Comprehensive Mutation Analysis of PMS2 in a Large Cohort of Probands Suspected of Lynch Syndrome or Constitutional Mismatch Repair Deficiency Syndrome

Monoallelic PMS2 germline mutations cause 5%–15% of Lynch syndrome, a midlife cancer predisposition, whereas biallelic PMS2 mutations cause approximately 60% of constitutional mismatch repair deficiency (CMMRD), a rare childhood cancer syndrome. Recently improved DNA‐ and RNA‐based strategies are applied to overcome problematic PMS2 mutation analysis due to the presence of pseudogenes and frequent gene conversion events. Here, we determined PMS2 mutation detection yield and mutation spectrum in a nationwide cohort of 396 probands. Furthermore, we studied concordance between tumor IHC/MSI (immunohistochemistry/microsatellite instability) profile and mutation carrier state. Overall, we found 52 different pathogenic PMS2 variants explaining 121 Lynch syndrome and nine CMMRD patients. In vitro mismatch repair assays suggested pathogenicity for three missense variants. Ninety‐one PMS2 mutation carriers (70%) showed isolated loss of PMS2 in their tumors, for 31 (24%) no or inconclusive IHC was available, and eight carriers (6%) showed discordant IHC (presence of PMS2 or loss of both MLH1 and PMS2). Ten cases with isolated PMS2 loss (10%; 10/97) harbored MLH1 mutations. We confirmed that recently improved mutation analysis provides a high yield of PMS2 mutations in patients with isolated loss of PMS2 expression. Application of universal tumor prescreening methods will however miss some PMS2 germline mutation carriers.     

The preliminary effectiveness of migraine lay trainers in a home-based behavioural management training

OBJECTIVES: This pilot study examined the effectiveness and trainer skills of the first migraine lay trainers (MLTs). METHODS: In a stepwise training program eight MLTs participated in a behavioural management training (BMT) aimed at the prevention of migraine attacks by proactive relaxation and trigger management. After successful reduction of their migraine attacks, three MLTs provided BMT under supervision at home to one fellow patient and subsequently to a small group. RESULTS: Migraine frequency was significantly reduced in five out of eight patients trained by MLTs (mean 48%) and medication use decreased substantially in four patients (mean 47%). Qualities of MLTs concerned their motivational assistance, knowledge of premonitory symptoms and exchange of disease specific problems. Pitfalls were that migraine symptoms hampered an active guidance of the sessions and providing tailored feedback was difficult. CONCLUSION: The first MLTs were successful in training fellow patients in behavioural prevention of migraine attacks. PRACTICE IMPLICATIONS: Continuous supervision of MLTs health and trainer skills is recommended but is likely to have implications for cost-effectiveness.     

Chronic Q Fever Diagnosis—Consensus Guideline versus Expert Opinion

Chronic Q fever, caused by Coxiella burnetii, has high mortality and morbidity rates if left untreated. Controversy about the diagnosis of this complex disease has emerged recently. We applied the guideline from the Dutch Q Fever Consensus Group and a set of diagnostic criteria proposed by Didier Raoult to all 284 chronic Q fever patients included in the Dutch National Chronic Q Fever Database during 2006–2012. Of the patients who had proven cases of chronic Q fever by the Dutch guideline, 46 (30.5%) would not have received a diagnosis by the alternative criteria designed by Raoult, and 14 (4.9%) would have been considered to have possible chronic Q fever. Six patients with proven chronic Q fever died of related causes. Until results from future studies are available, by which current guidelines can be modified, we believe that the Dutch literature-based consensus guideline is more sensitive and easier to use in clinical practice.     

Evaluation of yield and experiences of age-related molecular investigation for heritable and nonheritable causes of mismatch repair deficient colorectal cancer to identify Lynch syndrome

Universal mismatch repair deficiency (dMMR) testing of colorectal cancer (CRC) is promoted as routine diagnostics to prescreen for Lynch syndrome. We evaluated the yield and experience of age-related molecular investigation for heritable and nonheritable causes of dMMR in CRC below age 70 to identify Lynch Syndrome. In a prospective cohort of 3602 newly diagnosed CRCs below age 70 from 19 hospitals, dMMR, MLH1 promoter hypermethylation, germline MMR gene and somatic MMR gene testing was assessed in daily practice. Yield was evaluated using data from the Dutch Pathology Registry (PALGA) and two regional genetic centers. Experiences of clinicians were evaluated through questionnaires. Participating clinicians were overwhelmingly positive about the clinical workflow. Pathologists routinely applied dMMR-testing in 84% CRCs and determined 10% was dMMR, largely due to somatic MLH1 hypermethylation (66%). Of those, 69% with dMMR CRC below age 70 without hypermethylation were referred for genetic testing, of which 55% was due to Lynch syndrome (hereditary) and 43% to somatic biallelic pathogenic MMR (nonhereditary). The prevalence of Lynch syndrome was 18% in CRC < 40, 1.7% in CRC age 40-64 and 0.7% in CRC age 65-69. Age 65-69 represents most cases with dMMR, in which dMMR due to somatic causes (13%) is 20 times more prevalent than Lynch syndrome. In conclusion, up to age 65 routine diagnostics of (non-)heritable causes of dMMR CRCs effectively identifies Lynch syndrome and reduces Lynch-like diagnoses. Above age 64, the effort to detect one Lynch syndrome patient in dMMR CRC is high and germline testing rarely needed.     

89Zr-trastuzumab PET visualises HER2 downregulation by the HSP90 inhibitor NVP-AUY922 in a human tumour xenograft

NVP-AUY922, a potent heat shock protein (HSP) 90 inhibitor, downregulates the expression of many oncogenic proteins, including the human epidermal growth factor receptor-2 (HER2). Because HER2 downregulation is a potential biomarker for early response to HSP90-targeted therapies, we used the ⁸⁹Zr-labelled HER2 antibody trastuzumab to quantify the alterations in HER2 expression after NVP-AUY922 treatment with HER2 positron emission tomography (PET) imaging.The HER2 overexpressing human SKOV-3 ovarian tumour cell line was used for in vitro experiments and as xenograft model in nude athymic mice. In vitro HER2 membrane expression was assessed by flow cytometry and a radio-immuno assay with ⁸⁹Zr-trastuzumab. For in vivo evaluation, mice received 50 mg/kg NVP-AUY922 intraperitoneally every other day. ⁸⁹Zr-trastuzumab was injected intravenously 6 d before NVP-AUY922 treatment and after 3 NVP-AUY922 doses. MicroPET imaging was performed at 24, 72 and 144 h post tracer injection followed by ex-vivo biodistribution and immunohistochemical staining.After 24 h NVP-AUY922 treatment HER2 membrane expression showed profound reduction with flow cytometry (80%) and radio-immuno assay (75%). PET tumour quantification, showed a mean reduction of 41% (p = 0.0001) in ⁸⁹Zr-trastuzumab uptake at 144 h post tracer injection after NVP-AUY922 treatment. PET results were confirmed by ex-vivo ⁸⁹Zr-trastuzumab biodistribution and HER2 immunohistochemical staining.NVP-AUY922 effectively downregulates HER2, which can be monitored and quantified in vivo non-invasively with ⁸⁹Zr-trastuzumab PET. This technique is currently under clinical evaluation and might serve as an early biomarker for HSP90 inhibition in HER2 positive metastatic breast cancer patients.     

Implantation of Artisan toric phakic intraocular lenses for the correction of astigmatism and spherical errors in patients with keratoconus

PURPOSE: To evaluate the correction of astigmatism and spherical ametropia in patients with keratoconus through implantation of an Artisan toric phakic intraocular lens (PIOL) (Ophtec, Groningen, The Netherlands). METHODS: Artisan toric PIOLs were implanted uneventfully in both eyes of three patients with keratoconus with clear central corneas and contact lens intolerance. RESULTS: Best spectacle-corrected subjective visual acuity after lens implantation was unchanged in one eye and improved in five eyes. Spherical equivalent refraction was significantly reduced in all eyes (P=.03). The safety index was 1.49. CONCLUSIONS: The implantation of an Artisan toric PIOL may be an alternative for treating astigmatism and myopia in contact lens intolerant patients with keratoconus with clear central corneas. Especially in patients with associated myopia, this procedure is worth considering before planning a penetrating keratoplasty.     

Mutation and association analyses of the candidate genes ESR1, ESR2, MAX, PCNA, and KAT2A in patients with unexplained MSH2-deficient tumors

Lynch syndrome (Hereditary non-polyposis colorectal cancer/HNPCC) is a cancer susceptibility syndrome which is caused by germline mutations in DNA mismatch repair (MMR) genes, in particular MLH1 and MSH2. A pathogenic germline mutation in the respective MMR gene is suggested by the finding of a loss of a mismatch repair protein in tumor tissue on immunohistochemical staining combined with an early age of onset and/or the familial occurrence of colorectal cancer. Pathogenic germline mutations are identifiable in around 60% of patients suspected of Lynch syndrome, depending on the familial occurrence. The aim of the present study was to identify novel susceptibility genes for Lynch syndrome. 64 Healthy controls and 64 Lynch syndrome patients with no pathogenic MSH2 mutation but a loss of MSH2 expression in their tumor tissue were screened for rare and disease causing germline mutations in the functional candidate genes ESR1, ESR2, MAX, PCNA, and KAT2A. Thirty variants were identified, and these were then genotyped in an independent sample of 36 mutation negative Lynch syndrome patients and 234 controls. Since a trend towards association was observed for KAT2A, an additional set of 21 tagging SNPs was analyzed at this locus in a final case-control sample of 142 mutation negative Lynch syndrome patients and 298 controls. The mutation analysis failed to reveal any rare disease-causing mutations. No association was found at the single-marker or haplotypic level for any common disease-modifying variant. The present results suggest that neither rare nor common genetic variants in ESR1, ESR2, MAX, PCNA, or KAT2A contribute to the development of Lynch syndrome.