Absence of CHEK2 1100delC, R145W and I157T Mutations in Breast Cancer in a Moroccan Population

Amal ElAmrani, Khalid Moumad, Mohammed Attaleb, Mustapha Benhassou, Asta Försti, Moulay Mustapha Ennaji, Mohammed El Mzibri, Meriem Khyatti

  Open Access OPEN ACCESS  Peer Reviewed PEER-REVIEWED

Absence of CHEK2 1100delC, R145W and I157T Mutations in Breast Cancer in a Moroccan Population

Amal ElAmrani1, 2, Khalid Moumad1, Mohammed Attaleb3, Mustapha Benhassou4, Asta Försti5, 6, Moulay Mustapha Ennaji2, Mohammed El Mzibri3, Meriem Khyatti1,

1Oncovirology Laboratory, Institut Pasteur du Maroc, Casablanca, Morocco

2Laboratory of Virology, Microbiology and Quality/EBQ, Faculty of Sciences & Technics University Hassan II Mohammedia-Casablanca, Mohammedia, Morocco

3Unité de Biologie et Recherche Médicale, Centre National de l'Energie, des Sciences et des Techniques Nucléaires, (CNESTEN), Rabat, Morocco

4Obstetrics Service "A" Maternité Lalla Meryem, CHU Ibn Rochd Casablanca, Morocco

5Division of Molecular Genetic Epidemiology, German Cancer Research Center (DKFZ), Heidelberg, Germany

6Center for Primary Health Care Research, Clinical Research Center, Lund University, Malmö, Sweden


Mutations in the BRCA1 and BRCA2 genes confer a high risk of breast cancer (BC), although they account for only a small fraction of BC susceptibility. Rare mutations in genes conferring moderate risk may contribute to BC risk. Previous studies have shown that mutations in the CHEK2 gene, which encodes for an upstream regulator of BRCA1, may cause a moderately increased BC risk. In the current study we investigated the status of three founder mutations in the CHEK2 gene (c.1100delC, R145W and I157T) using direct sequencing in 50 BC and 50 control samples. No mutations were detected. This result is in line with the postulated existence of a c.1100delC frequency gradient from the North to the South in Europe with higher frequencies in the Northern countries.

Cite this article:

  • ElAmrani, Amal, et al. "Absence of CHEK2 1100delC, R145W and I157T Mutations in Breast Cancer in a Moroccan Population." Journal of Cancer Research and Treatment 2.1 (2014): 6-9.
  • ElAmrani, A. , Moumad, K. , Attaleb, M. , Benhassou, M. , Försti, A. , Ennaji, M. M. , Mzibri, M. E. , & Khyatti, M. (2014). Absence of CHEK2 1100delC, R145W and I157T Mutations in Breast Cancer in a Moroccan Population. Journal of Cancer Research and Treatment, 2(1), 6-9.
  • ElAmrani, Amal, Khalid Moumad, Mohammed Attaleb, Mustapha Benhassou, Asta Försti, Moulay Mustapha Ennaji, Mohammed El Mzibri, and Meriem Khyatti. "Absence of CHEK2 1100delC, R145W and I157T Mutations in Breast Cancer in a Moroccan Population." Journal of Cancer Research and Treatment 2, no. 1 (2014): 6-9.

Import into BibTeX Import into EndNote Import into RefMan Import into RefWorks

1. Introduction

Breast cancer (BC) is the most frequently diagnosed cancer and the leading cause of cancer death in females worldwide, accounting for 23% of total new cancer cases and 14% of total cancer deaths [1]. BC is a heterogeneous disease in which breast cells become abnormal and multiply to form a malignant tumor. In about 5 to 10% of cases, BC is due to hereditary genetic background caused by mutations in high penetrance susceptibility genes, which enhance familial risk [2]. About 16% of hereditary BCs can be attributed to germline mutations in the breast cancer 1 (BRCA1) and 2 (BRCA2) genes [3]. Mutations in other genes confer moderate risk of BC development. Among them, the Checkpoint kinase 2 (CHEK2) is a low penetrance gene mutated in less than 3% of BC cases [4].

CHEK2 is a functionally related DNA repair gene; it emerges as an important signal transducer that mediates cellular responses to DNA damage. In response to double-strand DNA breaks, CHEK2 activates p53 by phosphorylation [5]. CHEK2 is also a candidate tumor suppressor whose defects contribute to molecular pathogenesis of diverse types of human malignancies. Worldwide, different CHEK2 mutations have been reported to be associated with BC development. The c.1100delC is the first recurrent mutation in the CHEK2 gene to be reported as an important cause of BC [4]. Since then, numerous studies have documented the prevalence of this single founder mutation in various populations, but up to now few studies have been interested in its clinical impact. Thus far, five deleterious recurrent mutations in CHEK2 have been identified that confer about two-fold elevated risk of BC. These include in addition to the truncating mutation c.1100delC, the missense mutations R145W and I157T, the splice site mutation IVS2 + 1G > A and the large genomic 5,395 bp deletion (del5395) [6].

This preliminary study was planned to assess the frequencies of three CHEK2 mutations (c.1100delC, R145W and I157T) in a case-control study of 50 BC patients and 50 breast controls to evaluate the implication of these mutations in predisposition to BC in Morocco.

2. Materials and Methods

A total of 50 BC patients were recruited at the Obstetrics Service of the Ibn Rochd hospital (Casablanca, Morocco) between 2010 and 2012. Tumor tissue samples were obtained from biopsies collected for clinical diagnosis. The characteristics of the studied population and histopathological data are shown in Table 1. Fifty blood samples from healthy women, aged matched with BC patients, were collected to be used as a control group. The study protocol was approved by the Ethical Committee of Pasteur Institute of Morocco, and written informed consent was obtained from each study subject.

All the BC patients and controls were screened for the c.1100delC, R145W and I157T mutations. Mutations detection was done by PCR amplification and direct DNA sequencing as previously described [7]. PCR primers and the amplimers’ sizes are reported in Table 2.

Amplification reaction was performed in a total volume of 25 µl. The amplification mixture contained 50 pmol of each consensus primer, 200 µM of each dNTP, 0.5 units Taq DNA polymerase (Invitrogen / Life Technologies) and 2 µl of DNA sample in 1x Taq polymerase buffer. A touchdown PCR program was used for PCR amplification. PCR conditions were as follows: 94°C for 1 min; seven cycles of 94°C for 30 s, 60°C for 30 s, 68°C for 30 s; twenty eight cycles of 94°C for 30 s, 54°C for 30 s, 68°C for 30 s, followed by 70°C for 5 min and 12°C thereafter. Mutation positive and negative controls as well as a negative control for the PCR reaction were included in the analysis.

For DNA sequencing, the PCR products were purified by the Exo SaP-IT clean up system (USB, USA) and sequenced directly on an ABI 3130XL DNA analyzer (Applied Biosystems, Foster city, CA, USA), using Big Dye® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems, Foster city, CA, USA), according to manufacturer’s protocol. For each sample, PCR amplification and DNA sequencing were performed twice. The sequences obtained were then compared with the non-mutated reference sequence of CHEK2 gene available in the database Genatlas.

Table 1. Characteristics of the Breast cancer cases

3. Results and Discussion

The CHEK2 c.1100delC heterozygosity may be associated with an increased risk of BC; however, it is unclear whether the evidence is sufficient to recommend genotyping in the clinical practice. Thus, the relevance of CHEK2 mutations as a screening target for an elevated risk of BC is of interest. Indeed, it is widely accepted that the risk of BC may be higher for women who have both a CHEK2 gene mutation and a family history of BC [8].

To our knowledge, no study has been conducted on CHEK2 mutations in Moroccan patients with BC. Therefore, we assessed the prevalence of these mutations in a case–control study of BC among Moroccan women. In the present study, the interest was focused on three CHEK2 variants that are known to affect protein function (c.1100delC, R145W, and I175T). The c.1100delC variant is a protein truncating mutation that abrogates CHEK2 kinase activity. R145W has been reported to have disrupted kinase activity and I175T is deficient in binding to BRCA1 and p53 [9].

In our study, neither cancer cases nor controls showed any of the mutations analyzed. Our data on the absence of the c.1100delC mutation in Morocco are in line with those of the previous studies ([10, 11]) and support the notion that c.1100delC does not contribute to BC susceptibility in Mediterranean populations ([12, 13]). In Spain, Osorio et al. did not detect the c.1100delC mutation in 856 samples analyzed [10]. The c.1100delC was also reported to be absent in Asian population. In North American populations, a very low frequency of CHEK2 c.1100delC mutation was reported [14] (Table 3). Interestingly, the highest frequency of CHEK2 1100delC has been found in patients from the North and the West of Europe, as compared to the southern countries exhibiting the lowest frequency (Italy and Spain) ([10, 11]) (Table 3). These findings are in agreement with the hypothesis of the existence of a c.1100delC frequency gradient from the North-West to the South-East of Europe, caused by an ancestral common origin in the North [15]. This gradient may be more accentuated in the Mediterranean countries which may explain the absence of this mutation in the Moroccan population.

Table 3. Population’s frequencies of the studied Mutations in Chek2 gene

The CHEK2 I157T, a missense variant, alters the kinase activity of the CHEK2 protein. This variant has been reported in ethnically diverse populations and has been associated with a modest risk for developing BC among German and Belarusian populations [16] and for developing colon cancer among Polish and Finnish populations [17] (Table 3).

Other rare variants that adversely affect the CHEK2 protein function have been reported in ethnically diverse populations, and seem to marginally contribute to the overall burden of BC [18].

Our findings suggests that CHEK2 mutations is a rare event and play a negligible role in BC risk in Morocco and therefore the clinical relevance of CHEK2 mutations screening as a cancer predisposing gene is very limited.


This paper was supported by EU FP7/2007-2013 grant 260715.


[1]  Jemal A, Bray F, Center MM, Ferlay J, Ward E, Forman D. Global cancer statistics. CA: a cancer journal for clinicians 2011; 61: 69-90.
In article      CrossRefPubMed
[2]  Claus EB, Schildkraut JM, Thompson WD, Risch NJ. The genetic attributable risk of breast and ovarian cancer. Cancer 1996; 77: 2318-24.
In article      CrossRef
[3]  Narod SA, Foulkes WD. BRCA1 and BRCA2: 1994 and beyond. Nature reviews Cancer 2004; 4: 665-76.
In article      CrossRefPubMed
[4]  Meijers-Heijboer H, van den Ouweland A, Klijn J, Wasielewski M, de Snoo A, Oldenburg R, et al. Low-penetrance susceptibility to breast cancer due to CHEK2 (*) 1100delC in noncarriers of BRCA1 or BRCA2 mutations. Nature genetics 2002; 31: 55-9.
In article      CrossRefPubMed
[5]  Chehab NH, Malikzay A, Appel M, Halazonetis TD. Chk2/hCds1 functions as a DNA damage checkpoint in G (1) by stabilizing p53. Genes & development 2000; 14: 278-88.
In article      PubMed
[6]  Rashid MU, Muhammad N, Faisal S, Amin A, Hamann U. Constitutional CHEK2 mutations are infrequent in early-onset and familial breast/ovarian cancer patients from Pakistan. BMC cancer 2013; 13: 312.
In article      CrossRefPubMed
[7]  Kuusisto KM, Bebel A, Vihinen M, Schleutker J, Sallinen SL. Screening for BRCA1, BRCA2, CHEK2, PALB2, BRIP1, RAD50, and CDH1 mutations in high-risk Finnish BRCA1/2-founder mutation-negative breast and/or ovarian cancer individuals. Breast cancer research: BCR 2011; 13: R20.
In article      CrossRefPubMed
[8]  Cybulski C, Wokołorczyk D, Jakubowska A, Huzarski T, Byrski T, Gronwald J, et al. Risk of Breast Cancer in Women With a CHEK2 Mutation With and Without a Family History of Breast Cancer. Journal of Clinical Oncology 2011; 29: 3747-52.
In article      CrossRefPubMed
[9]  Wu X, Webster SR, Chen J. Characterization of tumor-associated Chk2 mutations. The Journal of biological chemistry 2001; 276: 2971-4.
In article      CrossRefPubMed
[10]  Osorio A, Rodriguez-Lopez R, Diez O, de la Hoya M, Ignacio Martinez J, Vega A, et al. The breast cancer low-penetrance allele 1100delC in the CHEK2 gene is not present in Spanish familial breast cancer population. International journal of cancer Journal international du cancer 2004; 108: 54-6.
In article      CrossRefPubMed
[11]  Caligo MA, Agata S, Aceto G, Crucianelli R, Manoukian S, Peissel B, et al. The CHEK2 c.1100delC mutation plays an irrelevant role in breast cancer predisposition in Italy. Human mutation 2004; 24: 100-1.
In article      CrossRefPubMed
[12]  Thirthagiri E, Cheong L, Yip C, Teo S-H. CHEK2* 1100delC does not contribute to risk to breast cancer among Malay, Chinese and Indians in Malaysia. Familial Cancer 2009; 8: 355-8.
In article      CrossRefPubMed
[13]  Weischer M, Bojesen SE, Nordestgaard BG. In Reply. Journal of Clinical Oncology 2008; 26: 2419-20.
In article      CrossRef
[14]  Iniesta MD, Gorin MA, Chien L-C, Thomas SM, Milliron KJ, Douglas JA, et al. Absence of CHEK2* 1100delC mutation in families with hereditary breast cancer in North America. Cancer genetics and cytogenetics 2010; 202: 136-40.
In article      CrossRefPubMed
[15]  González-Hormazábal P, Castro V, Blanco R, Gómez F, Peralta O, Waugh E, et al. Absence of CHEK2 1100delC mutation in familial breast cancer cases from a South American population. Breast Cancer Res Treat 2008; 110: 543-5.
In article      CrossRefPubMed
[16]  Bogdanova N, Enssen-Dubrowinskaja N, Feshchenko S, Lazjuk GI, Rogov YI, Dammann O, et al. Association of two mutations in the CHEK2 gene with breast cancer. International journal of cancer Journal international du cancer 2005; 116: 263-6.
In article      CrossRefPubMed
[17]  Kilpivaara O, Alhopuro P, Vahteristo P, Aaltonen LA, Nevanlinna H. CHEK2 I157T associates with familial and sporadic colorectal cancer. Journal of medical genetics 2006; 43: e34.
In article      CrossRefPubMed
[18]  Nevanlinna H, Bartek J. The CHEK2 gene and inherited breast cancer susceptibility. Oncogene 2006; 25: 5912-9.
In article      CrossRefPubMed
[19]  Consortium CBCC-C: CHEK2* 1100delC and susceptibility to breast cancer: a collaborative analysis involving 10,860 breast cancer cases and 9,065 controls from 10 studies. American journal of human genetics 2004; 74(6): 1175-1182.
In article      CrossRefPubMed
[20]  Friedrichsen DM, Malone KE, Doody DR, Daling JR, Ostrander EA: Frequency of CHEK2 mutations in a population based, case-control study of breast cancer in young women. Breast cancer research: BCR 2004; 6(6): R629-635.
In article      CrossRefPubMed
[21]  Offit K, Pierce H, Kirchhoff T, Kolachana P, Rapaport B, Gregersen P, Johnson S, Yossepowitch O, Huang H, Satagopan J et al: Frequency of CHEK2* 1100delC in New York breast cancer cases and controls. BMC medical genetics 2003; 4:1.
In article      CrossRefPubMed
[22]  Wagenius M, Borg A, Johansson L, Giwercman A, Bratt O: CHEK2* 1100delC is not an important high-risk gene in families with hereditary prostate cancer in southern Sweden. Scandinavian journal of urology and nephrology 2006; 40(1): 23-25.
In article      CrossRefPubMed
[23]  Einarsdottir K, Humphreys K, Bonnard C, Palmgren J, Iles MM, Sjolander A, Li Y, Chia KS, Liu ET, Hall P et al: Linkage disequilibrium mapping of CHEK2: common variation and breast cancer risk. PLoS medicine 2006; 3(6): e168.
In article      CrossRefPubMed
[24]  Shaag A, Walsh T, Renbaum P, Kirchhoff T, Nafa K, Shiovitz S, Mandell JB, Welcsh P, Lee MK, Ellis N et al: Functional and genomic approaches reveal an ancient CHEK2 allele associated with breast cancer in the Ashkenazi Jewish population. Human molecular genetics 2005; 14(4): 555-563.
In article      CrossRefPubMed
[25]  Kleibl Z, Novotny J, Bezdickova D, Malik R, Kleiblova P, Foretova L, Petruzelka L, Ilencikova D, Cinek P, Pohlreich P: The CHEK2 c.1100delC germline mutation rarely contributes to breast cancer development in the Czech Republic. Breast cancer research and treatment 2005; 90(2): 165-167.
In article      CrossRefPubMed
[26]  Bernstein JL, Teraoka SN, John EM, Andrulis IL, Knight JA, Lapinski R, Olson ER, Wolitzer AL, Seminara D, Whittemore AS et al: The CHEK2* 1100delC allelic variant and risk of breast cancer: screening results from the Breast Cancer Family Registry. Cancer epidemiology, biomarkers & prevention: a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology 2006; 15(2): 348-352.
In article      
  • CiteULikeCiteULike
  • MendeleyMendeley
  • StumbleUponStumbleUpon
  • Add to DeliciousDelicious
  • FacebookFacebook
  • TwitterTwitter
  • LinkedInLinkedIn