Genetic counselling legislation and practice in cancer in EU Member States.

BACKGROUND: Somatic and germline genetic alterations are significant drivers of cancer. Increasing integration of new technologies which profile these alterations requires timely, equitable and high-quality genetic counselling to facilitate accurate diagnoses and informed decision-making by patients and their families in preventive and clinical settings. This article aims to provide an overview of genetic counselling legislation and practice across European Union (EU) Member States to serve as a foundation for future European recommendations and action. METHODS: National legislative databases of all 27 Member States were searched using terms relevant to genetic counselling, translated as appropriate. Interviews with relevant experts from each Member State were conducted to validate legislative search results and provide detailed insights into genetic counselling practice in each country. RESULTS: Genetic counselling is included in national legislative documents of 22 of 27 Member States, with substantial variation in legal mechanisms and prescribed details (i.e. the 'who, what, when and where' of counselling). Practice is similarly varied. Workforce capacity (25 of 27 Member States) and genetic literacy (all Member States) were common reported barriers. Recognition and/or better integration of genetic counsellors and updated legislation and were most commonly noted as the 'most important change' which would improve practice. CONCLUSIONS: This review highlights substantial variability in genetic counselling across EU Member States, as well as common barriers notwithstanding this variation. Future recommendations and action should focus on addressing literacy and capacity challenges through legislative, regulatory and/or strategic approaches at EU, national, regional and/or local levels.

The heterogeneous cancer phenotype of individuals with biallelic germline pathogenic variants in CHEK2.

PURPOSE: Females with biallelic CHEK2 germline pathogenic variants (gPVs) more often develop multiple breast cancers than individuals with monoallelic CHEK2 gPVs. This study is aimed at expanding the knowledge on the occurrence of other malignancies. METHODS: Exome sequencing of individuals who developed multiple primary malignancies identified 3 individuals with the CHEK2 (NM_007194.4) c.1100del p.(Thr367MetfsTer15) loss-of-function gPV in a biallelic state. We collected the phenotypes of an additional cohort of individuals with CHEK2 biallelic gPVs (n = 291). RESULTS: In total, 157 individuals (53.4%; 157/294 individuals) developed ≥1 (pre)malignancy. The most common (pre)malignancies next to breast cancer were colorectal- (n = 19), thyroid- (n = 19), and prostate (pre)malignancies (n = 12). Females with biallelic CHEK2 loss-of-function gPVs more frequently developed ≥2 (pre)malignancies and at an earlier age compared with females biallelic for the CHEK2 c.470T>C p.(Ile157Thr) missense variant. Furthermore, 26 males (31%; 26/84 males) with CHEK2 biallelic gPVs developed ≥1 (pre)malignancies of 15 origins. CONCLUSION: Our study suggests that CHEK2 biallelic gPVs likely increase the susceptibility to develop multiple malignancies in various tissues, both in females and males. However, it is possible that a substantial proportion of individuals with CHEK2 biallelic gPVs is missed as diagnostic testing for CHEK2 often is limited to individuals who developed breast cancer.

Lessons learned from rapid exome sequencing for 575 critically ill patients across the broad spectrum of rare disease.

Introduction: Rapid exome sequencing (rES) has become the first-choice genetic test for critically ill patients, mostly neonates, young infants, or fetuses in prenatal care, in time-sensitive situations and when it is expected that the genetic test result may guide clinical decision making. The implementation of rES has revolutionized medicine by enabling timely identification of genetic causes for various rare diseases. The utilization of rES has increasingly been recognized as an essential diagnostic tool for the identification of complex and undiagnosed genetic disorders. Methods: We conducted a retrospective evaluation of our experiences with rES performed on 575 critically ill patients from various age groups (prenatal to adulthood), over a four-year period (2016-2019). These patients presented with a wide spectrum of rare diseases, including but not limited to neurological disorders, severe combined immune deficiency, and cancer. Results: During the study period, there was a significant increase in rES referrals, with a rise from a total of two referrals in Q1-2016 to 10 referrals per week in Q4-2019. The median turnaround time (TAT) decreased from 17 to 11 days in the period 2016-2019, with an overall median TAT of 11 days (IQR 8-15 days). The overall diagnostic yield for this cohort was 30.4%, and did not significantly differ between the different age groups (e.g. adults 22.2% vs children 31.0%; p-value 0.35). However, variability in yield was observed between clinical entities: craniofacial anomalies yielded 58.3%, while for three clinical entities (severe combined immune deficiency, aneurysm, and hypogonadotropic hypogonadism) no diagnoses were obtained. Discussion: Importantly, whereas clinical significance is often only attributed to a conclusive diagnosis, we also observed impact on clinical decision-making for individuals in whom no genetic diagnosis was established. Hence, our experience shows that rES has an important role for patients of all ages and across the broad spectrum of rare diseases to impact clinical outcomes.

Rapid whole exome sequencing in pregnancies to identify the underlying genetic cause in fetuses with congenital anomalies detected by ultrasound imaging.

OBJECTIVE: The purpose of this study was to explore the diagnostic yield and clinical utility of trio-based rapid whole exome sequencing (rWES) in pregnancies of fetuses with a wide range of congenital anomalies detected by ultrasound imaging. METHODS: In this observational study, we analyzed the first 54 cases referred to our laboratory for prenatal rWES to support clinical decision making, after the sonographic detection of fetal congenital anomalies. The most common identified congenital anomalies were skeletal dysplasia (n = 20), multiple major fetal congenital anomalies (n = 17) and intracerebral structural anomalies (n = 7). RESULTS: A conclusive diagnosis was identified in 18 of the 54 cases (33%). Pathogenic variants were detected most often in fetuses with skeletal dysplasia (n = 11) followed by fetuses with multiple major fetal congenital anomalies (n = 4) and intracerebral structural anomalies (n = 3). A survey, completed by the physicians for 37 of 54 cases, indicated that the rWES results impacted clinical decision making in 68% of cases. CONCLUSIONS: These results suggest that rWES improves prenatal diagnosis of fetuses with congenital anomalies, and has an important impact on prenatal and peripartum parental and clinical decision making.

Serum AMH levels in healthy women from BRCA1/2 mutated families: are they reduced?

STUDY QUESTION: Do BRCA1/2 mutation carriers have a compromised ovarian reserve compared to proven non-carriers, based on serum anti-Müllerian hormone (AMH) levels? SUMMARY ANSWER: BRCA1/2 mutation carriers do not show a lower serum AMH level in comparison to proven non-carriers, after adjustment for potential confounders. WHAT IS KNOWN ALREADY: It has been suggested that the BRCA genes play a role in the process of ovarian reserve depletion, although previous studies have shown inconsistent results regarding the association between serum AMH levels and BRCA mutation status. Hence, it is yet unclear whether BRCA1/2 mutation carriers may indeed be at risk of a reduced reproductive lifespan. STUDY DESIGN, SIZE, DURATION: A multicenter, cross-sectional study was performed between January 2012 and February 2015 in 255 women. We needed to include 120 BRCA1/2 mutation carriers and 120 proven non-carriers to demonstrate a difference in AMH levels of 0.40 µg/l (SD ± 0.12 µg/l, two-sided alpha-error 0.05, power 80%). PARTICIPANTS/MATERIALS, SETTING, METHOD: Healthy women aged 18-45 years who were referred to the Clinical Genetics Department and applied for predictive BRCA1/2 testing because of a familial BRCA1/2 mutation were asked to participate. A cross-sectional assessment was performed by measuring serum AMH levels and filling out a questionnaire. Multivariate linear regression analyses adjusted for age, current smoking and current hormonal contraceptive use were performed on log-transformed serum AMH levels. MAIN RESULTS AND THE ROLE OF CHANCE: Out of 823 potentially eligible women, 421 (51.2%) were willing to participate, and of those, 166 (39%) did not meet our inclusion criteria. Two hundred and fifty-five women were available for analyses; 124 BRCA1/2 mutation carriers and 131 proven non-carriers. The median [range] AMH level in carriers was 1.90 µg/l [0.11-19.00] compared to 1.80 µg/l [0.11-10.00] in non-carriers (P = 0.34). Adjusted linear regression analysis revealed no reduction in AMH level in the carriers (relative change = 0.98 (95%CI, 0.77-1.22); P = 0.76). LIMITATIONS, REASONS FOR CAUTION: Participants were relatively young. Power was insufficient to analyze BRCA1 and BRCA2 mutation carriers separately. AMH levels may have been influenced by the use of hormonal contraceptives, though similar proportions of carriers and non-carriers were current users and adjustments were made to correct for potential confounding in our analysis. WIDER IMPLICATIONS OF THE FINDINGS: Limitations of the current analysis and limitations of the existing literature argue for prospective, well-controlled follow-up studies with recurrent AMH measurements to determine whether carriers might be at risk for low ovarian reserve and to definitively guide care. STUDY FUNDING/COMPETING INTERESTS: This study was partially financially supported by a personal grant for Inge A.P. Derks-Smeets, kindly provided by the Dutch Cancer Society (Grant Number UM 2011-5249). Theodora C. van Tilborg, Inge A.P. Derks-Smeets, Anna M.E. Bos, Jan C. Oosterwijk, Christine E. de Die-Smulders, Lizet E. van der Kolk, Wendy A.G. van Zelst-Stams, Maria E. Velthuizen, Marinus J.C. Eijkemans and Margreet G.E.M. Ausems have nothing to disclose. Ron J. van Golde has received unrestricted research grants from Ferring and Merck Serono, outside the submitted work. Annemieke Hoek received an unrestricted educational grant from Ferring pharmaceutical BV, The Netherlands and a speaker's fee for post graduate education from MSD pharmaceutical company, outside the submitted work. Joop S.E. Laven has received unrestricted research grants from Ferring, Merck Serono, Merck Sharpe & Dome, Organon, and Schering Plough, outside the submitted work. Frank J.M. Broekmans is a member of the external advisory board for Merck Serono (The Netherlands), outside the submitted work. TRIAL REGISTRATION NUMBER: NTR no. 4324.

High Satisfaction and Low Distress in Breast Cancer Patients One Year after BRCA-Mutation Testing without Prior Face-to-Face Genetic Counseling.

According to standard practice following referral to clinical genetics, most high risk breast cancer (BC) patients in many countries receive face-to-face genetic counseling prior to BRCA-mutation testing (DNA-intake). We evaluated a novel format by prospective study: replacing the intake consultation with telephone, written and digital information sent home. Face-to-face counseling then followed BRCA-mutation testing (DNA-direct). One year after BRCA-result disclosure, 108 participants returned long-term follow-up questionnaires, of whom 59 (55 %) had previously chosen DNA-direct (intervention) versus DNA-intake (standard practice i.e., control: 45 %). Questionnaires assessed satisfaction and psychological distress. All participants were satisfied and 85 % of DNA-direct participants would choose this procedure again; 10 % would prefer DNA-intake and 5 % were undecided. In repeated measurements ANOVA, general distress (GHQ-12, p = 0.01) and BC-specific distress (IES-bc, p = 0.03) were lower in DNA-direct than DNA-intake at all time measurements. Heredity-specific distress (IES-her) did not differ significantly between groups. Multivariate regression analyses showed that choice of procedure did not significantly contribute to either general or heredity-specific distress. BC-specific distress (after BC diagnosis) did contribute to both general and heredity-specific distress. This suggests that higher distress scores reflected BC experience, rather than the type of genetic diagnostic procedure. In conclusion, the large majority of BC patients that used DNA-direct reported high satisfaction without increased distress both in the short term, and 1 year after conclusion of genetic testing.

[CHEK2-mutation in Dutch breast cancer families: expanding genetic testing for breast cancer]. / CHEK2-mutatie in Nederlandse borstkankerfamilies: uitbreiding van de genetische diagnostiek op borstkanker.

In the majority of breast cancer families, DNA testing does not show BRCA1 or BRCA2 mutations and the genetic cause of breast cancer remains unexplained. Routine testing for the CHEK2*1100delC mutation has recently been introduced in breast cancer families in the Netherlands. The 1100delC mutation in the CHEK2-gene may explain the occurrence of breast cancer in about 5% of non-BRCA1/2 families in the Netherlands. In the general population the CHEK2*1100delC mutation confers a slightly increased breast cancer risk, but in a familial breast cancer setting this risk is between 35-55% for first degree female carriers. Female breast cancer patients with the CHEK2*1100delC mutation are at increased risk of contralateral breast cancer and may have a less favourable prognosis. Female heterozygous CHEK2*1100delC mutation carriers are offered annual mammography and specialist breast surveillance between the ages of 35-60 years. Prospective research in CHEK2-positive families is essential in order to develop more specific treatment and screening strategies.

A germline homozygous mutation in the base-excision repair gene NTHL1 causes adenomatous polyposis and colorectal cancer.

The genetic cause underlying the development of multiple colonic adenomas, the premalignant precursors of colorectal cancer (CRC), frequently remains unresolved in patients with adenomatous polyposis. Here we applied whole-exome sequencing to 51 individuals with multiple colonic adenomas from 48 families. In seven affected individuals from three unrelated families, we identified a homozygous germline nonsense mutation in the base-excision repair (BER) gene NTHL1. This mutation was exclusively found in a heterozygous state in controls (minor allele frequency of 0.0036; n = 2,329). All three families showed recessive inheritance of the adenomatous polyposis phenotype and progression to CRC in at least one member. All three affected women developed an endometrial malignancy or premalignancy. Genetic analysis of three carcinomas and five adenomas from different affected individuals showed a non-hypermutated profile enriched for cytosine-to-thymine transitions. We conclude that a homozygous loss-of-function germline mutation in the NTHL1 gene predisposes to a new subtype of BER-associated adenomatous polyposis and CRC.

NRAS-mutated melanocytic BAP1-associated intradermal tumor (MBAIT): a case report.

Melanocytic BAP1-associated intradermal tumors (MBAITs) are epithelioid spitzoid looking, mostly intradermally located melanocytic tumors that often have tumor-infiltrating lymphocytes and a common nevus component. They occur sporadically but also in the context of an underlying BAP1 germline mutation. Recognition of these lesions is important because they can be a marker for an underlying BAP1-associated cancer syndrome. Most cases reported in the literature thus far were found to have both a BRAF and BAP1 mutation. Here, we report an unusual case of an MBAIT lesion with a combined NRAS and BAP1 mutation. A BAP1 germline mutation was excluded. Our case is the second case reported until now with this combination of mutations in this subset of lesions. In the other reported NRAS-/BAP1-mutated MBAIT case, presence of a BAP1 germline mutation was not tested. Our case confirms that the mutational spectrum in MBAITs is broader than previously thought. Just as in the BRAF-mutated cases, it is likely that a subset might be associated with a BAP1 germline mutation. In case of suspicion of an MBAIT lesion based on histological examination, diagnostic work-up should include assessment of protein expression and/or mutation analysis of at least BRAF, NRAS, and BAP1. Work-up should not be limited to analyzing only BRAF protein expression or mutation, since NRAS-mutated MBAITs might be missed.

Somatic mutations in MLH1 and MSH2 are a frequent cause of mismatch-repair deficiency in Lynch syndrome-like tumors.

Lynch syndrome is caused by germline mutations in the mismatch repair (MMR) genes. Tumors are characterized by microsatellite instability (MSI). However, a considerable number of MSI-positive tumors have no known molecular mechanism of development. By using Sanger and ion semiconductor sequencing, 25 MSI-positive tumors were screened for somatic mutations and loss of heterozygosity in mutL homolog 1 (MLH1) and mutS homolog 2 (MSH2). In 13 of 25 tumors (8 MLH1-deficient and 5 MSH2-deficient tumors), we identified 2 somatic mutations in these genes. We conclude that 2 acquired events explain the MMR-deficiency in more than 50% of the MMR-deficient tumors without causal germline mutations or promoter methylation.

More breast cancer patients prefer BRCA-mutation testing without prior face-to-face genetic counseling.

Currently, most breast cancer (BC) patients receive face-to-face genetic counseling (DNA-intake) prior to BRCA-mutation testing, with generic information regarding hereditary BC and BRCA-mutation testing. This prospective study evaluated a novel format: replacing the intake consultation with telephone, written and digital information sent home, and face-to-face contact following BRCA-mutation testing (DNA-direct). From August 2011 to February 2012, 161 of 233 eligible BC patients referred to our Human Genetics department chose between DNA-direct (intervention) or DNA-intake (control). Exclusion criteria were psychological problems (n = 33), difficulty with Dutch text (n = 5), known BRCA-family (n = 3), non-BRCA-referral (n = 1). 30 declined genetic counseling or study participation. Participants received questionnaires including satisfaction and psychological distress. 59 % chose DNA-direct (p = 0.03), of whom 90 % were satisfied and would choose DNA-direct again (including 6/8 BRCA-mutation carriers); although 27 % hesitated to recommend DNA-direct to other patients. General distress (GHQ-12, p = 0.001) and heredity-specific distress (IES, p = 0.02) scored lower in DNA-direct than DNA-intake, both at baseline and follow-up 2 weeks after BRCA-result disclosure; all scores remained below clinical relevance. DNA-direct participants reported higher website use (53 vs. 32 %, p = 0.01), more referrer information about personal consequences (41 vs. 20 %, p = 0.004) and lower decisional conflict (median 20 [0-88] vs. 25 [0-50], p = 0.01). Processing time in DNA-direct was reduced by 1 month. Mutation detection rate was 8 % in both groups. All BRCA-mutation carriers fulfilled current testing criteria. In conclusion, more BC patients preferred DNA-direct over intake consultation prior to BRCA-mutation testing, the majority being strongly to moderately satisfied with the procedure followed, without increased distress.

A post-hoc comparison of the utility of sanger sequencing and exome sequencing for the diagnosis of heterogeneous diseases.

The advent of massive parallel sequencing is rapidly changing the strategies employed for the genetic diagnosis and research of rare diseases that involve a large number of genes. So far it is not clear whether these approaches perform significantly better than conventional single gene testing as requested by clinicians. The current yield of this traditional diagnostic approach depends on a complex of factors that include gene-specific phenotype traits, and the relative frequency of the involvement of specific genes. To gauge the impact of the paradigm shift that is occurring in molecular diagnostics, we assessed traditional Sanger-based sequencing (in 2011) and exome sequencing followed by targeted bioinformatics analysis (in 2012) for five different conditions that are highly heterogeneous, and for which our center provides molecular diagnosis. We find that exome sequencing has a much higher diagnostic yield than Sanger sequencing for deafness, blindness, mitochondrial disease, and movement disorders. For microsatellite-stable colorectal cancer, this was low under both strategies. Even if all genes that could have been ordered by physicians had been tested, the larger number of genes captured by the exome would still have led to a clearly superior diagnostic yield at a fraction of the cost.

Focusing on patient needs and preferences may improve genetic counseling for colorectal cancer.

During cancer genetic counseling, different items which counselors consider important are discussed. However, relatively little empirical evidence exists regarding the needs and preferences of counselees. In this study needs and preferences were assessed from counselees with a personal and/or family history of colorectal cancer (CRC), who were referred for genetic counseling regarding CRC. They received a slightly modified version of the QUOTE-GENE(ca) questionnaire prior to their first visit to the Hereditary Cancer Clinic. Response rate was 60 % (48/80 participants). Counselees rated the importance of 45 items assessing their needs and preferences regarding the content and process of genetic counseling. Participants rated the items regarding discussion of information about their familial CRC risk (100 %) and preventive options (98 %) as important or very important. Fewer participants rated items concerning general information on genetics as important. Sensitive communication during counseling was considered very important by a large percentage of counselees. Generally, no major differences were seen between participants in relation to individual characteristics. Our data suggest that focusing on familial CRC risk and surveillance options, in combination with sensitive communication may lead to better satisfaction with genetic counseling.

DNA-testing for BRCA1/2 prior to genetic counselling in patients with breast cancer: design of an intervention study, DNA-direct.

BACKGROUND: Current practice for patients with breast cancer referred for genetic counseling, includes face-to-face consultations with a genetic counselor prior to and following DNA-testing. This is based on guidelines regarding Huntington's disease in anticipation of high psychosocial impact of DNA-testing for mutations in BRCA1/2 genes. The initial consultation covers generic information regarding hereditary breast cancer and the (im)possibilities of DNA-testing, prior to such testing. Patients with breast cancer may see this information as irrelevant or unnecessary because individual genetic advice depends on DNA-test results. Also, verbal information is not always remembered well by patients. A different format for this information prior to DNA-testing is possible: replacing initial face-to-face genetic counseling (DNA-intake procedure) by telephone, written and digital information sent to patients' homes (DNA-direct procedure). METHODS/DESIGN: In this intervention study, 150 patients with breast cancer referred to the department of Clinical Genetics of the Radboud University Nijmegen Medical Centre are given the choice between two procedures, DNA-direct (intervention group) or DNA-intake (usual care, control group). During a triage telephone call, patients are excluded if they have problems with Dutch text, family communication, or of psychological or psychiatric nature. Primary outcome measures are satisfaction and psychological distress. Secondary outcome measures are determinants for the participant's choice of procedure, waiting and processing times, and family characteristics. Data are collected by self-report questionnaires at baseline and following completion of genetic counseling. A minority of participants will receive an invitation for a 30 min semi-structured telephone interview, e.g. confirmed carriers of a BRCA1/2 mutation, and those who report problems with the procedure. DISCUSSION: This study compares current practice of an intake consultation (DNA-intake) to a home informational package of telephone, written and digital information (DNA-direct) prior to DNA-testing in patients with breast cancer. The aim is to determine whether DNA-direct is an acceptable procedure for BRCA1/2 testing, in order to provide customized care to patients with breast cancer, cutting down on the period of uncertainty during this diagnostic process.

Quantification of sequence exchange events between PMS2 and PMS2CL provides a basis for improved mutation scanning of Lynch syndrome patients.

Heterozygous mutations in PMS2 are involved in Lynch syndrome, whereas biallelic mutations are found in Constitutional mismatch repair-deficiency syndrome patients. Mutation detection is complicated by the occurrence of sequence exchange events between the duplicated regions of PMS2 and PMS2CL. We investigated the frequency of such events with a nonspecific polymerase chain reaction (PCR) strategy, co-amplifying both PMS2 and PMS2CL sequences. This allowed us to score ratios between gene and pseudogene-specific nucleotides at 29 PSV sites from exon 11 to the end of the gene. We found sequence transfer at all investigated PSVs from intron 12 to the 3' end of the gene in 4 to 52% of DNA samples. Overall, sequence exchange between PMS2 and PMS2CL was observed in 69% (83/120) of individuals. We demonstrate that mutation scanning with PMS2-specific PCR primers and MLPA probes, designed on PSVs, in the 3' duplicated region is unreliable, and present an RNA-based mutation detection strategy to improve reliability. Using this strategy, we found 19 different putative pathogenic PMS2 mutations. Four of these (21%) are lying in the region with frequent sequence transfer and are missed or called incorrectly as homozygous with several PSV-based mutation detection methods.