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Spectrum of mismatch repair gene mutations and clinical presentation of Hispanic individuals with Lynch syndrome
Accepted Manuscript Title: Spectrum of Mismatch Repair Gene Mutations and Clinical Presentation of Hispanic Individuals with Lynch syndrome Author: Annette Y. Sunga, Charité Ricker, Carin R. Espenschied, Danielle Castillo, Marilena Melas, Josef Herzog, Sarah Bannon, Marcia Cruz-Correa, Patrick Lynch, Ilana Solomon, Stephen B. Gruber, Jeffrey N. Weitzel PII: DOI: Reference:
S2210-7762(17)30021-2 http://dx.doi.org/doi: 10.1016/j.cancergen.2017.01.003 CGEN 514
To appear in:
Cancer Genetics
Received date: Revised date: Accepted date:
19-4-2016 13-1-2017 15-1-2017
Please cite this article as: Annette Y. Sunga, Charité Ricker, Carin R. Espenschied, Danielle Castillo, Marilena Melas, Josef Herzog, Sarah Bannon, Marcia Cruz-Correa, Patrick Lynch, Ilana Solomon, Stephen B. Gruber, Jeffrey N. Weitzel, Spectrum of Mismatch Repair Gene Mutations and Clinical Presentation of Hispanic Individuals with Lynch syndrome, Cancer Genetics (2017), http://dx.doi.org/doi: 10.1016/j.cancergen.2017.01.003. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Title: Spectrum of Mismatch Repair Gene Mutations and Clinical Presentation of Hispanic Individuals with Lynch syndrome
Short Title: Mismatch Repair Mutations in Hispanics Authors: Annette Y. Sunga*1 and Charité Ricker*2, Carin R. Espenschied1, Danielle Castillo1, Marilena Melas2, Josef Herzog1, Sarah Bannon3, Marcia Cruz-Correa4, Patrick Lynch3, Ilana Solomon1, Stephen B. Gruber2, Jeffrey N. Weitzel1 Author affiliations: 1. Clinical Cancer Genetics, City of Hope National Medical Center, Duarte, CA, 91010, USA 2. USC Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA 90089, USA 3. Clinical Cancer Genetics, MD Anderson Cancer Center, Houston, TX 77030, USA 4. Clinical Cancer Genetics, University of Puerto Rico Comprehensive Cancer Center, Rio Piedras, Puerto Rico 00935 *Contributed equally Corresponding Author: Jeffrey N. Weitzel Division of Clinical Cancer Genetics Mod 173 City of Hope 1500 E. Duarte Rd., Duarte, CA 91010 Phone: (626) 256-8662 Fax: (626) 930-5495 [email protected] Conflict of Interest Statement: The authors have no conflicts of interest to disclose. Carin R. Espenschied is currently employed by Ambry Genetics, however, work on this project was completed while employed at City of Hope.
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Highlights Nine recurrent Hispanic mutations were observed in sample of a Hispanic patients seen for genetic cancer risk assessment Three of these were previously reported in Spain, confirming the influence of Spanish ancestry of Hispanic populations This is the largest report of Hispanic MMR mutations in North America Abstract Lynch syndrome (LS), the most common hereditary colorectal cancer syndrome, is caused by mismatch repair (MMR) gene mutations. However, data about MMR mutations in Hispanics are limited. This study aims to describe the spectrum of MMR mutations in Hispanics with LS and explore ancestral origins. This case series involved an IRB-approved retrospective chart review of self-identified Hispanic patients (n=397) seen for genetic cancer risk assessment at four collaborating academic institutions in California, Texas, and Puerto Rico who were evaluated by MMR genotyping and/or tumor analysis. A literature review was conducted for all mutation identified. Of those who underwent clinical genetic testing (n=176), 71 had MMR gene mutations. Nine mutations were observed more than once. One third (3/9) of recurrent mutations and two additional mutations (seen only once) were previously reported in Spain, confirming the influence of Spanish ancestry on MMR mutations in Hispanic populations. The recurrent mutations identified (n=9) included both previously reported mutations as well as unique mutations not in the literature. This is the largest report of Hispanic MMR mutations in North America; however, a larger sample and haplotype analyses are needed to better understand recurrent MMR mutations in Hispanic populations.
Keywords: MMR Mutations, Lynch Syndrome, Hispanics, MLH1, MSH2, MSH6, PMS2, Colon Cancer 2
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Abbreviations
GCRA: Genetic Cancer Risk Assessment HNPCC: Hereditary non-Polyposis Colon Cancer IHC: Immunohistochemistry LS: Lynch syndrome MMR: Mismatch Repair MSI: Microsatellite instability testing VUS: Variants of Uncertain Significance
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Introduction Individuals of Hispanic or Latino ancestry represent the largest and fastest growing ethnic minority group in the United States [1]. More than 50 million Americans self-identify as Hispanic or Latino, accounting for 16% of the population [2]. It is projected that 30% of the total U.S. population will identify as Hispanic by 2050. There is great heterogeneity amongst individuals identifying as Hispanic. The term is used in aggregate for individuals of Mexican, Central and South American, Caribbean, or Spanish ancestry. The underlying population varies greatly not only by country of origin, but also by cultural identity. As the population continues to grow, along with the corresponding cancer burden, it is increasingly important to expand our understanding of cancer risk factors and etiologies in the Hispanic population. Colorectal cancer is the second most commonly diagnosed cancer in Hispanics and endometrial cancer is the most common gynecologic cancer in Hispanic women [3]. While most colorectal and endometrial cancers are not associated with inherited cancer syndromes, it is estimated that approximately 3-5% are attributable to Lynch syndrome (LS) or hereditary non-polyposis colorectal cancer (HNPCC) (OMIM#120435). LS is caused by mutations in MLH1, MSH2, MSH6, PMS2, and EPCAM. The range of lifetime cancer risks faced by individuals with LS are 40-80% for colorectal cancer, 1560% for endometrial cancer, 11-24% for ovarian cancer, and 3-13% for gastric cancer, all of which are substantially elevated over the general population [4-9]. Despite these elevated risks, appropriate cancer screening and risk-reducing interventions can
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substantially decrease the probability of developing cancer and increase the chance of a favorable prognosis for those who do develop cancer [10, 11]. LS has been reported worldwide, occurring in individuals of all ethnic and racial backgrounds [12-18]. However, data are limited about the mutations associated with LS in Hispanics, as the majority of the extant literature has been focused on non-Hispanic individuals of European ancestry. A large body of literature on Spanish LS cohorts exists [14, 19-22], but the literature outside of Spain has been limited to just a few South American studies and one in the Caribbean [23-28]. While data from Hispanic BRCA1/2 cohorts demonstrate that Spanish ancestral ties are manifested through specific recurrent mutations, there are other common mutations that are likely Amerindian in origin [29]. We hypothesized that this may also be true in LS. In order to better understand the spectrum of mutations associated with LS in Hispanics in the US and Puerto Rico, a collaborative clinic-based cohort was assembled from four institutions. Materials and Methods Our sample was ascertained from four centers serving Hispanic populations. City of Hope (COH) cases included patients seen for genetic cancer risk assessment (GCRA) at collaborating institutions of the COH Clinical Cancer Genomics Community Research Network between 1996 and 2014. University of Southern California (USC) cases included patients who received cancer genetic services at two facilities, USC Norris Comprehensive Cancer Center and Los Angeles County + USC Medical Center, from 2007-2014. The University of Texas: MD Anderson Cancer Clinical Cancer Genetics Program (MDA) maintains a comprehensive database of all patients seen for genetic counseling and genetic testing since 1999. The Puerto Rico Familiar Colorectal 5
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Cancer Registry (PURIFICAR) identifies newly diagnosed colorectal cancer cases through the Puerto Rico Cancer Registry (PRCCR). We retrospectively collected data under IRB approved protocols at each institution (COH-#96144; USC 0S-12-4; MDA MTA#: 10334; PR #A2210207) on all probands that were of Hispanic ancestry and who were assessed for LS at their respective facilities as part of their clinical care. This assessment was defined as microsatellite instability testing (MSI) and/or immunohistochemistry (IHC) of the MMR gene protein products and/or genotyping for one or more of the MMR genes (MLH1, MSH2, MSH6, PMS2) and EpCAM. Germline genetic testing was ordered as deemed clinically indicated, based on abnormal IHC staining patterns or clinical testing criteria. Clinical criteria utilized included Amsterdam criteria, Bethesda guidelines and NCCN consensus guidelines that are annually updated [30-34]. All germline genetic testing was performed at CLIA approved laboratories. The number of MMR genes analyzed varied based on the results of IHC, insurance coverage, and genes clinically available at the time of testing. Sequence alterations were classified by each CLIA-approved laboratory, based on their established variant assessment process. For consistency, all alterations were converted to the current Human Genome Variation Society and the American College of Medical Genetics and Genomics for variant nomenclature and interpretation [35, 36]. A comprehensive literature review was conducted for all mutations in the cohort using PubMed, Google Scholar, the InSiGHT database, and the Mismatch Repair Genes Variant database [37, 38]. Ethnicity was recorded for all cases reported in the literature. When the specific ethnicity of the case was not described in a publication, the 6
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country(ies) of the authors was/were recorded. Descriptive statistics were performed using SPSS version 19 (IBM Corp. Released 2010. IBM SPSS Statistics for Windows, Version 19.0 Armonk, NY: IBM Corp.). Results Demographics and clinical features Tumor studies (MSI and/or IHC) and/or genotyping of one or more of the MMR genes were performed in 397 individuals of Hispanic ancestry (Table 1 and Figure 1). Most participants reported Mexico as their country of origin (n=124, 31.2%), followed by Puerto Rico (n=64, 16.1%), Guatemala (n=20, 5.0%), and El Salvador (n=19, 4.8%). A majority of participants (n=381, 96.0%) were diagnosed with at least one tumor, with colorectal cancer being the most common (n=296, 74.6%). The mean age of cancer diagnosis for participants was 43.6 years. The majority of individuals with a cancer diagnosis (n=344, 90.2%) had MSI and/or IHC performed on their tumor. The results of these tumor studies were classified as MMR proficient if all proteins were expressed by IHC and/or the tumor was MSI stable or low. Tumors that exhibited loss of one of more of the MMR proteins by IHC or were MSI high were classified as MMR deficient. Figure 1 details these studies across the cohort, as well as subsequent germline findings. In our multi-center cohort, 79 MMR gene sequence alterations were identified in 77 probands from unrelated Hispanic families. Seventy-one alternations were classified as pathogenic or likely pathogenic and nine as variants of unknown significance (VUS; Table 2, Appendix). Subsequent analyses focused on the 71 individuals with pathogenic and likely pathogenic mutations. Sixty-one had a cancer diagnosis; 66.2% 7
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had colorectal cancer, 12.7% (n=9) had uterine cancer, 1.4% (n=1) had gastric cancer, and 5.6% (n=4) had other cancers (Table 1). The majority (57.7%) of the affected mutation carriers had one LS associated cancer, while 18.3% (n=13) individuals had two, and seven (9.9%) had three or more LS associated cancers (Table 1). MMR gene mutations In our cohort, 71 people were identified to have pathogenic or likely pathogenic mutations (Table 2), representing 57 distinct mutations (Appendix). Of these, 29 were in MLH1, 17 in MSH2, 7 in MSH6, and 4 in PMS2. The mutations were mostly frameshift (n=17, 29.8%) and nonsense (n=15, 26.3%), followed by splice site (n=12, 21.1%), large exonic deletions (n=7, 12.3%), and missense mutations (n=6, 10.5%) (Appendix). Nine mutations (MLH1 c.350C>T, c.588+5G>A, c.1024_1039del16, c.1790_1791del2ins9, c.1852_1854delAAG, c.2041G>A, and MSH2 c.425C>G, c.1216C>T, c.1705_1706delGA) were observed more than once (Table 3). MSH2 c.1216C>T was seen five times (at three different institutions) with all families reporting Mexico as their country of origin. MLH1 c.1852_1854delAAG was seen four times, with all families reportedly from El Salvador. Though seen in two unrelated families at one institution in our series, to our knowledge, MLH1 c.1790_1791del2ins9 has not been previously reported in the literature or MMR mutation databases, nor have 17 other mutations identified in our cohort (Appendix). Discussion
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This multi-institutional collaboration is the first to describe the spectrum of mutations in Hispanic LS patients in North America. MLH1 c.350C>T and MSH2 c.1216C>T have been reported multiple times in European studies (including Spain), as well as in US, South American, and Chinese studies [18, 21, 23, 39-42]. These two mutations are within CpG dinucleotides, which are known to cause mutational hotspots, and may explain why they were seen previously in multiple ethnicities as well as our cohort [43]. MLH1 c.1852_1854delAAG has been reported in France, South America, the Netherlands and others [8,15,24,44,45]. MLH1 c.350C>T,[21] c.2041G>A [42], c.332C>T [46], and c.676C>T [47] and MSH2 c.1216C>T [20, 21, 41, 48], deletion of exons 4-8 [20, 49], and deletion of exon 8 [21], and PMS2 deletion of exon 14 have all been reported previously in Spain [50]. The MSH2 deletion of exons 4-8 was seen in one Mexican family in our series and has been reported as a Spanish founder mutation [20]. The fact that several mutations in our cohort were previously reported in Spain is consistent with established migration patterns of Europeans, especially those from Spain, settling in Mexico and Central America. Obtaining a better understanding of penetrance and expression of MMR gene mutations in Hispanic families is a natural extension of this collaborative project. Many of the individuals included in this cohort were ascertained from colorectal cancer clinics and/or registries, such that other non-colon cancer, LS associated tumors may be underrepresented. A more expanded cohort with wider representation of individuals with other LS tumors is of interest.
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While all cases in this clinical series had a test performed to assess for LS (either tumor studies and/or germline genetic testing), both types of analyses were not available for all individuals. In addition, due to the retrospective nature of this study not all individuals in the cohort underwent complete germline analysis of the MMR genes because of the variability in institutional protocols for tumor testing, insurance coverage or accessibility, and availability of germline testing for specific genes over time (e.g. PMS2). Therefore, there may be individuals in this cohort with germline mutations that have not been identified. A limitation of this study is that clinical information, such as multigenerational family history and tumor histological features, was incomplete for two sites, so it is not reported. Finally, as this is a clinical study cohort, mutation prevalence cannot be estimated. Conclusion Findings from this collaborative study add to the understanding of recurrent MMR mutations in Hispanic populations. The presence of mutations in this cohort that were previously reported in Spanish and European populations reflects the ancestral connections of much of Latin America with Spain and Europe. However, the number of unique mutations, which have not been reported previously, suggest a rich area of further research to determine if there are potential Amerindian founder mutations. While this is the largest reported cohort of Hispanic patients with MMR mutations in North America, a larger sample and haplotype analyses are needed to better define the spectrum and origin of mutations in Hispanic populations.
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Acknowledgements: Research reported in this publication was supported by the National Cancer Institute of the National Institutes of Health under award number P30CA33572. Patients were recruited for study from the COH Clinical Cancer Genomics Community Research Network (CCGCRN), supported in part by award number RC4CA153828 (PI: J. Weitzel) from the National Cancer Institute and the Office of the Director, National Institutes of Health. The Hispanic MMR project is supported in part by a grant from The Safeway Foundation. USC research activities are supported by the Norris Cancer Center Core Grant (NCI P30CA014089) and the Anton B. Burg Foundation. The Puerto Rico Familiar Colorectal Cancer Registry was funded in part by a pilot NCI grant under award number R03 CA130034. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Cancer Institute or the National Institutes of Health. We would like to acknowledge the efforts of Lacolle Robinson and Kai Yang for data collection and Lily R Van Tongeren and Christina Ryback for manuscript preparation and review.
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[25] Alvarez K, Hurtado C, Hevia MA, Wielandt AM, de la Fuente M, Church J, Carvallo P, Lopez-Kostner F. Spectrum of MLH1 and MSH2 mutations in Chilean families with suspected Lynch syndrome. Dis Colon Rectum 2010;53:450-9. [26] da Silva FC, de Oliveira LP, Santos EM, Nakagawa WT, Aguiar Junior S, Valentin MD, Rossi BM, de Oliveira Ferreira F. Frequency of extracolonic tumors in Brazilian families with Lynch syndrome: analysis of a hereditary colorectal cancer institutional registry. Fam Cancer 2010;9:563-70. [27] Giraldo A, Gomez A, Salguero G, Garcia H, Aristizabal F, Gutierrez O, Angel LA, Padron J, Martinez C, Martinez H, Malaver O, Florez L, Barvo R. MLH1 and MSH2 mutations in Colombian families with hereditary nonpolyposis colorectal cancer (Lynch syndrome)-description of four novel mutations. Fam Cancer 2005;4:285-90. [28] Cruz-Correa M, Diaz-Algorri Y, Perez-Mayoral J, Suleiman-Suleiman W, Del Mar Gonzalez-Pons M, Bertran C, Casellas N, Rodriguez N, Pardo S, Rivera K, Mosquera R, Rodriguez-Quilichini S. Clinical characterization and mutation spectrum in Caribbean Hispanic families with Lynch syndrome. Fam Cancer 2015;14:415-25. [29] Weitzel JN, Clague J, Martir-Negron A, Ogaz R, Herzog J, Ricker C, Jungbluth C, Cina C, Duncan P, Unzeitig G, Saldivar JS, Beattie M, Feldman N, Sand S, Port D, Barragan DI, John EM, Neuhausen SL, Larson GP. Prevalence and type of BRCA mutations in Hispanics undergoing genetic cancer risk assessment in the southwestern United States: a report from the Clinical Cancer Genetics Community Research Network. J Clin Oncol 2013;31:210-6. [30] Provenzale D, Gupta S, Ahnen DJ, Bray T, Cannon JA, Cooper G, David DS, Early DS, Erwin D, Ford JM, Giardiello FM, Grady W, Halverson AL, Hamilton SR, Hampel H, Ismail MK, Klapman JB, Larson DW, Lazenby AJ, Lynch PM, Mayer RJ, Ness RM, Regenbogen SE, Samadder NJ, Shike M, Steinbach G, Weinberg D, Dwyer M, Darlow S. Genetic/Familial HighRisk Assessment: Colorectal Version 1.2016, NCCN Clinical Practice Guidelines in Oncology. J Natl Compr Canc Netw 2016;14:1010-30. [31] Vasen HF, Mecklin JP, Khan PM, Lynch HT. The International Collaborative Group on Hereditary Non-Polyposis Colorectal Cancer (ICG-HNPCC). Dis Colon Rectum 1991;34:424-5. [32] Vasen HFA, Watson P, Mecklin J-P, Lynch HT, (ICG-HNPCC). New clinical criteria for hereditary nonpolyposis colorectal cancer (HNPCC, Lynch Syndrome) proposed by the International Collaborative Group on HNPCC. Gastroenterology 1999;116:1453-6. [33] Rodriguez-Bigas MA, Boland CR, Hamilton SR, Henson DE, Jass JR, Khan PM, Lynch H, Perucho M, Smyrk T, Sobin L, Srivastava S. A National Cancer Institute Workshop on Hereditary Nonpolyposis Colorectal Cancer Syndrome: meeting highlights and Bethesda guidelines. J Natl Cancer Inst 1997;89:1758-62. [34] Umar A, Boland CR, Terdiman JP, Syngal S, de la Chapelle A, Ruschoff J, Fishel R, Lindor NM, Burgart LJ, Hamelin R, Hamilton SR, Hiatt RA, Jass J, Lindblom A, Lynch HT, Peltomaki P, Ramsey SD, Rodriguez-Bigas MA, Vasen HF, Hawk ET, Barrett JC, Freedman AN, Srivastava S. Revised Bethesda Guidelines for hereditary nonpolyposis colorectal cancer (Lynch Syndrome) and microsatellite instability. J Natl Cancer Inst 2004;96:261-8. [35] Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, Grody WW, Hegde M, Lyon E, Spector E, Voelkerding K, Rehm HL. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet Med 2015;17:40524. [36] den Dunnen JT, Dalgleish R, Maglott DR, Hart RK, Greenblatt MS, McGowan-Jordan J, Roux AF, Smith T, Antonarakis SE, Taschner PE. HGVS Recommendations for the Description of Sequence Variants: 2016 Update. Human mutation 2016;37:564-9. [37] International Society for Gastrointestinal Hereditary Tumours. Colon cancer gene variant databases. InSiGHT Variants databases: Leiden University Medical Center, 2008. 14
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[38] Woods MO, Williams P, Careen A, Edwards L, Bartlett S, McLaughlin JR, Younghusband HB. A new variant database for mismatch repair genes associated with Lynch syndrome. Hum Mutat 2007;28:669-73. [39] Sarroca C, Valle AD, Fresco R, Renkonen E, Peltomaki P, Lynch H. Frequency of hereditary non-polyposis colorectal cancer among Uruguayan patients with colorectal cancer. Clin Genet 2005;68:80-7. [40] Auclair J, Busine MP, Navarro C, Ruano E, Montmain G, Desseigne F, Saurin JC, Lasset C, Bonadona V, Giraud S, Puisieux A, Wang Q. Systematic mRNA analysis for the effect of MLH1 and MSH2 missense and silent mutations on aberrant splicing. Hum Mutat 2006;27:145-54. [41] Grindedal EM, Blanco I, Stormorken A, Maehle L, Clark N, Gonzalez S, Capella G, Vasen H, Burn J, Moller P. High risk of endometrial cancer in colorectal cancer kindred is pathognomonic for MMR-mutation carriers. Fam Cancer 2009;8:145-51. [42] Rodriguez-Soler M, Perez-Carbonell L, Guarinos C, Zapater P, Castillejo A, Barbera VM, Juarez M, Bessa X, Xicola RM, Clofent J, Bujanda L, Balaguer F, Rene JM, de-Castro L, MarinGabriel JC, Lanas A, Cubiella J, Nicolas-Perez D, Brea-Fernandez A, Castellvi-Bel S, Alenda C, Ruiz-Ponte C, Carracedo A, Castells A, Andreu M, Llor X, Soto JL, Paya A, Jover R. Risk of cancer in cases of suspected lynch syndrome without germline mutation. Gastroenterology 2013;144:926-32 e1; quiz e13-4. [43] Maliaka YK, Chudina AP, Belev NF, Alday P, Bochkov NP, Buerstedde JM. CpG dinucleotides in the hMSH2 and hMLH1 genes are hotspots for HNPCC mutations. Hum Genet 1996;97:251-5. [44] Syngal S, Fox EA, Li C, Dovidio M, Eng C, Kolodner RD, Garber JE. Interpretation of genetic test results for hereditary nonpolyposis colorectal cancer: Implications for clinical predisposition testing. JAMA-J Am Med Assoc 1999;282:247-53. [45] Wagner A, Tops C, Wijnen JT, Zwinderman K, van der Meer C, Kets M, Niermeijer MF, Klijn JG, Tibben A, Vasen HF, Meijers-Heijboer H. Genetic testing in hereditary non-polyposis colorectal cancer families with a MSH2, MLH1, or MSH6 mutation. J Med Genet 2002;39:833-7. [46] Caldes T, Godino J, de la Hoya M, Garcia Carbonero I, Perez Segura P, Eng C, Benito M, Diaz-Rubio E. Prevalence of germline mutations of MLH1 and MSH2 in hereditary nonpolyposis colorectal cancer families from Spain. Int J Cancer 2002;98:774-9. [47] Marcos I, Borrego S, Urioste M, Garcia-Valles C, Antinolo G. Mutations in the DNA mismatch repair gene MLH1 associated with early-onset colon cancer. J Pediatr 2006;148:8379. [48] Martinez-Bouzas C, Beristain E, Ojembarrena E, Errasti J, Mujika K, Viguera N, Tejada MI. A study on MSH2 and MLH1 mutations in hereditary nonpolyposis colorectal cancer families from the Basque Country, describing four new germline mutations. Fam Cancer 2009;8:533-9. [49] Perez-Cabornero L, Sanz MI, Sampedro EV, Aras EL, Becares AA, Pino CM, Dominguez MD. Frequency of rearrangements in Lynch syndrome cases associated with MSH2: characterization of a new deletion involving both EPCAM and the 5' part of MSH2. Cancer Prev Res (Phila) 2011;4:1556-62. [50] Brea-Fernandez AJ, Cameselle-Teijeiro JM, Alenda C, Fernandez-Rozadilla C, Cubiella J, Clofent J, Rene JM, Anido U, Mila M, Balaguer F, Castells A, Castellvi-Bel S, Jover R, Carracedo A, Ruiz-Ponte C. High incidence of large deletions in the PMS2 gene in Spanish Lynch syndrome families. Clin Genet 2014;85:583-8.
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Figure 1: Flowsheet of Tumor Studies and Germline Genetic Testing Performed in Cohort
Individuals Evaluated for Lynch Syndromea N=397
Family History / NO Personal History of Cancer N=16
Personal History of Cancer N=381
IHC and/or MSI on Tumor N=344
No Tumor Studies N=37
MMR Proficient N=243
NO Genetic Testing N=193
Genetic Testing N=37
b
VUS N=1 Negative N=18
Positive N=18
c
Genetic Testing N=50
VUS N=2 Negative N=47
Positive N=1
MMR Deficient N=101
NO Genetic Testing N=28
VUS N=4 Negative N=27
Genetic Testing N=73
Positive d N=42
Genetic Testing N=16
VUS N=0 Negative N=6
Positive N=10
16
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a Each
individual is only represented once = Variant of uncertain significance c Positive = Likely pathogenic and pathogenic alterations d Two individuals with a positive result also had a VUS, but are categorized in this figure based on the positive result b VUS
TABLE 1. Demographics of Study Cohort
Variable
Sequence negative or Only tumor studies [n (%)] 326 (82.4)
MMR gene mutation carrier [n (%)] 71 (17.9)
[n (%)]
42.98 (10.8)
43.39 (9.3)
43.1
≤ 40
102 (31.3)
28 (39.4)
130 (32.7)
>40
224 (68.7)
43 (60.6)
267 (67.3)
Female
162 (49.7)
40 (56.3)
202 (50.9)
Male
106 (32.5)
31 (43.7)
137 (34.5)
COH
70 (21.5)
19 (26.8)
89 (22.4)
MDA
92 (28.2)
14 (19.7)
106 (26.7)
USC
106 (32.5)
31 (43.7)
137 (34.5)
PR
58 (17.8)
7 (9.9)
65 (16.4)
Affected
315 (96.6)
61 (85.9)
381 (96.0)
Unaffected
6 (1.8)
10 (14.1)
16 (4.0)
Colorectal Uterine Ovarian Gastric Breast Other tumorc
249 (76.4) 31 (9.5) 8 (2.5) 8 (2.5) 6 (1.8) 13 (4.0)
47 (66.2) 9 (12.7) 0 1 (1.4) 0 4 (5.6)
296 (74.6) 40 (10.1) 8 (2.0) 9 (2.3) 6 (1.5) 22 (5.5)
1 2 ≥3
278 (85.3) 31 (9.5) 5 (1.5)
41 (57.7) 13 (18.3) 7 (9.9)
319 (80.4) 44 (11.1) 12 (3.0)
Category
N (%) Mean age (SD) Age
Gendera
Institution
Disease Status
First Cancer Diagnosis in Probandsb
Total
397
Number of Lynch syndrome Associated Tumorsd
17
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Country of Origin
a
Mexico
84 (25.8)
40 (56.3)
124 (31.2)
Puerto Rico El Salvador Guatemala Othere
59 (18.1) 13 (4.0) 14 (4.3) 156 (47.9)
5 (7.0) 6 (8.5) 6 (8.5) 14 (19.7)
64 (16.1) 19 (4.8) 20 (5.0) 170 (42.8)
Gender was missing for 58 individuals Individuals with multiple primaries were classified based on their first cancer diagnosis c Includes bladder cancer, glioblastoma multiforme, Hodgkin's lymphoma, kidney cancer, sarcoma, sebaceous carcinoma, small bowel cancer, ureter cancer, and unknown. d Non-LS associated tumors were excluded from the multiple primary cancers e Includes individuals from the Dominican Republic, Ecuador, Peru, Colombia, Spain, Honduras, Cuba, Chile, Nicaragua and those with partial or unspecified Hispanic ancestry b
TABLE 2. Characteristics of MMR Gene Mutations Affected
Unaffected
Total
61
10
71
Pathogenic
54
10
64
Likely pathogenic
7
0
7
MLH1 MSH2 MSH6 PMS2 Yes No
31 22 7 1 22 39
6 1 0 3 1 9
37 23 7 4 23 48
Probands
Classification
Gene
Recurrent
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TABLE 3: Recurrent and Previously Published Pathogenic/Likely Pathogenic Mutations Gene
Mutation
Frequency
Institution
Reported Country of Origin in Study Cohort
MLH1
c.350C>T
2
COH
Mexico
c.588+5G>A
2
MDA, USC
Mexico
c.676C>T
1*
USC
Mexico
c.1024_1039del16 c.1790_1791del2ins9
2 2
PR MDA
Puerto Rico Mexico
c.1852_1854delAAG
4
MDA, USC
El Salvador
c.2041G>A
2
COH, USC
c.425C>G
2
COH, USC
Exon 4-8 del
1*
COH
Mexico & Guatemala Mexico & Guatemala Mexico
c.1216C>T
5
COH, MDA, USC
Mexico
c.1705_1706delGA
2
MDA, PR
Exon 8 del Exon 14 del
1* 1*
MDA COH
MSH2
PMS2
Mexico & Puerto Rico El Salvador Mexico
Reported Country of Origin in Previous Publications Spain, South America, Europe, Japan, China Europe Spain, South America, Europe, USA Italy None South America, Europe, Japan, US South America, Spain, Chile, Columbia, Europe, Japan None Spain Spain, Uruguay, South America, Europe, China, US Europe Spain, Europe, USA, Canada Spain
Italicized highlights Spain and South American countries. * Not recurrent in this study, but previously cited in publications linked to South America and/or Spain. Abbreviations: COH, City of Hope; USC, University of Southern California; PR, Puerto Rico; MDA, MD Anderson
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S2210-7762(17)30021-2 http://dx.doi.org/doi: 10.1016/j.cancergen.2017.01.003 CGEN 514
To appear in:
Cancer Genetics
Received date: Revised date: Accepted date:
19-4-2016 13-1-2017 15-1-2017
Please cite this article as: Annette Y. Sunga, Charité Ricker, Carin R. Espenschied, Danielle Castillo, Marilena Melas, Josef Herzog, Sarah Bannon, Marcia Cruz-Correa, Patrick Lynch, Ilana Solomon, Stephen B. Gruber, Jeffrey N. Weitzel, Spectrum of Mismatch Repair Gene Mutations and Clinical Presentation of Hispanic Individuals with Lynch syndrome, Cancer Genetics (2017), http://dx.doi.org/doi: 10.1016/j.cancergen.2017.01.003. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Title: Spectrum of Mismatch Repair Gene Mutations and Clinical Presentation of Hispanic Individuals with Lynch syndrome
Short Title: Mismatch Repair Mutations in Hispanics Authors: Annette Y. Sunga*1 and Charité Ricker*2, Carin R. Espenschied1, Danielle Castillo1, Marilena Melas2, Josef Herzog1, Sarah Bannon3, Marcia Cruz-Correa4, Patrick Lynch3, Ilana Solomon1, Stephen B. Gruber2, Jeffrey N. Weitzel1 Author affiliations: 1. Clinical Cancer Genetics, City of Hope National Medical Center, Duarte, CA, 91010, USA 2. USC Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA 90089, USA 3. Clinical Cancer Genetics, MD Anderson Cancer Center, Houston, TX 77030, USA 4. Clinical Cancer Genetics, University of Puerto Rico Comprehensive Cancer Center, Rio Piedras, Puerto Rico 00935 *Contributed equally Corresponding Author: Jeffrey N. Weitzel Division of Clinical Cancer Genetics Mod 173 City of Hope 1500 E. Duarte Rd., Duarte, CA 91010 Phone: (626) 256-8662 Fax: (626) 930-5495 [email protected] Conflict of Interest Statement: The authors have no conflicts of interest to disclose. Carin R. Espenschied is currently employed by Ambry Genetics, however, work on this project was completed while employed at City of Hope.
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Highlights Nine recurrent Hispanic mutations were observed in sample of a Hispanic patients seen for genetic cancer risk assessment Three of these were previously reported in Spain, confirming the influence of Spanish ancestry of Hispanic populations This is the largest report of Hispanic MMR mutations in North America Abstract Lynch syndrome (LS), the most common hereditary colorectal cancer syndrome, is caused by mismatch repair (MMR) gene mutations. However, data about MMR mutations in Hispanics are limited. This study aims to describe the spectrum of MMR mutations in Hispanics with LS and explore ancestral origins. This case series involved an IRB-approved retrospective chart review of self-identified Hispanic patients (n=397) seen for genetic cancer risk assessment at four collaborating academic institutions in California, Texas, and Puerto Rico who were evaluated by MMR genotyping and/or tumor analysis. A literature review was conducted for all mutation identified. Of those who underwent clinical genetic testing (n=176), 71 had MMR gene mutations. Nine mutations were observed more than once. One third (3/9) of recurrent mutations and two additional mutations (seen only once) were previously reported in Spain, confirming the influence of Spanish ancestry on MMR mutations in Hispanic populations. The recurrent mutations identified (n=9) included both previously reported mutations as well as unique mutations not in the literature. This is the largest report of Hispanic MMR mutations in North America; however, a larger sample and haplotype analyses are needed to better understand recurrent MMR mutations in Hispanic populations.
Keywords: MMR Mutations, Lynch Syndrome, Hispanics, MLH1, MSH2, MSH6, PMS2, Colon Cancer 2
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Abbreviations
GCRA: Genetic Cancer Risk Assessment HNPCC: Hereditary non-Polyposis Colon Cancer IHC: Immunohistochemistry LS: Lynch syndrome MMR: Mismatch Repair MSI: Microsatellite instability testing VUS: Variants of Uncertain Significance
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Introduction Individuals of Hispanic or Latino ancestry represent the largest and fastest growing ethnic minority group in the United States [1]. More than 50 million Americans self-identify as Hispanic or Latino, accounting for 16% of the population [2]. It is projected that 30% of the total U.S. population will identify as Hispanic by 2050. There is great heterogeneity amongst individuals identifying as Hispanic. The term is used in aggregate for individuals of Mexican, Central and South American, Caribbean, or Spanish ancestry. The underlying population varies greatly not only by country of origin, but also by cultural identity. As the population continues to grow, along with the corresponding cancer burden, it is increasingly important to expand our understanding of cancer risk factors and etiologies in the Hispanic population. Colorectal cancer is the second most commonly diagnosed cancer in Hispanics and endometrial cancer is the most common gynecologic cancer in Hispanic women [3]. While most colorectal and endometrial cancers are not associated with inherited cancer syndromes, it is estimated that approximately 3-5% are attributable to Lynch syndrome (LS) or hereditary non-polyposis colorectal cancer (HNPCC) (OMIM#120435). LS is caused by mutations in MLH1, MSH2, MSH6, PMS2, and EPCAM. The range of lifetime cancer risks faced by individuals with LS are 40-80% for colorectal cancer, 1560% for endometrial cancer, 11-24% for ovarian cancer, and 3-13% for gastric cancer, all of which are substantially elevated over the general population [4-9]. Despite these elevated risks, appropriate cancer screening and risk-reducing interventions can
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substantially decrease the probability of developing cancer and increase the chance of a favorable prognosis for those who do develop cancer [10, 11]. LS has been reported worldwide, occurring in individuals of all ethnic and racial backgrounds [12-18]. However, data are limited about the mutations associated with LS in Hispanics, as the majority of the extant literature has been focused on non-Hispanic individuals of European ancestry. A large body of literature on Spanish LS cohorts exists [14, 19-22], but the literature outside of Spain has been limited to just a few South American studies and one in the Caribbean [23-28]. While data from Hispanic BRCA1/2 cohorts demonstrate that Spanish ancestral ties are manifested through specific recurrent mutations, there are other common mutations that are likely Amerindian in origin [29]. We hypothesized that this may also be true in LS. In order to better understand the spectrum of mutations associated with LS in Hispanics in the US and Puerto Rico, a collaborative clinic-based cohort was assembled from four institutions. Materials and Methods Our sample was ascertained from four centers serving Hispanic populations. City of Hope (COH) cases included patients seen for genetic cancer risk assessment (GCRA) at collaborating institutions of the COH Clinical Cancer Genomics Community Research Network between 1996 and 2014. University of Southern California (USC) cases included patients who received cancer genetic services at two facilities, USC Norris Comprehensive Cancer Center and Los Angeles County + USC Medical Center, from 2007-2014. The University of Texas: MD Anderson Cancer Clinical Cancer Genetics Program (MDA) maintains a comprehensive database of all patients seen for genetic counseling and genetic testing since 1999. The Puerto Rico Familiar Colorectal 5
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Cancer Registry (PURIFICAR) identifies newly diagnosed colorectal cancer cases through the Puerto Rico Cancer Registry (PRCCR). We retrospectively collected data under IRB approved protocols at each institution (COH-#96144; USC 0S-12-4; MDA MTA#: 10334; PR #A2210207) on all probands that were of Hispanic ancestry and who were assessed for LS at their respective facilities as part of their clinical care. This assessment was defined as microsatellite instability testing (MSI) and/or immunohistochemistry (IHC) of the MMR gene protein products and/or genotyping for one or more of the MMR genes (MLH1, MSH2, MSH6, PMS2) and EpCAM. Germline genetic testing was ordered as deemed clinically indicated, based on abnormal IHC staining patterns or clinical testing criteria. Clinical criteria utilized included Amsterdam criteria, Bethesda guidelines and NCCN consensus guidelines that are annually updated [30-34]. All germline genetic testing was performed at CLIA approved laboratories. The number of MMR genes analyzed varied based on the results of IHC, insurance coverage, and genes clinically available at the time of testing. Sequence alterations were classified by each CLIA-approved laboratory, based on their established variant assessment process. For consistency, all alterations were converted to the current Human Genome Variation Society and the American College of Medical Genetics and Genomics for variant nomenclature and interpretation [35, 36]. A comprehensive literature review was conducted for all mutations in the cohort using PubMed, Google Scholar, the InSiGHT database, and the Mismatch Repair Genes Variant database [37, 38]. Ethnicity was recorded for all cases reported in the literature. When the specific ethnicity of the case was not described in a publication, the 6
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country(ies) of the authors was/were recorded. Descriptive statistics were performed using SPSS version 19 (IBM Corp. Released 2010. IBM SPSS Statistics for Windows, Version 19.0 Armonk, NY: IBM Corp.). Results Demographics and clinical features Tumor studies (MSI and/or IHC) and/or genotyping of one or more of the MMR genes were performed in 397 individuals of Hispanic ancestry (Table 1 and Figure 1). Most participants reported Mexico as their country of origin (n=124, 31.2%), followed by Puerto Rico (n=64, 16.1%), Guatemala (n=20, 5.0%), and El Salvador (n=19, 4.8%). A majority of participants (n=381, 96.0%) were diagnosed with at least one tumor, with colorectal cancer being the most common (n=296, 74.6%). The mean age of cancer diagnosis for participants was 43.6 years. The majority of individuals with a cancer diagnosis (n=344, 90.2%) had MSI and/or IHC performed on their tumor. The results of these tumor studies were classified as MMR proficient if all proteins were expressed by IHC and/or the tumor was MSI stable or low. Tumors that exhibited loss of one of more of the MMR proteins by IHC or were MSI high were classified as MMR deficient. Figure 1 details these studies across the cohort, as well as subsequent germline findings. In our multi-center cohort, 79 MMR gene sequence alterations were identified in 77 probands from unrelated Hispanic families. Seventy-one alternations were classified as pathogenic or likely pathogenic and nine as variants of unknown significance (VUS; Table 2, Appendix). Subsequent analyses focused on the 71 individuals with pathogenic and likely pathogenic mutations. Sixty-one had a cancer diagnosis; 66.2% 7
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had colorectal cancer, 12.7% (n=9) had uterine cancer, 1.4% (n=1) had gastric cancer, and 5.6% (n=4) had other cancers (Table 1). The majority (57.7%) of the affected mutation carriers had one LS associated cancer, while 18.3% (n=13) individuals had two, and seven (9.9%) had three or more LS associated cancers (Table 1). MMR gene mutations In our cohort, 71 people were identified to have pathogenic or likely pathogenic mutations (Table 2), representing 57 distinct mutations (Appendix). Of these, 29 were in MLH1, 17 in MSH2, 7 in MSH6, and 4 in PMS2. The mutations were mostly frameshift (n=17, 29.8%) and nonsense (n=15, 26.3%), followed by splice site (n=12, 21.1%), large exonic deletions (n=7, 12.3%), and missense mutations (n=6, 10.5%) (Appendix). Nine mutations (MLH1 c.350C>T, c.588+5G>A, c.1024_1039del16, c.1790_1791del2ins9, c.1852_1854delAAG, c.2041G>A, and MSH2 c.425C>G, c.1216C>T, c.1705_1706delGA) were observed more than once (Table 3). MSH2 c.1216C>T was seen five times (at three different institutions) with all families reporting Mexico as their country of origin. MLH1 c.1852_1854delAAG was seen four times, with all families reportedly from El Salvador. Though seen in two unrelated families at one institution in our series, to our knowledge, MLH1 c.1790_1791del2ins9 has not been previously reported in the literature or MMR mutation databases, nor have 17 other mutations identified in our cohort (Appendix). Discussion
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This multi-institutional collaboration is the first to describe the spectrum of mutations in Hispanic LS patients in North America. MLH1 c.350C>T and MSH2 c.1216C>T have been reported multiple times in European studies (including Spain), as well as in US, South American, and Chinese studies [18, 21, 23, 39-42]. These two mutations are within CpG dinucleotides, which are known to cause mutational hotspots, and may explain why they were seen previously in multiple ethnicities as well as our cohort [43]. MLH1 c.1852_1854delAAG has been reported in France, South America, the Netherlands and others [8,15,24,44,45]. MLH1 c.350C>T,[21] c.2041G>A [42], c.332C>T [46], and c.676C>T [47] and MSH2 c.1216C>T [20, 21, 41, 48], deletion of exons 4-8 [20, 49], and deletion of exon 8 [21], and PMS2 deletion of exon 14 have all been reported previously in Spain [50]. The MSH2 deletion of exons 4-8 was seen in one Mexican family in our series and has been reported as a Spanish founder mutation [20]. The fact that several mutations in our cohort were previously reported in Spain is consistent with established migration patterns of Europeans, especially those from Spain, settling in Mexico and Central America. Obtaining a better understanding of penetrance and expression of MMR gene mutations in Hispanic families is a natural extension of this collaborative project. Many of the individuals included in this cohort were ascertained from colorectal cancer clinics and/or registries, such that other non-colon cancer, LS associated tumors may be underrepresented. A more expanded cohort with wider representation of individuals with other LS tumors is of interest.
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While all cases in this clinical series had a test performed to assess for LS (either tumor studies and/or germline genetic testing), both types of analyses were not available for all individuals. In addition, due to the retrospective nature of this study not all individuals in the cohort underwent complete germline analysis of the MMR genes because of the variability in institutional protocols for tumor testing, insurance coverage or accessibility, and availability of germline testing for specific genes over time (e.g. PMS2). Therefore, there may be individuals in this cohort with germline mutations that have not been identified. A limitation of this study is that clinical information, such as multigenerational family history and tumor histological features, was incomplete for two sites, so it is not reported. Finally, as this is a clinical study cohort, mutation prevalence cannot be estimated. Conclusion Findings from this collaborative study add to the understanding of recurrent MMR mutations in Hispanic populations. The presence of mutations in this cohort that were previously reported in Spanish and European populations reflects the ancestral connections of much of Latin America with Spain and Europe. However, the number of unique mutations, which have not been reported previously, suggest a rich area of further research to determine if there are potential Amerindian founder mutations. While this is the largest reported cohort of Hispanic patients with MMR mutations in North America, a larger sample and haplotype analyses are needed to better define the spectrum and origin of mutations in Hispanic populations.
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Acknowledgements: Research reported in this publication was supported by the National Cancer Institute of the National Institutes of Health under award number P30CA33572. Patients were recruited for study from the COH Clinical Cancer Genomics Community Research Network (CCGCRN), supported in part by award number RC4CA153828 (PI: J. Weitzel) from the National Cancer Institute and the Office of the Director, National Institutes of Health. The Hispanic MMR project is supported in part by a grant from The Safeway Foundation. USC research activities are supported by the Norris Cancer Center Core Grant (NCI P30CA014089) and the Anton B. Burg Foundation. The Puerto Rico Familiar Colorectal Cancer Registry was funded in part by a pilot NCI grant under award number R03 CA130034. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Cancer Institute or the National Institutes of Health. We would like to acknowledge the efforts of Lacolle Robinson and Kai Yang for data collection and Lily R Van Tongeren and Christina Ryback for manuscript preparation and review.
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[25] Alvarez K, Hurtado C, Hevia MA, Wielandt AM, de la Fuente M, Church J, Carvallo P, Lopez-Kostner F. Spectrum of MLH1 and MSH2 mutations in Chilean families with suspected Lynch syndrome. Dis Colon Rectum 2010;53:450-9. [26] da Silva FC, de Oliveira LP, Santos EM, Nakagawa WT, Aguiar Junior S, Valentin MD, Rossi BM, de Oliveira Ferreira F. Frequency of extracolonic tumors in Brazilian families with Lynch syndrome: analysis of a hereditary colorectal cancer institutional registry. Fam Cancer 2010;9:563-70. [27] Giraldo A, Gomez A, Salguero G, Garcia H, Aristizabal F, Gutierrez O, Angel LA, Padron J, Martinez C, Martinez H, Malaver O, Florez L, Barvo R. MLH1 and MSH2 mutations in Colombian families with hereditary nonpolyposis colorectal cancer (Lynch syndrome)-description of four novel mutations. Fam Cancer 2005;4:285-90. [28] Cruz-Correa M, Diaz-Algorri Y, Perez-Mayoral J, Suleiman-Suleiman W, Del Mar Gonzalez-Pons M, Bertran C, Casellas N, Rodriguez N, Pardo S, Rivera K, Mosquera R, Rodriguez-Quilichini S. Clinical characterization and mutation spectrum in Caribbean Hispanic families with Lynch syndrome. Fam Cancer 2015;14:415-25. [29] Weitzel JN, Clague J, Martir-Negron A, Ogaz R, Herzog J, Ricker C, Jungbluth C, Cina C, Duncan P, Unzeitig G, Saldivar JS, Beattie M, Feldman N, Sand S, Port D, Barragan DI, John EM, Neuhausen SL, Larson GP. Prevalence and type of BRCA mutations in Hispanics undergoing genetic cancer risk assessment in the southwestern United States: a report from the Clinical Cancer Genetics Community Research Network. J Clin Oncol 2013;31:210-6. [30] Provenzale D, Gupta S, Ahnen DJ, Bray T, Cannon JA, Cooper G, David DS, Early DS, Erwin D, Ford JM, Giardiello FM, Grady W, Halverson AL, Hamilton SR, Hampel H, Ismail MK, Klapman JB, Larson DW, Lazenby AJ, Lynch PM, Mayer RJ, Ness RM, Regenbogen SE, Samadder NJ, Shike M, Steinbach G, Weinberg D, Dwyer M, Darlow S. Genetic/Familial HighRisk Assessment: Colorectal Version 1.2016, NCCN Clinical Practice Guidelines in Oncology. J Natl Compr Canc Netw 2016;14:1010-30. [31] Vasen HF, Mecklin JP, Khan PM, Lynch HT. The International Collaborative Group on Hereditary Non-Polyposis Colorectal Cancer (ICG-HNPCC). Dis Colon Rectum 1991;34:424-5. [32] Vasen HFA, Watson P, Mecklin J-P, Lynch HT, (ICG-HNPCC). New clinical criteria for hereditary nonpolyposis colorectal cancer (HNPCC, Lynch Syndrome) proposed by the International Collaborative Group on HNPCC. Gastroenterology 1999;116:1453-6. [33] Rodriguez-Bigas MA, Boland CR, Hamilton SR, Henson DE, Jass JR, Khan PM, Lynch H, Perucho M, Smyrk T, Sobin L, Srivastava S. A National Cancer Institute Workshop on Hereditary Nonpolyposis Colorectal Cancer Syndrome: meeting highlights and Bethesda guidelines. J Natl Cancer Inst 1997;89:1758-62. [34] Umar A, Boland CR, Terdiman JP, Syngal S, de la Chapelle A, Ruschoff J, Fishel R, Lindor NM, Burgart LJ, Hamelin R, Hamilton SR, Hiatt RA, Jass J, Lindblom A, Lynch HT, Peltomaki P, Ramsey SD, Rodriguez-Bigas MA, Vasen HF, Hawk ET, Barrett JC, Freedman AN, Srivastava S. Revised Bethesda Guidelines for hereditary nonpolyposis colorectal cancer (Lynch Syndrome) and microsatellite instability. J Natl Cancer Inst 2004;96:261-8. [35] Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, Grody WW, Hegde M, Lyon E, Spector E, Voelkerding K, Rehm HL. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet Med 2015;17:40524. [36] den Dunnen JT, Dalgleish R, Maglott DR, Hart RK, Greenblatt MS, McGowan-Jordan J, Roux AF, Smith T, Antonarakis SE, Taschner PE. HGVS Recommendations for the Description of Sequence Variants: 2016 Update. Human mutation 2016;37:564-9. [37] International Society for Gastrointestinal Hereditary Tumours. Colon cancer gene variant databases. InSiGHT Variants databases: Leiden University Medical Center, 2008. 14
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[38] Woods MO, Williams P, Careen A, Edwards L, Bartlett S, McLaughlin JR, Younghusband HB. A new variant database for mismatch repair genes associated with Lynch syndrome. Hum Mutat 2007;28:669-73. [39] Sarroca C, Valle AD, Fresco R, Renkonen E, Peltomaki P, Lynch H. Frequency of hereditary non-polyposis colorectal cancer among Uruguayan patients with colorectal cancer. Clin Genet 2005;68:80-7. [40] Auclair J, Busine MP, Navarro C, Ruano E, Montmain G, Desseigne F, Saurin JC, Lasset C, Bonadona V, Giraud S, Puisieux A, Wang Q. Systematic mRNA analysis for the effect of MLH1 and MSH2 missense and silent mutations on aberrant splicing. Hum Mutat 2006;27:145-54. [41] Grindedal EM, Blanco I, Stormorken A, Maehle L, Clark N, Gonzalez S, Capella G, Vasen H, Burn J, Moller P. High risk of endometrial cancer in colorectal cancer kindred is pathognomonic for MMR-mutation carriers. Fam Cancer 2009;8:145-51. [42] Rodriguez-Soler M, Perez-Carbonell L, Guarinos C, Zapater P, Castillejo A, Barbera VM, Juarez M, Bessa X, Xicola RM, Clofent J, Bujanda L, Balaguer F, Rene JM, de-Castro L, MarinGabriel JC, Lanas A, Cubiella J, Nicolas-Perez D, Brea-Fernandez A, Castellvi-Bel S, Alenda C, Ruiz-Ponte C, Carracedo A, Castells A, Andreu M, Llor X, Soto JL, Paya A, Jover R. Risk of cancer in cases of suspected lynch syndrome without germline mutation. Gastroenterology 2013;144:926-32 e1; quiz e13-4. [43] Maliaka YK, Chudina AP, Belev NF, Alday P, Bochkov NP, Buerstedde JM. CpG dinucleotides in the hMSH2 and hMLH1 genes are hotspots for HNPCC mutations. Hum Genet 1996;97:251-5. [44] Syngal S, Fox EA, Li C, Dovidio M, Eng C, Kolodner RD, Garber JE. Interpretation of genetic test results for hereditary nonpolyposis colorectal cancer: Implications for clinical predisposition testing. JAMA-J Am Med Assoc 1999;282:247-53. [45] Wagner A, Tops C, Wijnen JT, Zwinderman K, van der Meer C, Kets M, Niermeijer MF, Klijn JG, Tibben A, Vasen HF, Meijers-Heijboer H. Genetic testing in hereditary non-polyposis colorectal cancer families with a MSH2, MLH1, or MSH6 mutation. J Med Genet 2002;39:833-7. [46] Caldes T, Godino J, de la Hoya M, Garcia Carbonero I, Perez Segura P, Eng C, Benito M, Diaz-Rubio E. Prevalence of germline mutations of MLH1 and MSH2 in hereditary nonpolyposis colorectal cancer families from Spain. Int J Cancer 2002;98:774-9. [47] Marcos I, Borrego S, Urioste M, Garcia-Valles C, Antinolo G. Mutations in the DNA mismatch repair gene MLH1 associated with early-onset colon cancer. J Pediatr 2006;148:8379. [48] Martinez-Bouzas C, Beristain E, Ojembarrena E, Errasti J, Mujika K, Viguera N, Tejada MI. A study on MSH2 and MLH1 mutations in hereditary nonpolyposis colorectal cancer families from the Basque Country, describing four new germline mutations. Fam Cancer 2009;8:533-9. [49] Perez-Cabornero L, Sanz MI, Sampedro EV, Aras EL, Becares AA, Pino CM, Dominguez MD. Frequency of rearrangements in Lynch syndrome cases associated with MSH2: characterization of a new deletion involving both EPCAM and the 5' part of MSH2. Cancer Prev Res (Phila) 2011;4:1556-62. [50] Brea-Fernandez AJ, Cameselle-Teijeiro JM, Alenda C, Fernandez-Rozadilla C, Cubiella J, Clofent J, Rene JM, Anido U, Mila M, Balaguer F, Castells A, Castellvi-Bel S, Jover R, Carracedo A, Ruiz-Ponte C. High incidence of large deletions in the PMS2 gene in Spanish Lynch syndrome families. Clin Genet 2014;85:583-8.
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Figure 1: Flowsheet of Tumor Studies and Germline Genetic Testing Performed in Cohort
Individuals Evaluated for Lynch Syndromea N=397
Family History / NO Personal History of Cancer N=16
Personal History of Cancer N=381
IHC and/or MSI on Tumor N=344
No Tumor Studies N=37
MMR Proficient N=243
NO Genetic Testing N=193
Genetic Testing N=37
b
VUS N=1 Negative N=18
Positive N=18
c
Genetic Testing N=50
VUS N=2 Negative N=47
Positive N=1
MMR Deficient N=101
NO Genetic Testing N=28
VUS N=4 Negative N=27
Genetic Testing N=73
Positive d N=42
Genetic Testing N=16
VUS N=0 Negative N=6
Positive N=10
16
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a Each
individual is only represented once = Variant of uncertain significance c Positive = Likely pathogenic and pathogenic alterations d Two individuals with a positive result also had a VUS, but are categorized in this figure based on the positive result b VUS
TABLE 1. Demographics of Study Cohort
Variable
Sequence negative or Only tumor studies [n (%)] 326 (82.4)
MMR gene mutation carrier [n (%)] 71 (17.9)
[n (%)]
42.98 (10.8)
43.39 (9.3)
43.1
≤ 40
102 (31.3)
28 (39.4)
130 (32.7)
>40
224 (68.7)
43 (60.6)
267 (67.3)
Female
162 (49.7)
40 (56.3)
202 (50.9)
Male
106 (32.5)
31 (43.7)
137 (34.5)
COH
70 (21.5)
19 (26.8)
89 (22.4)
MDA
92 (28.2)
14 (19.7)
106 (26.7)
USC
106 (32.5)
31 (43.7)
137 (34.5)
PR
58 (17.8)
7 (9.9)
65 (16.4)
Affected
315 (96.6)
61 (85.9)
381 (96.0)
Unaffected
6 (1.8)
10 (14.1)
16 (4.0)
Colorectal Uterine Ovarian Gastric Breast Other tumorc
249 (76.4) 31 (9.5) 8 (2.5) 8 (2.5) 6 (1.8) 13 (4.0)
47 (66.2) 9 (12.7) 0 1 (1.4) 0 4 (5.6)
296 (74.6) 40 (10.1) 8 (2.0) 9 (2.3) 6 (1.5) 22 (5.5)
1 2 ≥3
278 (85.3) 31 (9.5) 5 (1.5)
41 (57.7) 13 (18.3) 7 (9.9)
319 (80.4) 44 (11.1) 12 (3.0)
Category
N (%) Mean age (SD) Age
Gendera
Institution
Disease Status
First Cancer Diagnosis in Probandsb
Total
397
Number of Lynch syndrome Associated Tumorsd
17
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Country of Origin
a
Mexico
84 (25.8)
40 (56.3)
124 (31.2)
Puerto Rico El Salvador Guatemala Othere
59 (18.1) 13 (4.0) 14 (4.3) 156 (47.9)
5 (7.0) 6 (8.5) 6 (8.5) 14 (19.7)
64 (16.1) 19 (4.8) 20 (5.0) 170 (42.8)
Gender was missing for 58 individuals Individuals with multiple primaries were classified based on their first cancer diagnosis c Includes bladder cancer, glioblastoma multiforme, Hodgkin's lymphoma, kidney cancer, sarcoma, sebaceous carcinoma, small bowel cancer, ureter cancer, and unknown. d Non-LS associated tumors were excluded from the multiple primary cancers e Includes individuals from the Dominican Republic, Ecuador, Peru, Colombia, Spain, Honduras, Cuba, Chile, Nicaragua and those with partial or unspecified Hispanic ancestry b
TABLE 2. Characteristics of MMR Gene Mutations Affected
Unaffected
Total
61
10
71
Pathogenic
54
10
64
Likely pathogenic
7
0
7
MLH1 MSH2 MSH6 PMS2 Yes No
31 22 7 1 22 39
6 1 0 3 1 9
37 23 7 4 23 48
Probands
Classification
Gene
Recurrent
18
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TABLE 3: Recurrent and Previously Published Pathogenic/Likely Pathogenic Mutations Gene
Mutation
Frequency
Institution
Reported Country of Origin in Study Cohort
MLH1
c.350C>T
2
COH
Mexico
c.588+5G>A
2
MDA, USC
Mexico
c.676C>T
1*
USC
Mexico
c.1024_1039del16 c.1790_1791del2ins9
2 2
PR MDA
Puerto Rico Mexico
c.1852_1854delAAG
4
MDA, USC
El Salvador
c.2041G>A
2
COH, USC
c.425C>G
2
COH, USC
Exon 4-8 del
1*
COH
Mexico & Guatemala Mexico & Guatemala Mexico
c.1216C>T
5
COH, MDA, USC
Mexico
c.1705_1706delGA
2
MDA, PR
Exon 8 del Exon 14 del
1* 1*
MDA COH
MSH2
PMS2
Mexico & Puerto Rico El Salvador Mexico
Reported Country of Origin in Previous Publications Spain, South America, Europe, Japan, China Europe Spain, South America, Europe, USA Italy None South America, Europe, Japan, US South America, Spain, Chile, Columbia, Europe, Japan None Spain Spain, Uruguay, South America, Europe, China, US Europe Spain, Europe, USA, Canada Spain
Italicized highlights Spain and South American countries. * Not recurrent in this study, but previously cited in publications linked to South America and/or Spain. Abbreviations: COH, City of Hope; USC, University of Southern California; PR, Puerto Rico; MDA, MD Anderson
1
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