Recurrent genetic variants and prioritization of variants of uncertain clinical significance associated with hereditary breast and ovarian cancer in families from the Region of Murcia

© 2023 the author(s). This manuscript version is made available under the CC-BY 4.0 license http://creativecommons.org/licenses/by/4.0/. This document is the Published version of a Published Work that appeared in final form in Advances in Laboratory Medicine. To access the final edited and published work see https://doi.org/10.1515/almed-2023-0103Objectives Hereditary breast and ovarian cancer (HBOC) follows an autosomal dominant inheritance pattern of cancer susceptibility genes. The risk of developing this disease is primarily associated with germline mutations in the BRCA1 and BRCA2 genes. The advent of massive genetic sequencing technologies has expanded the mutational spectrum of this hereditary syndrome, thereby increasing the number of variants of uncertain clinical significance (VUS) detected by genetic testing. Methods A prevalence study of HBOC was performed within 2,928 families from the Region of Murcia, in southeastern Spain. Genetic testing enabled the identification of recurrent pathogenic variants and founder mutations, which were mainly related to the BRCA1 and BRCA2 genes. VUS testing was performed using a prioritization algorithm designed by our working group. Results Variants c.68_69del, c.212+1G>A, and c.5123C>A were detected in 30 % of BRCA1 carriers, whereas exon 2 deletion concurrent with c.3264dupT, c.3455T>G and c.9117G>A variants were found in 30 % of BRCA2 carriers. A total of 16 VUS (15 %) were prioritized. Conclusions The genotype-phenotype correlation observed in our study is consistent with the scientific literature. Furthermore, the founder effect of c.1918C>T (BRCA1) and c.8251_8254del (ATM) was verified in the Murcian population, whereas exon 2 deletion (BRCA2) was proven to be a Spanish founder mutation. Our algorithm enabled us to prioritize potentially pathogenic VUS that required further testing to determine their clinical significance and potential role in HBOC

Poliposis adenomatosa familiar hiperatenuada

En este trabajo se realiza un análisis descriptivo clínico de 28 familias portadoras de una misma alteración en el gen APC que conlleva un fenotipo muy atenuado de poliposis familiar, asociado a un incremento moderado del riesgo de cáncer colorrectal. Desde la Unidad de Consejo Genético en Cáncer del Hospital Morales Meseguer de Murcia entre los años 2007 y 2014 detectamos 28 sujetos (casos índice) aparentemente no relacionadas, que compartían una misma mutación en el exón 2 de APC (APC: c.147_150delACAA). Todas las familias provenían de un área geográfica concreta dentro de la Región de Murcia: el Valle de Ricote y la Vega Media del Segura. Los objetivos principales de la tesis fueron: 1) demostrar el origen común o efecto fundador de esta mutación en el área. 2) caracterizar de forma detallada el fenotipo en los portadores, haciendo hincapié en el carácter especialmente atenuado del mismo en comparación con otras familias similares descritas en la literatura. 3) realizar una estimación de la datación de dicha mutación en la Región de Murcia. 4) proponer un protocolo de diagnóstico y seguimiento específico para los portadores de esta mutación. Metodología. Tras oportuno Consejo Genético se procedió a testar a los familiares de los casos índices hasta un total de 354 sujetos, detectando 194 portadores, los cuales fueron objeto de un análisis exhaustivo de sus características clínicas (historia clínica, datos de colonoscopias, casos de CCR, manifestaciones extracolónicas, etcétera). Para confirmar el origen común de la mutación identificada, se realizó análisis de ligamiento con marcadores genéticos próximos al gen APC. Para hacer una aproximación de la datación de la mutación se emplearon dos métodos diferentes, uno calculando el número de generaciones transcurridas desde el antecesor común más reciente (ACMR) de todos los alelos mutados analizados y otro empleando no un solo marcador sino el haplotipo completo. Resultados y Conclusiones. Se pudo comprobar el carácter fundacional de la mutación, así como realizar un cálculo aproximado de la aparición de dicha mutación en esta área geográfica concreta entre 875 y 950 años. Se determinaron las características fenotípicas que la presencia de dicha mutación confería a los portadores, principalmente en cuanto al número y tipo de pólipos colónicos y la presencia o no de cáncer colorrectal, con un hallazgo fundamental: el marcado carácter atenuado que esta alteración genética provocaba, dando lugar a un menor número de adenomas, con un menor riesgo y una edad más elevada de diagnóstico de cáncer colorrectal en los portadores, comparado con otras series de Poliposis Adenomatosa Familiar Atenuada (PAFA) publicadas. Por último planteamos una propuesta de protocolo de diagnóstico de nuevos casos y de seguimiento clínico/endoscópico en los pacientes con PAFA de estas áreas de salud. In this work we carried out a descriptive clinical analysis of 28 families carrying the same alteration in the APC gene, that leads to a very attenuated phenotype of familial polyposis, associated with a moderate increase in the risk of colorectal cáncer. From the Cancer Genetic Counseling Unit of Hospital Morales Meseguer in Murcia between 2007 and 2014, we detected 28 subjects (index cases) apparently unrelated, who shared the same mutation in exon 2 of APC (APC: c.147_150delACAA). All the families came from a specific geographical area within the Region of Murcia: Ricote Valley and the Vega Media del Segura. The main objectives of this work were: 1) to demonstrate the common origin or founding effect of this mutation in the area. 2) characterize in detail the phenotype in the carriers, emphasizing the especially attenuated nature of it, in comparison with other similar families described in the literature. 3) make an estimate of the dating of the mutation in the Region of Murcia. 4) propose a specific diagnosis and monitoring protocol for the carriers of this mutation. Methodology. After opportune Genetic Counsel we proceeded to test the relatives of the index cases up to a total of 354 subjects, detecting 194 carriers, who underwent an exhaustive analysis of their clinical characteristics (clinical history, colonoscopy data, cases of CRC, extracolonic manifestations, etc.). To confirm the common origin of the identified mutation, linkage analysis was performed with genetic markers next to the APC gene. To make an approximation of the mutation dating, two different methods were used, one calculating the number of generations passed from the most recent common ancestor (ACMR) of all the mutated alleles analyzed and another using not a single marker but the complete haplotype. Results and conclusions The foundational character of the mutation could be verified, as well as an approximate calculation of the appearance of the mutation in this specific geographical area between 875 and 950 years. We determined the phenotypic characteristics that the presence of this mutation conferred to the carriers, mainly regarding the number and type of colonic polyps and the presence or not of colorectal cancer, with a fundamental finding: the marked attenuated character that this genetic alteration caused, giving rise to a lower number of adenomas, with a lower risk and a higher age of diagnosis of colorectal cancer in the carriers, compared with other published series of Attenuated Familial Adenomatous Polyposis (AFAP) Finally, we propose a protocol for the diagnosis of new cases and clinical / endoscopic follow-up in patients with AFAP in this geographic área

Mutational analysis of RAD51C and RAD51D genes in hereditary breast and ovarian cancer families from Murcia (southeastern Spain)

© 2018 Elsevier Masson SAS. All rights reserved. This document is the Published version of a Published Work that appeared in final form in European Journal of Medical Genetics. To access the final edited and published work see https://doi.org/10.1016/j.ejmg.2018.01.015RAD51C and RAD51D have been defined as susceptibility genes for hereditary breast and ovarian cancer syndrome in several studies. In the present study, a mutation analysis of these genes was performed on non BRCA1/2 families. RAD51C and RAD51D genes were analyzed in 141 and 77 families, respectively. The analysis included direct sequencing and multiple ligation probe analysis. The RAD51C pathogenic variant c.404G > A was identified in a breast and ovarian cancer family (0.7%), while the RAD51D pathogenic variant c.694C > T was described in an ovarian cancer family (1.3%). Moreover, three unknown clinical significance variants were detected: c.307T > G in RAD51C, and c.413A > G and c.715C > T in RAD51D. No large genomic rearrangements (LGRs) were found. RAD51D carriers suffered from premenopausal ovarian tumors. These results increase our knowledge about the RAD51C and RAD51D mutation spectrum and support the notion that these genes should be included in the gene panel testing performed on patients with hereditary breast and ovarian cancer syndrome

Recurrent genetic variants and prioritization of variants of uncertain clinical significance associated with hereditary breast and ovarian cancer in families from the Region of Murcia

Hereditary breast and ovarian cancer (HBOC) follows an autosomal dominant inheritance pattern of cancer susceptibility genes. The risk of developing this disease is primarily associated with germline mutations in the BRCA1 and BRCA2 genes. The advent of massive genetic sequencing technologies has expanded the mutational spectrum of this hereditary syndrome, thereby increasing the number of variants of uncertain clinical significance (VUS) detected by genetic testing

Variantes genéticas recurrentes y priorización de variantes de significado clínico desconocido asociadas al síndrome de cáncer de mama y ovario hereditario en familias de la Región de Murcia

El síndrome de cáncer de mama y ovario hereditario (SCMOH) presenta un patrón de herencia autosómica dominante en genes de susceptibilidad al cáncer y su riesgo está principalmente vinculado a mutaciones germinales en BRCA1 y BRCA2. Sin embargo, la implementación de paneles genéticos mediante secuenciación masiva en la práctica asistencial, ha ampliado el espectro mutacional de este síndrome hereditario y el número de variantes de significado clínico desconocido (VUS) detectadas en los estudios genéticos

Next step in molecular genetics of hereditary breast/ovarian cancer: Multigene panel testing in clinical actionably genes and prioritization algorithms in the study of variants of uncertain significance

© 2022 Elsevier Masson SAS. All rights reserved. This document is the Published version of a Published Work that appeared in final form in European Journal of Medical Genetics. To access the final edited and published work see https://doi.org/10.1016/j.ejmg.2022.104468Introduction: BRCA1 and BRCA2 are the two main genes causing hereditary breast and ovarian cancer (HBOC). However, thanks to the development of Next Generation Sequencing (NGS), other genes linked to this syndrome (CHEK2, BRIP1, ATM and PALB2 among others) can be analysed. Material and methods: an analysis by multigene panel testing was performed in 138 index cases (ICs) from HBOC Spanish families with a previous non-informative result for BRCA1/2. The BRCA Hereditary Cancer Master™ Plus kit, including 26 actionable and candidate genes related to HBOC was employed. Once classified, an algorithm was employed to prioritized those variants of unknown significance with a higher risk of having a deleterious effect. Moreover, a mRNA splicing assay was performed for the prioritized VUS c.3402+3A > C in ATM, located at intron 23. Results: A total of 82 variants were found: 70 VUS and 12 pathogenic or probably pathogenic variants. The diagnostic yield in actionable genes non-BRCA was 7.97% of the total tested ICs. Overall, 19 VUS were prioritized, which meant 27% of the 70 total VUS. RNA analysis of the variant 3402+3A > C confirmed a deleterious impact on splicing. Discussion: The implementation of a multigene panel in HBOC studied families improved the diagnostic yield, concordant with results obtained in previous publications. Due to the important number of VUS obtained in NGS, the application of a prioritization algorithm is needed in order to select those variants in which it is necessary to conduct further studies

Molecular characterization and clinical interpretation of BRCA1/BRCA2 variants in families from Murcia (south-eastern Spain) with hereditary breast and ovarian cancer: clinical–pathological features in BRCA carriers and non-carriers

© Springer Science+Business Media Dordrecht 2017. This document is the Published version of a Published Work that appeared in final form in Familial Cancer. To access the final edited and published work see https://doi.org/10.1007/s10689-017-9985-xThis is the first study performed in Murcia (south-eastern Spain) in which 592 families with hereditary breast and ovarian cancer were identified thanks to Genetic Counselling Units from this area over 6 years. Diagnostic performance was 18.1% and 194 different genetic variants were obtained. Variants with uncertain significance accounted for only 5.6% of the total number of reports, so our population has been well characterised. In BRCA1 gene, two novel variants were found (c.1859delT and c.3205C > T) and the most frequently detected mutations were c.68_69delAG, c.212 + 1G > A, c.5123C > A, c.211A > G and c.1918C > T, which together represented 56.67% of total pathogenic mutations. In BRCA2 gene, four recurrent variants were described (deletion of entire exon 2, c.9117G > A, c.3264dupT and c.3455T > G) representing 43.5% of the mutations in this gene. Mutation c.68_69delAG and deletion of entire exon 2 in BRCA1 and BRCA2 genes respectively were the most prevalent variants in our population. Regarding the genotype-phenotype relation, mutation c.212 + 1G > A appeared in an important percentage of breast and ovarian cancer cases, c.5123C > A in bilateral breast cancer and c.9117G > A in bilateral breast cancer and ovarian cancer. With respect to clinical–pathological characteristic, BRCA1/BRCA2 mutation carriers showed earlier onset age of breast tumour and higher risk of developing contra lateral breast cancer than non-informative cases. Moreover, association between either molecular subtype triple negative breast cancer or ovarian cancer and BRCA1 carriers was obtained