Familial Cancer

, Volume 17, Issue 2, pp 261–268 | Cite as

Mutational analysis of the RB1 gene and the inheritance patterns of retinoblastoma in Jordan

  • Yacoub A. Yousef
  • Abdelghani Tbakhi
  • Maysa Al-Hussaini
  • Ibrahim AlNawaiseh
  • Ala Saab
  • Amal Afifi
  • Maysa Naji
  • Mona Mohammad
  • Rasha Deebajah
  • Imad Jaradat
  • Iyad Sultan
  • Mustafa Mehyar
Original Article


Retinoblastoma (RB) is a childhood cancer developing in the retina due to RB1 pathologic variant. Herein we are evaluating the oncogenic mutations in the RB1 gene and the inheritance patterns of RB in the Jordanian patients. In this prospective study, the peripheral blood of 50 retinoblastoma patients was collected, genomic DNA was extracted, mutations were identified using Quantitative multiplex PCR (QM-PCR), Allele-specific PCR, Next Generation Sequencing analysis, and Sanger sequencing. In this cohort of 50 patients, 20(40%) patients had unilateral RB and 30(60%) were males. Overall, 36(72%) patients had germline disease, 17(47%) of whom had the same RB1 pathologic variant detected in one of the parents (inherited disease). In the bilateral group, all (100%) patients had germline disease; 13(43%) of them had inherited mutation. In the unilateral group, 6(30%) had germline disease, 4(20%) of them had inherited mutation. Nonsense mutation generating a stop codon and producing a truncated non-functional protein was the most frequent detected type of mutations (n = 15/36, 42%). Only one (2%) of the patients had mosaic mutation, and of the 17 inherited cases, 16(94%) had an unaffected carrier parent. In conclusion, in addition to all bilateral RB patients in our cohort, 30% of unilateral cases showed germline mutation. Almost half (47%) of germline cases had inherited disease from affected (6%) parent or unaffected carrier (94%). Therefore molecular screening is critical for the genetic counseling regarding the risk for inherited RB in both unilateral and bilateral cases including those with no family history.


Germline Mutation RB1 gene Retinoblastoma 



We want to thank The Impact Genetics Lab Inc. (Ontario, Canada), mainly Prof. B, Gallie (Laboratory Director and RB Medical Director), F. Jewett (Managing Director), and H. Racher, PHD (Scientific and Laboratory Director) for providing us with mutational analysis for RB1 gene in our patients. Also we thank Mr. Raed Marashdeh, and Mr. Ahmad Qawas from King Hussein Cancer Center for solving all issues related to logistics.


This research was supported by an intra-mural grant from the Research Council, King Hussein Cancer Center (Grant No. 13KHCC25) (Amman, Jordan).

Compliance with ethical standards

Conflict of interest

All authors declare that they have no conflict of interest.


  1. 1.
    Yun J, Li Y, Xu C-T, Pan B-R (2011) Epidemiology and Rb1 gene of retinoblastoma. Int J Ophthalmol 4(1):103–109PubMedPubMedCentralGoogle Scholar
  2. 2.
    Broaddus E, Topham A, Singh AD (2009) Incidence of retinoblastoma in the USA: 1975–2004. Br J Ophthalmol 93(1):21–23CrossRefPubMedGoogle Scholar
  3. 3.
    Dimaras H, Corson TW, Cobrinik D et al (2015) Retinoblastoma. Nat Rev Dis Primers 27(1):15021CrossRefGoogle Scholar
  4. 4.
    Taylor M, Dehainault C, Desjardins L et al (2006) Genotype–phenotype correlation in hereditary familial retinoblastoma. Hum Mutat 28:284–293CrossRefGoogle Scholar
  5. 5.
    Jaradat I, Yousef YA, Mehyar M et al (2011) Retinoblastoma in Jordan: an epidemiological study (2006–2010). Hematol Oncol Stem Cell Ther 4(3):126–131CrossRefPubMedGoogle Scholar
  6. 6.
    Corson TW, Gallie BL (2007) One hit, two hits, three hits, more? Genomic changes in the development of retinoblastoma. Genes Chromosomes Cancer 46:617–634CrossRefPubMedGoogle Scholar
  7. 7.
    Knudson AG (1971) Mutation and cancer: statistical study of retinoblastoma. Proc Natl Acad Sci USA 68:820–823CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Harbour JW (2001) Molecular basis of low-penetrance retinoblastoma. Arch Ophthalmol 119(11):1699–1704 (Review)CrossRefPubMedGoogle Scholar
  9. 9.
    Valverde JR, Alonso J, Palacios I et al (2005) RB1 gene mutation update. A meta-analysis based on 932 reported mutations available in searchable database. BMC Genet 6:53CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    de Andrade AF, da Hora Barbosa R, Vargas FR et al (2006) A molecular study of first and second RB1 mutational hits in retinoblastoma patients. Cancer Genet Cytogenet 167(1):43–46CrossRefPubMedGoogle Scholar
  11. 11.
    Houdayer C, Gauthier-Villars M, Lauge A et al (2004) Comprehensive screening for constitutional RB1 mutations by DHPLC and QMPSF. Hum Mutat 23:193–202CrossRefPubMedGoogle Scholar
  12. 12.
    Devarajan B, Prakash L, Kannan TR et al (2015) Targeted next generation sequencing of RB1 gene for the molecular diagnosis of Retinoblastoma. BMC Cancer 28(15):320CrossRefGoogle Scholar
  13. 13.
    Soliman SE, Dimaras H, Khetan V, Gardiner JA, Chan HS, Héon E, Gallie BL (2016) Prenatal versus postnatal screening for familial retinoblastoma. Ophthalmology 123(12):2610–2617CrossRefPubMedGoogle Scholar
  14. 14.
    Gallie B, Erraguntla V, Heon E, Chan H (2004) Retinoblastoma. In: Taylor D, Hoyt C (eds) Pediatric ophthalmology and strabismus, vol Chap. 50. Elsevier, London, p. 486–504Google Scholar
  15. 15.
    Rebbeck TR, Rogatko A, Viana MA (1997) Evaluation of genotype data in clinical risk assessment: methods and application to BRCA1, BRCA2, and N-acetyl transferase-2 genotypes in breast cancer. Genet Test 1:157–164CrossRefPubMedGoogle Scholar
  16. 16.
    Ganguly A (2002) An update on conformation sensitive gel electrophoresis. Hum Mutat 19(4):334–342CrossRefPubMedGoogle Scholar
  17. 17.
    Price EA, Price K, Kolkiewicz K, Hack S, Reddy MA, Hungerford JL, Kingston JE, Onadim Z (2014) Spectrum of RB1 mutations identified in 403 retinoblastoma patients. J Med Genet 51:208–214CrossRefPubMedGoogle Scholar
  18. 18.
    Rushlow D, Piovesan B, Zhang K et al (2009) Detection of mosaic RB1 mutations in families with retinoblastoma. Hum Mutat 30:842–851CrossRefPubMedGoogle Scholar
  19. 19.
    Nichols KE, Walther S, Chao E, Shields C, Ganguly A (2004) Recent advances in retinoblastoma genetic research. Comprehensive screening for constitutional RB1 mutations by DHPLC and QMPSF. Hum Mutat 23:193–202CrossRefGoogle Scholar
  20. 20.
    Nichols KE, Waltherb S, Chaob E, Shieldsc C, Gangulyb A (2009) Current opinion in ophthalmology. Recent advances in retinoblastoma genetic research. Curr Opin Ophthalmol 20:351–355CrossRefPubMedGoogle Scholar
  21. 21.
    Richter S, Vandezande K, Chen N et al (2003) Sensitive and efficient detection of RB1 gene mutations enhances care for families with retinoblastoma. Am J Hum Genet 72(2):253–269CrossRefPubMedGoogle Scholar
  22. 22.
    Sugano K, Yoshida T, Izumi H et al (2004) Outpatient clinic for genetic counseling and gene testing of retinoblastoma. Int J Clin Oncol 9:25–30CrossRefPubMedGoogle Scholar
  23. 23.
    Lohmann DR, Brandt B, Hopping W, Passarge E, Horsthemke B (1996) The spectrum of RB1 germ-line mutations in hereditary retinoblastoma. Am J Hum Genet 58:940–949PubMedPubMedCentralGoogle Scholar
  24. 24.
    Alonso J, Frayle H, Menéndez I et al (2005) Identification of 26 new constitutional RB1 gene mutations in Spanish, Colombian, and Cuban retinoblastoma patients. Hum Mutat 25:99CrossRefPubMedGoogle Scholar
  25. 25.
    Abidi O, Knari S, Sefri H et al (2011) Mutational analysis of the RB1 gene in Moroccan patients with retinoblastoma. Mol Vis 17:3541–3547PubMedPubMedCentralGoogle Scholar
  26. 26.
    Sampieri K, Hadjistilianou T, Mari F et al (2006) Mutational screening of the RB1 gene in Italian patients with retinoblastoma reveals 11 novel mutations. J Hum Genet 51:209–216CrossRefPubMedGoogle Scholar
  27. 27.
    Macías M, Dean M, Atkinson A et al (2008) Spectrum of RB1 gene mutations and loss of heterozygosity in Mexican patients with retinoblastoma: identification of six novel mutations. Cancer Biomark 4:93–99CrossRefPubMedGoogle Scholar
  28. 28.
    Dalamón V, Surace E, Giliberto F, Ferreiro V, Fernandez C, Szijan I (2004) Detection of germline mutations in argentine retinoblastoma patients: low and full penetrance retinoblastoma caused by the same germline truncating mutation. J Biochem Mol Biol 37:246–253PubMedGoogle Scholar
  29. 29.
    Nichols KE, Houseknecht MD, Godmilow L et al (2005) Sensitive multistep clinical molecular screening of 180 unrelated individuals with retinoblastoma detects 36 novel mutations in the RB1 gene. Hum Mutat 25:566–574CrossRefPubMedGoogle Scholar
  30. 30.
    Alonso J, García-Miguel P, Abelairas J et al (2001) Spectrum of germline RB1 gene mutations in Spanish retinoblastoma patients: phenotypic and molecular epidemiological implications. Hum Mutat 17:412–422CrossRefPubMedGoogle Scholar
  31. 31.
    Mallipatna A, Marino M, Singh AD (2016) Genetics of Retinoblastoma. Asia Pac J Ophthalmol 5(4):260–264CrossRefGoogle Scholar
  32. 32.
    Houdayer C, Gauthier-Villars M, Laugé A et al (2004) Comprehensive screening for constitutional RB1 mutations by DHPLC and QMPSF. Hum Mutat 23:193–202CrossRefPubMedGoogle Scholar
  33. 33.
    Parsam VL, Kannabiran C, Honavar S, Vemuganti GK, Ali MJ (2009) A comprehensive, sensitive and economical approach for the detection of mutations in the RB1 gene in retinoblastoma. J Genet 88(4):517–527CrossRefPubMedGoogle Scholar
  34. 34.
    Pradhan MA, Ng Y, Strickland A, George PM, Raizis A, Warrington J, Vincent AL (2010) Role of genetic testing in retinoblastoma management at a tertiary referral centre. Clin Exp Ophthalmol 38:231–236PubMedGoogle Scholar
  35. 35.
    Lohmann DR, Gallie BL (2004). Retinoblastoma: revisiting the model prototype of inherited cancer. Am J Med Genet 129C(1):23–28CrossRefPubMedGoogle Scholar
  36. 36.
    Houdayer C, Gauthier-Villars M, Lauge A et al (2004) Comprehensive screening for constitutional RB1 mutations by DHPLC and QMPSF. Hum Mutat 23(2):193–202CrossRefPubMedGoogle Scholar
  37. 37.
    Joseph B, Shanmugam MP, Srinivasan MK, Kumaramanickavel G (2004) Retinoblastoma: genetic testing versus conventional clinical screening in India. Mol Diagn 8(4):237–243PubMedGoogle Scholar
  38. 38.
    Soliman SE, Racher H, Zhang C, MacDonald H, Gallie BL (2017) Genetics and molecular diagnostics in retinoblastoma—an update. Asia Pac J Ophthalmol 6(2):197–207CrossRefGoogle Scholar
  39. 39.
    Fletcher O, Easton D, Anderson K, Gilham C, Jay M, Peto J (2004) Lifetime risks of common cancers among retinoblastoma survivors. J Natl Cancer Inst 96(5):357–363CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2017

Authors and Affiliations

  • Yacoub A. Yousef
    • 1
  • Abdelghani Tbakhi
    • 2
  • Maysa Al-Hussaini
    • 3
  • Ibrahim AlNawaiseh
    • 1
  • Ala Saab
    • 1
  • Amal Afifi
    • 2
  • Maysa Naji
    • 4
  • Mona Mohammad
    • 1
  • Rasha Deebajah
    • 4
  • Imad Jaradat
    • 5
  • Iyad Sultan
    • 4
  • Mustafa Mehyar
    • 1
  1. 1.Department of Surgery/OphthalmologyKing Hussein Cancer CenterAmmanJordan
  2. 2.Department of Cell Therapy and Applied GenomicsKing Hussein Cancer CenterAmmanJordan
  3. 3.Department of PathologyKing Hussein Cancer CenterAmmanJordan
  4. 4.Department of Pediatric OncologyKing Hussein Cancer CenterAmmanJordan
  5. 5.Department of RadiotherapyKing Hussein Cancer CenterAmmanJordan

Personalised recommendations