Lars Fredrik Engebretsen

CHEK2 mutations affecting kinase activity together with mutations in TP53 indicate a functional pathway associated with resistance to epirubicin in primary breast cancer

Background Chemoresistance is the main obstacle to cure in most malignant diseases. Anthracyclines are among the main drugs used for breast cancer therapy and in many other malignant conditions. Single parameter analysis or global gene expression profiles have failed to identify mechanisms causing in vivo resistance to anthracyclines. While we previously found TP53 mutations in the L2/L3 domains to be associated with drug resistance, some tumors harboring wild-type TP53 were also therapy resistant. The aim of this study was; 1) To explore alterations in the TP53 gene with respect to resistance to a regular dose epirubicin regimen (90 mg/m2 every 3 week) in patients with primary, locally advanced breast cancer; 2) Identify critical mechanisms activating p53 in response to DNA damage in breast cancer; 3) Evaluate in vitro function of Chk2 and p14 proteins corresponding to identified mutations in the CHEK2 and p14(ARF) genes; and 4) Explore potential CHEK2 or p14(ARF) germline mutations with respect to family cancer incidence. Methods and Findings Snap-frozen biopsies from 109 patients collected prior to epirubicin (as preoperative therapy were investigated for TP53, CHEK2 and p14(ARF) mutations by sequencing the coding region and p14(ARF) promoter methylations. TP53 mutastions were associated with chemoresistance, defined as progressive disease on therapy (p = 0.0358; p = 0.0136 for mutations affecting p53 loop domains L2/L3). Germline CHEK2 mutations (n = 3) were associated with therapy resistance (p = 0.0226). Combined, mutations affecting either CHEK2 or TP53 strongly predicted therapy resistance (p = 0.0101; TP53 mutations restricted to the L2/L3 domains: p = 0.0032). Two patients progressing on therapy harbored the CHEK2 mutation, Arg95Ter, completely abrogating Chk2 protein dimerization and kinase activity. One patient (Epi132) revealed family cancer occurrence resembling families harboring CHEK2 mutations in general, the other patient (epi203) was non-conclusive. No mutation or promoter hypermethylation in p14(ARF) were detected. Conclusion This study is the first reporting an association between CHEK2 mutations and therapy resistance in human cancers and to document mutations in two genes acting direct up/down-stream to each other to cause therapy failure, emphasizing the need to investigate functional cascades in future studies.

Current clinical criteria for Lynch syndrome are not sensitive enough to identify MSH6 mutation carriers

Background Reported prevalence, penetrance and expression of deleterious mutations in the mismatch repair (MMR) genes, MLH1, MSH2, MSH6 and PMS2, may reflect differences in the clinical criteria used to select families for DNA testing. The authors have previously reported that clinical criteria are not sensitive enough to identify MMR mutation carriers among incident colorectal cancer cases. Objective To describe the sensitivity of the criteria when applied to families with a demonstrated MMR mutation. Methods Families with an aggregation of colorectal cancers were examined for deleterious MMR mutations according to the Mallorca guidelines. All families with a detected MMR mutation as of November 2009 were reclassified according to the Amsterdam and Bethesda criteria. Results Sixty-nine different DNA variants were identified in a total of 129 families. The original Amsterdam clinical criteria were met by 38%, 12%, 78% and 25% of families with mutations in MSH2, MSH6, MLH1 and PMS2, respectively. Corresponding numbers for the revised Amsterdam criteria were 62%, 48%, 87% and 38%. Similarly, each of the four clinical Bethesda criteria had low sensitivity for identifying MSH6 or PMS2 mutations. Conclusion Amsterdam criteria and each of the Bethesda criteria were inadequate for identifying MSH6 mutation-carrying kindreds. MSH6 mutations may be more common than currently assumed, and the penetrance/expression of MSH6 mutations, as derived from families meeting current clinical criteria, may be misleading. To increase detection rate of MMR mutation carriers, all cancers in the Lynch syndrome tumour spectrum should be subjected to immunohistochemical analysis and/or analysis for microsatellite instability.

Psychosocial Factors Associated with Quality of Life Among Individuals Attending Genetic Counseling for Hereditary Cancer

The aim of this multicenter study was to explore associations between psychosocial factors (general self-efficacy, perceived availability of social support, cancer-related distress) and health-related quality of life, among individuals at risk for hereditary cancer. One-hundred and twenty one participants with a family history of breast-cancer or colorectal cancer answered a questionnaire 2–4 weeks prior to genetic counseling. The two dimensions of the health-related quality of life measure, mental and physical health were both used as outcome variables. Multiple regression (linear) analyses revealed that increasing degrees of cancer-related distress was related to decreasing degrees of mental health whereas increasing degrees of self-efficacy and social support were related to increasing degrees of this outcome variable. Self-efficacy, self-esteem support and tangible aid seemed to moderate the relationship between cancer-related distress and mental health. These results suggest that self-efficacy and certain resources of social support buffer the negative association between cancer-related distress and mental health, and might be suitable for interventional efforts. Implications for genetic counseling practice are discussed.

A homozygote splice site PMS2 mutation as cause of Turcot syndrome gives rise to two different abnormal transcripts

Turcot syndrome is a rare, inherited disease predisposing of tumours in the central nerve system and in the colorectal system. This report describes a Turcot patient with an extraordinary clinical history. The patient is still alive at the age of 43. She was operated at the age of 10 by brain tumour and at the age of 16 by colorectal cancer. She has since then been treated for multiple cancers (gastrointestinal, endometrial, basal cell carcinomas), and removal of adenomatous polyps at several occasions. The aim of this work was to investigate if there was any specific genotype that explains her remarkable clinical history. Microsatellite instability and immunohistochemistry analysis for four DNA mismatch repair proteins were performed. DNA mutation analysis was done for genes involved in polyposis and mismatch repair by denaturing high performance liquid chromatography and sequencing. cDNA analysis was carried out for the mismatch repair gene PMS2. The patients genotype was found to be a homozygous splice site mutation in the PMS2 gene, c.989-1G<T, which resulted in two abnormal transcripts, not one as expected. The patient’s long time survival may in part be explained by meticulous follow up by health care professionals. The other importing factor is probably the nature of here genotype. cDNA analysis showed that the homozygous mutation led to two abnormal transcripts, of which one is perhaps less detrimental. Thus cDNA analysis is of prime importance for the full evaluation of the effect of putative splicing mutations.

Registration rate in the Norwegian Cruciate Ligament Register: Large-volume hospitals perform better

Background and purpose The Norwegian Cruciate Ligament Register (NCLR) was founded in 2004. The purpose of the NCLR is to provide representative and reliable data for future research. In this study we evaluated the development of the registration rate in the NCLR. Methods The Norwegian Patient Register (NPR) and the electronic patient charts (EPCs) were used as reference data for public and private hospitals, respectively. Data were retrieved for all primary and revision anterior cruciate ligament (ACL) surgery during 2008–2009 in public hospitals and during 2008 in private hospitals. The NOMESCO classification of surgical procedures was used for identification of ACL surgeries. Public hospitals were divided into subgroups according to the annual number of operations in the NPR: small hospitals (< 30 operations) and large hospitals (≥ 30 operations). Results For the 2-year data extracted from public hospitals, 2,781 and 2,393 operations met the inclusion criteria according to the NPR and the NCLR, respectively, giving an average registration rate of 86% (95% CI: 0.85–0.87). The registration rate for small public hospitals was 69% (CI: 0.65–0.73), which was significantly less than for large public hospitals (89%, CI: 0.88–0.90; p < 0.001). In 2008, private hospitals reported 548 operations to the NCLR while 637 were found in the EPCs, giving a registration rate of 86% (CI: 0.83–0.89). In that year, the registration rate for public hospitals was 86%, which was similar to that for private hospitals. Interpretation The NCLR registration rate for the period 2008–09 was similar in both 2008 and 2009, and is satisfactory for research. There is room for improvement of registration rates, particularly in hospitals with a small volume of ACL operations.

Prospective findings in breast cancer kindreds: annual incidence rates according to age, stage at diagnosis, mean sojourn time, and incidence rates for contralateral cancer

Abstract 2102 women over 55 years of age from breast cancer kindreds as clinically defined were examined with annual clinical examination and mammography every second year. Initially, all family histories were obtained and all women were invited for genetic counselling. During screening, 37 infiltrating cancers (CA) and 9 carcinomas in situ (CIS) were found. Annual incidence rates for CA or CIS were calculated to be 0.82% for those aged 30 years or more. Annual incidence rate for contralateral cancer among those who had contracted one cancer was 6%. In the first round, 17 of 23 cancers (74%) were without spread, and this increased to 20 of 23 (87%) during follow-up. Two of 23 cancers (18%) were CIS at the first round, increasing to 7 of 23 (30%) at follow-up visits. CIS was diagnosed at a younger age than CA. Employing figures for mean sojourn time from sporadic cancer to calculate the annual incidence rate underlying the observation in the first round gave the same incidence as observed during actual follow-up. This indicates that inherited breast cancer progression is comparable with that of sporadic cancer.

Stigmatization and Male Identity: Norwegian Males’ Experience after Identification as BRCA1/2 Mutation Carriers

In families with hereditary breast and ovarian cancer, there is limited knowledge about the reactions of BRCA1/2 mutation positive males. In the present qualitative study, fifteen BRCA1/2 mutation positive men in Norway participated in two successive, in-depth interviews. Seven female partners participated in the second interview. The men reported strong emotional reactions to their positive test results, and they expressed a desire to keep the genetic information private. They considered discussing their test results or health related information with other males as difficult, and they perceived females as their sources of social and emotional support. Interestingly, the second interview revealed important information not communicated during the first interview. The findings of this study contribute to the discussion of whether men who test positive for a BRCA1/2 mutation should receive tailored genetic counseling sessions. Health care providers should be aware of psychological vulnerability in these men, likely stemming from fewer emotional supports in their social networks.

High penetrances of BRCA1 and BRCA2 mutations confirmed in a prospective series

Penetrances of BRCA1 and BRCA2 mutations have been derived from retrospective studies, implying the possibility of ascertainment biases to influence the results.We have followed women at risk for breast and/or ovarian cancer for two decades, and report the prospectively observed age-related annual incidence rates to contract breast or ovarian cancer for women with deleterious BRCA1 or BRCA2 mutations based on 4830 observation years. Patients were grouped according to mutation, age and having/not having had previous cancer.In women not having had previous cancer and aged 40-59 years, the annual incidence rate to contract breast or ovarian cancer in those having the most frequent BRCA1 founder mutations was 4.0%, for women in this age group and with less frequent BRCA1 mutations annual incidence rate was 5.9%, and for women with BRCA2 mutations 3.5%.The observed figures may be used for genetic counseling of healthy mutation carriers in the respective age groups. The results may indicate that less frequent BRCA1 mutations have higher penetrances than BRCA1 founder mutations.

Living with Multiple Endocrine Neoplasia Type 1: Decent Care-Insufficient Medical and Genetic Information A Qualitative Study of MEN 1 Patients in a Swedish Hospital

This qualitative study explores how 29 Swedish patients with Multiple Endocrine Neoplasia type 1 (MEN1) experience living with the condition, appraisal of the clinical follow-up program, and surveys their future expectations. The aim of this study is to build knowledge about this patient group in order to provide optimal care. The participants describe physical, psychological, and social limitations in their daily activities and how these limitations influence quality of life. Our findings indicate that a majority of patients have adjusted to their situation, describing themselves as being healthy despite physical symptoms and treatment. The participants received decent care in the clinical follow-up program, - however, greater effort should be put into patient information. These patients might benefit from genetic counseling. Health professionals involved should recognize their potential impact and influence on a patient’s ability to adjust to these circumstances. Antonovsky`s Sense of Coherence theory is used to discuss these findings.

A Novel Deep Intronic Mutation Introducing a Cryptic Exon Causing Neurofibromatosis Type 1 in a Family with Highly Variable Phenotypes: A Case Study

Neurofibromatosis type 1 (NF1) is a common dominant inherited disorder with highly variable expressivity. Genetic testing for this condition has been more available the last decade. Here we present a case report of a NF1 family including seven affected family members, some of them with a very severe phenotype. When searching for a causative mutation in the NF1 gene, no mutation was found at DNA level. However, a misspliced transcript including a subsequence of intron 3 appeared when screening RNA. The underlying cause at DNA level was determined to be a deep intronic variant (c.288+1137C>T). This intronic point mutation creates a new splice site causing the insertion of a cryptic exon (r.288_289ins288+1018_1135), leading to reading frameshift at the protein level. Deep intronic mutations introducing a cryptic exon are known to be a cause of NF1, and we reviewed the literature to evaluate how common this mutation is in NF1 syndrome. We found 20 different deep intronic NF1 splice mutations, including the one found in the present study. In conclusion, this case illustrates the value of RNA analysis to detect the cause of genetic diseases, and we decided to use RNA based mutation screening as standard procedure for NF1 genetic testing in our laboratory.