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To determine the prevalence and type of BRCA1 and BRCA2 (BRCA) mutations among Hispanics in the Southwestern United States and their potential impact on genetic cancer risk assessment (GCRA).
Hispanics (n = 746) with a personal or family history of breast and/or ovarian cancer were enrolled in an institutional review board–approved registry and received GCRA and BRCA testing within a consortium of 14 clinics. Population-based Hispanic breast cancer cases (n = 492) enrolled in the Northern California Breast Cancer Family Registry, negative by sequencing for BRCA mutations, were analyzed for the presence of the BRCA1 ex9-12del large rearrangement.
Deleterious BRCA mutations were detected in 189 (25%) of 746 familial clinic patients (124 BRCA1, 65 BRCA2); 21 (11%) of 189 were large rearrangement mutations, of which 62% (13 of 21) were BRCA1 ex9-12del. Nine recurrent mutations accounted for 53% of the total. Among these, BRCA1 ex9-12del seems to be a Mexican founder mutation and represents 10% to 12% of all BRCA1 mutations in clinic- and population-based cohorts in the United States.
BRCA mutations were prevalent in the largest study of Hispanic breast and/or ovarian cancer families in the United States to date, and a significant proportion were large rearrangement mutations. The high frequency of large rearrangement mutations warrants screening in every case. We document the first Mexican founder mutation (BRCA1 ex9-12del), which, along with other recurrent mutations, suggests the potential for a cost-effective panel approach to ancestry-informed GCRA.
Hispanics, the fastest growing group in the United States, comprise 15.1% of the population. (Although we use the term “Hispanic” in this article, the more common census term for individuals of Spanish, Mexican, and Central and South American descent, referring to “ethnicity,” is “Latino.” Latino is generally considered a more ethnically/culturally based term for individuals of the aforementioned groups.) Breast cancer (BC) is the most commonly diagnosed cancer in Hispanic women and leading cause of cancer death. Although the incidence of BC in Hispanics is less than in non-Hispanic whites, our initial studies on the prevalence of deleterious mutations in BRCA1 and BRCA2 (BRCA) suggested they may account for a higher proportion of BC in Hispanics than other non–Ashkenazi Jewish populations.1,2 We and others have documented that BRCA1 185delAG is a recurrent mutation in Hispanics,1,3,4 occurring on the Jewish haplotype.1,5 Deleterious large rearrangement BRCA mutations are not detectable by standard sequencing.6–8 The prevalence of BRCA1 ex9-12del, a recurrent large rearrangement mutation initially identified in a small Mexican-American high-risk clinic cohort, is unknown.2
The risk for BRCA mutation carriers to develop BC varies from 57% by age 70 years9 to 85% lifetime risk among high-risk clinic patients, with lower risks reported from population-based studies.10 They also have a 20% to 50% risk for ovarian cancer (OC).11 The availability of effective screening, treatment, and risk reduction interventions makes BRCA testing a standard of care for patients with a personal and/or family history suggestive of an inherited predisposition to breast and/or ovarian cancer.12–16 However, low-income, underinsured, and racial/ethnic minority individuals have a significant burden of cancer and have limited access to genetic cancer risk assessment (GCRA). In addition, there is a dearth of Hispanic-specific research, particularly in the area of genetic predisposition to BC.
The Hispanic population in the Southwestern United States is primarily of Mexican ancestry, whereas individuals of Puerto Rican, Dominican, and Cuban ancestry predominate in the Eastern United States. Admixture studies indicate significantly different ancestral populations among US Hispanics.17,18 Because of the design of their test requisition form, the exclusive BRCA testing vendor in the United States cannot distinguish between Hispanics of Caribbean ancestry and those of Mexican and/or Central American ancestry.19 Therefore, we assembled two large cohorts of US Hispanics with a focus on the latter groups—a clinic-based cohort of patients referred for GCRA in the Southwestern United States and a cohort selected from a cancer registry population-based study—to determine the prevalence and type of BRCA mutations and explore the potential to translate the findings into ancestry-informed strategies for cost-effective GCRA.
The City of Hope Clinical Cancer Genetics Community Research Network includes a cross-section of cancer center and community-based clinics, primarily in the Southwestern United States, that provide GCRA to individuals with a personal or family history of cancer.20 All GCRA patients are invited to participate in an institutional review board–approved prospective Hereditary Cancer Registry at the time of consultation (> 90% participation). Between May 1998 and June 2010, 746 probands with self-reported Hispanic origin, mostly from Mexico and Central America (Table 1), were seen for GCRA, enrolled in the registry, and underwent clinical BRCA testing after informed consent. Only one individual from each family was included in the analyses. Participants with mixed ancestry were eligible only if pedigree analysis indicated that the Hispanic lineage was the likely origin of the familial cancer pattern. Blood samples, demographic data, and five-generation pedigrees were obtained, including reported ethnicity and country/state of origin for each grandparental lineage. Clinical details were obtained for relatives affected with BC and/or OC. A bilingual cancer risk counselor or translator conducted GCRA sessions for Spanish-speaking patients, with adapted counseling aides and consent forms.21,22
Tested solely for BRCA1 ex9-12del were DNA samples from 492 patients with BC of Mexican ancestry, negative for sequence-detected BRCA mutations, age less than 65 years with a family history of cancer, identified through the population-based Greater San Francisco Bay Area Cancer Registry and enrolled in the Northern California Breast Cancer Family Registry (NC-BCFR).3,23
Genetic testing was offered to women in the clinic-based cohort who met National Comprehensive Cancer Network criteria.12 BRCA testing was performed at Myriad Genetic Laboratories (Salt Lake City, UT) and included full sequencing of exons and flanking intronic segments,24 five specific BRCA1 rearrangements for testing after August 12, 2002, and multiplex quantitative differential polymerase chain reaction (PCR; BRCA Analysis Rearrangement Testing [BART])2 after August 1, 2006, for large rearrangement mutation testing for cases that met the vendor's automatic criteria (~BRCA mutation probability ≥ 30%). Because of the frequency of the BRCA1 ex9-12del mutation, as a triage step, a separate PCR analysis was performed for all cases in the clinic-based cohort that did not receive automatic BART. It was cost effective to test for that mutation specifically on a research basis (less than $5 per sample) and then obtain a “single site” rate for clinical grade testing from Myriad for the known mutation. For all remaining cases not meeting the vendor's criteria, BART was conducted electively when covered by private insurance or patient payment; BRCA1 was screened in the remaining uninformative cases by multiplex ligation-dependent probe amplification assay (MRC-Holland, Amsterdam, the Netherlands).25
To screen for the BRCA1 ex9-12del large rearrangement, a three-primer PCR assay was used.2 It resulted in coamplification of the mutant allele 742-bp breakpoint fusion product and a 1,145-bp wild-type allele product. As indicated in Figure 1, all BRCA-negative clinic-based cases and NC-BCFR3 population-based samples were tested.
Probabilities of carrying a BRCA mutation were estimated using the Myriad Tables (February 2010), BOADICEA (v2), and BRCAPRO (v2.0-5) models.24,26–28 Pedigrees were created electronically using Progeny 8 (Progeny Software, Delray Beach, FL) and uploaded to the BOADICEA Web Application29 and to Hughes riskApps30 for BRCAPRO probabilities.
DNA samples from 18 BRCA1 R1443X carriers and 20 BRCA1 ex9-12del carriers were genotyped at 12 microsatellite markers spanning 4.1 Mb of chromosome 17q encompassing the BRCA locus. When possible, haplotypes associated with each mutation were inferred by determining phase from related individuals within each kindred with the same mutation. Primer sequence design and PCR amplifications were previously described,1,2,31,32 with additional microsatellite markers (D17S649, D17S1787, D17S1801, D17S750, D17S951, D17S1860, D17S1861) from the University of California, Santa Cruz, genome database.33 Mutation age estimation was performed using the statistical model used in Neuhausen et al.31
Among the 746 clinic-based probands, there were 590 with BC, 39 with OC, 20 with both BC and OC, and 97 unaffected (Table 1). The average age at first BC diagnosis was 40 years. The majority of probands reported Mexico as their grandparents' country of origin (n = 582). Central America (n = 80), South America (n = 36), the Caribbean (n = 13), and Spain (n = 35) were also reported.
Overall, 189 (25%) had deleterious mutations (124 in BRCA1, 65 in BRCA2) in the clinic-based cohort; of these, 21 (11%) were large rearrangements (13 BRCA1 ex9-12del, seven unique rearrangements in BRCA1, and one in BRCA2). Fewer than half of the large rearrangement mutation carriers in the present study met Myriad Genetic Laboratories criteria (~30% prior probability) for automatic large rearrangement testing. Thirty-four (5%) had one or more unclassified variants, and 523 (70%) had negative/uninformative results (Table 1).
For 745 cases with complete pedigree data, the mean probability of a mutation across the clinic-based cohort, a cross-section of cancer center and community-based clinics, was calculated at 18.7% by BOADICEA, 12.8% by BRCAPRO, and 9.2% by Myriad.
In Table 2, nine recurrent BRCA mutations (seen in four or more unrelated families) along with grandparental country of origin (Mexican state specified when known) are shown. This subset accounted for 53% of all detected BRCA mutations. Eighteen had a BRCA1 185delAG mutation (15% of BRCA1 mutation carriers) and 13 had an ex9-12del mutation (10% of BRCA1 carriers). This subset also included R71G (n = 9), a Spanish founder mutation.34 The six unrelated Hispanic BRCA1 R1443X mutation carriers shared four distinct haplotypes: two independent haplotypes of Mexican ancestry, one of Columbian, and one of Peruvian ancestry. These were distinct from the haplotype seen in French-Canadian samples (samples courtesy of Dr. W. Foulkes).35,36 The BRCA2 3492insT mutation (n = 10) accounted for 15% of the BRCA2 mutations. Two recurrent BRCA1 mutations, 917delTT (n = 5) and IVS5+1 G more than A (n = 4), were observed exclusively in probands with family origins in El Salvador and Guatemala; they were reported previously in Italy37 and Spain,38 respectively. Probands with the BRCA2 9254del5 mutation (n = 5), reported previously in Spain,39 were exclusively of El Salvadoran origin.
In addition, the BRCA1 ex9-12del mutation was detected in three of 492 BRCA sequence negative families of Mexican ancestry identified through the population-based Greater San Francisco Bay Area Cancer Registry and enrolled in the NC-BCFR; this represents 12% (three of 25) of the BRCA1 mutations in the cohort (22 BRCA1 mutations were previously reported in the overall cohort).3
BRCA1ex9-12del mutation carriers (n = 13) were genotyped, and mutational age analyses estimated the BRCA1 ex9-12del mutation to have arisen 74 generations, or 1,480 years ago (95% CI, 920 to 2,260 years).
To date, this is the largest study of Hispanic breast/ovarian cancer families in the United States, confirming a high prevalence of BRCA mutations (25%), as well as a pattern of multiple recurrent mutations in this mostly Mexican-American population. Large rearrangement mutations, not detectable on standard sequencing, represented a significant proportion of the carriers. Nine recurrent mutations accounted for 53% of the total, suggesting the potential for more cost-effective, ancestry-informed genetic screening. Currently, the sensitivity of a Hispanic-specific BRCA panel is being evaluated prospectively.
As highlighted in Figure 2, the spectrum of mutations in Hispanic cohorts is similar in Texas, New Mexico, Arizona, and California,3,40 and the relative proportions of specific recurrent mutations such as BRCA1 185delAG and ex9-12del are the same as those in the population-based series of patients with BC enrolled in the NC-BCFR,3 suggesting that the pattern is generalizable and not due to referral bias. The persistence of village life and low rates of relocation among the Mexican population may account in part for persistent ancestral patterns of recurrent mutations.41 Although the ancestry-driven pattern is evident in the immigrant Mexican-American population, acculturation and further admixture with majority populations likely would ultimately diffuse the predictive value of a panel approach to testing. We would suggest that the patterns we observed in the immigrant Mexican-American population may be a relatively unbiased representation of the Mexican population, wherein there is currently little access to GCRA and BRCA testing. This hypothesis should be tested prospectively in Mexico.
Although most of the recurrent mutations are likely Spanish in origin, the BRCA1 ex9-12del mutation has never been observed in Spain or South America.42,43 Representing 10% to 12% of BRCA1 mutations in clinic- and population-based cohorts, all ex9-12del carriers reported Mexican ancestry, and the mutation was estimated to have arisen 1,480 years ago, predating Spanish colonization. Thus BRCA1 ex9-12del is clinically significant and one of the most frequent population-specific large rearrangement mutations in the world, as well as the first reported Mexican founder mutation.
Commercial BRCA rearrangement testing using the multiplex quantitative differential polymerase chain reaction method became available in 2006. Less than half of the large rearrangement carriers in the present study met Myriad's criteria (~30% prior probability) for large rearrangement testing. Furthermore, recent data from Myriad indicated that BRCA large rearrangement mutations are frequent (21%) in patients of Latin American/Caribbean ancestry who were tested for BRCA mutations and that BRCA1 ex9-12del represented a significant proportion of these.44
Eighteen probands had a BRCA1 185delAG, a known Jewish founder mutation, in our Hispanic population (9.5% of BRCA carriers). The term Hispanos has been applied to the Colonial-Hispanic population45 in the San Luis Valley, encompassing parts of Colorado and New Mexico, and it is suggested that their ancestral origins stem from the immigration of Spanish Conversos and Crypto-Jews.46 Although five 185delAG carriers in our study were from New Mexico, and thus likely Hispanos, they reported grandparental ancestry as Mexican or Spanish. Given that the majority of 185delAG carriers in our cohorts were recruited from areas of the United States outside of known Colonial-Hispanic settlements, the high prevalence is clinically relevant and may represent a greater than appreciated diaspora of people with Converso and Crypto-Jewish ancestry. In other words, the BRCA1 185delAG mutation is of Jewish origin and prevalent across the Mexican-American Hispanic population.
Previously reported as a French-Canadian founder mutation with highly conserved haplotypes,35,36 BRCA1 R1443X was observed six times in our study population, with ancestry reported from Mexico (n = 4), Colombia (n = 1), and Peru (n = 1). Our demonstration of distinct haplotypes, none in common with the French-Canadian samples, supports the hypothesis that there are multiple independent origins, possibly due to hypermutability of the CG dinucleotide to TG.35
The BRCA mutation prevalence of 25% in our high-risk population was higher than expected compared with the output of all three mutation probability models (BOADICEA, BRCAPRO, Myriad) that were applied. Reports of the predictive accuracy of the BRCAPRO, BOADICEA, and Myriad BRCA mutation probability models in Hispanic populations are conflicting.15,40,47,48 Our data support the possibility that the underlying prevalence of BRCA mutations in the Mexican-American population may be higher than the reference populations used to validate the models. We also observed a higher prevalence than previous reports. One small clinic-based study of Hispanics reported a sequencing-detected BRCA mutation prevalence of 17.9%,40 and a prevalence of 14.8% was reported among 1,936 cases with reported Latin American/Caribbean ancestry who received commercial BRCA sequencing, including the five-site large rearrangement panel.19 The latter study was not able to segregate ancestry data according to Hispanic subsets (eg, Caribbean Islanders v Mexican or South American) because it was based on limited categorical information volunteered on a commercial test requisition form. Neither of these studies screened for other rearrangements such as BRCA1 ex9-12del, which accounted for 10% of all BRCA1 mutations in our cohort. With the exception of Ashkenazi-Jewish subjects, Hispanics had the highest rate of BRCA1 mutations (10.8%) among women younger than 65 years with BC and with a family history of cancer, selected from a population-based cancer registry (NC-BCFR).3 This was likely an underestimate given that only BRCA1 was screened,3 and the testing would have missed genomic rearrangements, some of which were captured in our studies of this cohort. Thus the mutation prevalence observed in our study may be the closest approximation of the mutation prevalence in Hispanics who meet the criteria for BRCA testing and suggests that BRCA mutations may account for a higher percentage of familial BC in those of Mexican descent than other ethnic groups. This observation would be strengthened by future studies of the prevalence of BRCA mutations in other ethnic groups, both within the Clinical Cancer Genetics Community Research Network consortium of cancer center and community-based clinics and in other clinic- and population-based studies. Once validated, it may be appropriate to consider adjusting the threshold for recommending BRCA testing among Mexican-American Hispanics, similar to the situation in the Ashkenazi Jewish population. In addition, given the relatively higher proportion of BRCA mutations, future BC epidemiology studies among Hispanics with Mexican ancestry may need to consider analysis and stratification by BRCA status.
GCRA is a medical standard-of-care option for high-risk families and may identify persons at increased risk for cancer before the onset of disease, when early detection or prevention strategies are most effective.12,14,15 For example, salpingo-oophorectomy substantially lowers BC risk and all-cause mortality in premenopausal BRCA1 carriers.13,49 We previously demonstrated that there is interest in genetics and cancer prevention among underserved Hispanic patients in Los Angeles50 and that high-risk Hispanic women in an indigent care setting will attend their clinic visits.21 Furthermore, culturally adapted GCRA protocols seem to be effective in promoting risk-appropriate follow-up behaviors.22,51 Consequently, population-specific GCRA protocols and ancestry-informed genetic testing may have significant potential to be cost effective by superior allocation of health care resources to prevention and early detection of cancer in high-risk individuals, especially in a population in which families tend to be larger.
In this study of Hispanic breast/ovarian cancer families in the United States, the largest to date, we report a high prevalence of BRCA mutations, many of which were recurrent, and a significant proportion of which were large rearrangements. Many of these women and their family members would potentially be left unaware of extraordinary risk, as half of those with large rearrangement mutations did not meet the commercial vendor's criteria for automatic inclusion of comprehensive large rearrangement screening. From our professional education programs,52 we are aware of a significant gap in clinicians' knowledge about large rearrangement mutations, ultimately resulting in inadequate patient care and potential liability. In addition to the possibility of a relatively higher prevalence of BRCA mutations in Mexican-Americans, incomplete family cancer history reporting can influence the performance of probability models. The reasons for lack of apparent family cancer history may be a combination of limited family structure53 and limited family knowledge. Although formal assessment of multigenerational pedigrees was employed in this study, the depth of information about the extended family was sometimes limited in part because of separation from their ancestors as an immigrant population or cultural influences regarding health communication, with implications for ancestry-informed genetic screening.
Regardless of the factors influencing the prevalence and type of BRCA mutations, our study affirms the need for access to BRCA testing for Hispanics, with inclusion of full large rearrangement screening (ie, BART) for all patients. The latter recommendation was included in the 2012 National Comprehensive Cancer Network guidelines, wherein BRCA gene analysis was defined as the combination of sequencing and large rearrangement analyses.54
We thank David E. Goldgar, PhD, Department of Dermatology, University of Utah, Salt Lake City, UT, for expert assistance with the estimation of mutation age; William Foulkes, MD, Departments of Medicine and Human Genetics, McGill University, Montreal, Quebec, Canada, for sharing reference DNAs from French-Canadian BRCA1 R1443X carriers for haplotype studies; and Jane Congleton, MS, RN, CGC, for recruitment of study patients at Banner Good Samaritan Medical Center. Alexander Miron was responsible for previously published BRCA1 heteroduplex analysis of the NC-BCFR samples. Some of the genotyping was performed by Myriad Genetics Laboratory (Salt Lake City, UT) under research contract No. MGA 138. We would also like to thank Tracy Sulkin for assistance with manuscript preparation. Samples from the NC-BCFR were processed and distributed by the Coriell Institute for Medical Research.
Clinical Cancer Genetics Community Research Network: City of Hope (Jeffrey N. Weitzel, MD, Jessica Clague, PhD, Deborah J. MacDonald, PhD, Kathleen R. Blazer, EdD, Julie O. Culver, MS, Carin Espenschied, MS, Deborah I. Barragan, MS), Banner Good Samaritan Medical Center (Shaun R. Opie, PhD, Chelsy Jungbluth, MS), Cancer Center of Santa Barbara (Frederic Kass, MD, Megan McKenna, MS), Doctors Hospital of Laredo (Gary Unzeitig, MD), Edwards Comprehensive Cancer Center (Lisa Muto, MSN), Hematology Oncology Associates of New Mexico (Paul Duncan, MD), Holy Cross Hospital–Michael and Dianne Bienes Cancer Center (Zdenka Segota, MD, Carol Brudenell, MSN), Hunterdon Regional Cancer Center (Brian Quinn, MD, Jacqueline Hale, RN), John H. Stroger, Jr. Hospital of Cook County (Pamela Ganschow, MD, Christina Seelaus, MS), NM Oncology and Hematology Consultants (Annette Fontaine, MD, Darling Horcasitas, PA), North Valley Breast Clinic (Ian Grady, MD, Lauren Strickland, DO), Olive View–UCLA Medical Center (Nancy Feldman, MD, Lori Viveros, MPH), Saddleback Memorial Medical Center (Michelle Fajardo, DO, Sandra Brown, MS), St. Alphonsus Regional Medical Center (Kerry Pulver, MD, Patricia Dock, MS), St. Joseph Hospital (David Margileth, MD, Kimberly Banks, MS, Cheryl Cina, MS), St Jude Medical Center (William Lawler, M.D, John Lee, MS), Sutter Roseville Medical Center (Kristie Bobolis, MD, Kim Van Ysseldyk, MSN), Texas Tech University (J. Salvador Saldivar, MD, Kayla Castaneda, MSN), University of Southern California Norris Comprehensive Cancer Center (Darcy Spicer, MD, Charité Ricker, MS).
Written on behalf of the Clinical Cancer Genetics Community Research Network. See the Appendix (online only) for a list of participants.
Supported in part by USC Norris Comprehensive Cancer Center, American Cancer Society Grant No. RSGT-00-263-01 and Grants No. 1R03CA139588 and 1RC4CA153828 from the National Cancer Institute (principal investigator [PI], J.N.W.). In addition, J.C. and A.M.-N. were supported by Grant No. R25CA085771 from the National Cancer Institute (PI, J.N.W.). The Northern California Breast Cancer Family Registry (NC-BCFR) was supported by the National Cancer Institute, National Institutes of Health, under Grant No. RFA-CA-06-503 and through cooperative agreements with members of the BCFR and PIs, including the Cancer Prevention Institute of California (grant No. U01CA69417). M.B. was funded by the Avon Foundation, the San Francisco Bay Area Breast Specialized Programs of Research Excellence (SPORE Grant No. 2P50CA058207), and the Center for Translational and Policy Research in Personalized Medicine (Grant No. P01CA130818-02A1).
The content of this article does not necessarily reflect the views or policies of the National Cancer Institute or any of the collaborating centers in the BCFR, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government or the BCFR.
Authors' disclosures of potential conflicts of interest and author contributions are found at the end of this article.
The author(s) indicated no potential conflicts of interest.
Conception and design: Jeffrey N. Weitzel, Jessica Clague, Josef Herzog, Sharon Sand, Garrett P. Larson
Financial support: Jeffrey N. Weitzel, Mary Beattie, Esther M. John
Administrative support: Jeffrey N. Weitzel, Arelis Martir-Negron, Josef Herzog, Charité Ricker, Chelsy Jungbluth, Cheryl Cina, Paul Duncan, Gary Unzeitig, J. Salvador Saldivar, Sharon Sand, Danielle Port, Garrett P. Larson
Provision of study materials or patients: Jeffrey N. Weitzel, Jessica Clague, Charité Ricker, Chelsy Jungbluth, Cheryl Cina, Paul Duncan, Gary Unzeitig, J. Salvador Saldivar, Mary Beattie, Nancy Feldman, Deborah I. Barragan, Esther M. John
Collection and assembly of data: Jeffrey N. Weitzel, Jessica Clague, Arelis Martir-Negron, Raquel Ogaz, Josef Herzog, Charité Ricker, Chelsy Jungbluth, Cheryl Cina, Paul Duncan, Gary Unzeitig, J. Salvador Saldivar, Mary Beattie, Nancy Feldman, Sharon Sand, Danielle Port, Deborah I. Barragan, Esther M. John, Garrett P. Larson
Data analysis and interpretation: Jeffrey N. Weitzel, Jessica Clague, Arelis Martir-Negron, Josef Herzog, Susan L. Neuhausen, Garrett P. Larson
Manuscript writing: All authors
Final approval of manuscript: All authors