Germline mutations in checkpoint kinase 2 (CHEK2), a multiple cancer‐predisposing gene, increase breast cancer (BC) risk; however, risk estimates differ substantially in published studies. We analyzed germline CHEK2 variants in 1,928 high‐risk Czech breast/ovarian cancer (BC/OC) patients and 3,360 population‐matched controls (PMCs). For a functional classification of VUS, we developed a complementation assay in human nontransformed RPE1‐CHEK2‐knockout cells quantifying CHK2‐specific phosphorylation of endogenous protein KAP1. We identified 10 truncations in 46 (2.39%) patients and in 11 (0.33%) PMC (p = 1.1 × 10−14). Two types of large intragenic rearrangements (LGR) were found in 20/46 mutation carriers. Truncations significantly increased unilateral BC risk (OR = 7.94; 95%CI 3.90–17.47; p = 1.1 × 10−14) and were more frequent in patients with bilateral BC (4/149; 2.68%; p = 0.003), double primary BC/OC (3/79; 3.80%; p = 0.004), male BC (3/48; 6.25%; p = 8.6 × 10−4), but not with OC (3/354; 0.85%; p = 0.14). Additionally, we found 26 missense VUS in 88 (4.56%) patients and 131 (3.90%) PMC (p = 0.22). Using our functional assay, 11 variants identified in 15 (0.78%) patients and 6 (0.18%) PMC were scored deleterious (p = 0.002). Frequencies of functionally intermediate and neutral variants did not differ between patients and PMC. Functionally deleterious CHEK2 missense variants significantly increased BC risk (OR = 3.90; 95%CI 1.24–13.35; p = 0.009) and marginally OC risk (OR = 4.77; 95%CI 0.77–22.47; p = 0.047); however, carriers low frequency will require evaluation in larger studies. Our study highlights importance of LGR detection for CHEK2 analysis, careful consideration of ethnicity in both cases and controls for risk estimates, and demonstrates promising potential of newly developed human nontransformed cell line assay for functional CHEK2 VUS classification.

Background: Several reports indicate that inherited mutations in the PALB2 gene predispose to breast cancer. However, there is little agreement about the clinical relevance and usefulness of mutation screening in this gene. We analyzed the prevalence and spectrum of germline mutations in PALB2 to estimate their contribution to hereditary breast and/or ovarian cancer in the Czech Republic.Methods: The entire PALB2 coding region was sequenced in 409 breast/ovarian cancer patients negative for BRCA1 and BRCA2 mutations. Testing for large genomic rearrangements (LGR) was performed by multiplex ligation-dependent probe amplification (MLPA) analysis.Results: We have identified 13 different pathogenic alterations including 10 truncating mutations and three LGRs in 16 of 409 patients (3.9%), whereas one truncating mutation was found in a group of 1,226 controls (0.08%; P ¼ 2.6 Â 10

Objective: To analyze methodological influences and characterize the concentrations of cell-free fetal DNA (cffDNA) circulating in maternal plasma at different gestational ages in physiological pregnancies. Methods: We investigated 238 independent samples from single male-bearing pregnancies of different gestation age. In the other 50 pregnancies, the samples were collected three times during pregnancy (at all trimesters) to evaluate the kinetics of cffDNA. The manual and automated DNA extraction methods (Roche) were compared. cffDNA was amplified using real-time PCR method and Y-specific sequences SRY and DYS14. Total cell-free DNA circulating in maternal plasma was determined by the use of the GADPH sequence. Results: The elevation in the concentration of cffDNA during pregnancy with the highest value in the third trimester was observed independently on the DNA extraction method and on the Y-specific amplified sequence. The same is documented for the percentage of fetal DNA in total cell-free DNA in maternal plasma. It increases also in successive trimesters (8.3, 10.7 and 23.2%). Conclusions: We discuss methodological problems and describe statistical parameters of cffDNA concentrations in maternal plasma during pregnancy as the basic information for comparison with pregnancies having a pathological outcome.

BackgroundChoosing the optimal season for conception is a part of family planning since it can positively influence the pregnancy outcome. Changes in the monthly number of infants born with a birth defect can signal prenatal damage - death or malformation – related to a harmful seasonal factor. The aim of our paper was to search for possible seasonal differences in the numbers of new-borns with an orofacial cleft and thus for a period of conception that can increase the risk of orofacial cleft development.MethodsMean monthly numbers of live births in the Bohemia region of the Czech Republic during the years 1964–2000 were compared within a group of 5619 new-borns with various types of orofacial clefts and the control group derived from natality data on 3,080,891 new-borns.ResultsThe control group exhibited regular seasonal variation in the monthly numbers of new-borns: significantly more babies born during March–May and fewer babies born during October–December. Similar natural seasonal variation was also found in the group of babies with an orofacial cleft. However, after subdividing the cleft group according to gender and cleft type, in comparison to controls, significant differences appeared in the number of new-born girls with cleft lip during January–March and in the number of boys born with cleft palate in April – May.ConclusionsWe found significant differences from controls in the number of new-born girls with CL and boys with CP, whose dates of birth correspond to conception from April to August and to the estimated prenatal critical period for cleft formation from May to October. The latter period includes the warm season, when various injurious physical, chemical and biological factors may act on a pregnant woman. This finding should be considered in pregnancy planning. Future studies are necessary to investigate the putative injurious factors during the warm season that can influence pregnancy outcome.Electronic supplementary materialThe online version of this article (10.1186/s12884-018-1981-0) contains supplementary material, which is available to authorized users.