCR technology. Biomed Res Int. 2013;2013:385087. doi: 10.1155/2013/385087

9. Barbano R, Pasculli B, Coco M, et al. Competitive allele-specific TaqMan PCR (Cast-PCR) is a sensitive, specific and fast method for BRAF V600 mutation detection in Melanoma patients. Sci Rep. 2015;5:18592. doi: 10.1038/srep18592

10. Bourne HR, Sanders DA, McCormick F. The GTPase superfamily: a conserved switch for diverse cell functions. Nature. 1990;348(6297):125-132. doi: 10.1038/348125a0

11. Landis CA, Masters SB, Spada A, et al. GTPase inhibiting mutations activate the alpha chain of Gs and stimulate adenylyl cyclase in human pituitary tumours. Nature. 1989;340(6236):692-696. doi: 10.1038/340692a0

12. Clementi E, Malgaretti N, Meldolesi J, Taramelli R. A new constitutively activating mutation of the Gs protein alpha subunit-gsp oncogene is found in human pituitary tumours. Oncogene. 1990;5(7):1059-1061.

13. Idowu BD, Al-Adnani M, O’Donnell P, et al. A sensitive mutation-specific screening technique for GNAS1 mutations in cases of fibrous dysplasia: the first report of a codon 227 mutation in bone. Histopathology. 2007;50(6):691-704. doi: 10.1111/j.1365-2559.2007.02676.x

14. O’Hayre M, Vazquez-Prado J, Kufareva I, et al. The emerging mutational landscape of G proteins and G-protein-coupled receptors in cancer. Nat Rev Cancer. 2013;13(6):412-424. doi: 10.1038/nrc3521

15. Weinstein LS, Shenker A, Gejman PV, et al. Activating mutations of the stimulatory G protein in the McCune—Albright syndrome. N Engl J Med. 1991;325(24):1688-1695. doi: 10.1056/NEJM199112123252403

16. Candeliere GA, Roughley PJ, Glorieux FH. Polymerase chain reaction-based technique for the selective enrichment and analysis of mosaic arg201 mutations in Gαs from patients with fibrous dysplasia of bone. Bone. 1997;21(2):201-206. doi: 10.1016/s8756-3282(97)00107-5

17. Narumi S, Matsuo K, Ishii T, et al. Quantitative and sensitive detection of GNAS mutations causing mccune-albright syndrome with next generation sequencing. PLoS One. 2013;8(3):e60525. doi: 10.1371/journal.pone.0060525

18. de Sanctis L, Galliano I, Montanari P, et al. Combining real-time COLD- and MAMA-PCR TaqMan techniques to detect and quantify R201 GNAS mutations in the McCune—Albright syndrome. Horm Res Paediatr. 2017;87(5):342-349. doi: 10.1159/000463384