Single loss of a Trp53 allele triggers an increased oxidative, DNA damage and cytokine inflammatory responses through deregulation of IκBα expression

Dose of Trp53, the main keeper of genome stability, influences tumorigenesis; however, the causes underlying and driving tumorigenesis over time by the loss of a single p53 allele are still poorly characterized. Here, we found that single p53 allele loss specifically impacted the oxidative, DNA damage and inflammatory status of hematopoietic lineages. In particular, single Trp53 allele loss in mice triggered oxidative stress in peripheral blood granulocytes and spleenocytes, whereas lack of two Trp53 alleles produced enhanced oxidative stress in thymus cells, resulting in a higher incidence of lymphomas in the Trp53 knockout (KO) mice compared with hemizygous (HEM). In addition, single or complete loss of Trp53 alleles, as well as p53 downregulation, led to a differential increase in basal, LPS- and UVB-induced expression of a plethora of pro-inflammatory cytokine, such as interleukin-12 (Il-12a), TNFα (Tnfa) and interleukin (Il-23a) in bone marrow-derived macrophage cells (BMDMs) compared to WT cells. Interestingly, p53-dependent increased inflammatory gene expression correlated with deregulated expression of the NF-κB pathway inhibitor IκBα. Chromatin immunoprecipitation data revealed decreased p65 binding to Nfkbia in the absence of p53 and p53 binding to Nfkbia promoter, uncovering a novel crosstalk mechanism between p53 and NF-κB transcription factors. Overall, our data suggest that single Trp53 allele loss can drive a sustained inflammatory, DNA damage and oxidative stress response that, over time, facilitate and support carcinogenesis.

Monitoring of genetic diversity in autochthonous Czech poultry breeds assessed by genealogical data

Czech local poultry breeds face high risks of extinction. Because these populations are closed, they are more likely to lose genetic diversity. The aim of this analysis was to determine the loss of genetic diversity in three Czech autochthonous poultry breeds. Pedigree data from a total of 1 932 Czech Gold Speckled Hens, 325 Czech White Geese and 111 Czech Crested Geese registered in studbooks between 2000 and 2018 were evaluated. Data were analysed to determine the major factors that affect the genetic variability of these breeds. The average numbers of equivalent complete generations ranged from 2.53 to 4.82. The effective numbers of founders were from 29 to 59, representing from 43% to 62% of the total number of founders. The effective number of ancestors was estimated in the range of 21 to 41. The average inbreeding coefficient and relatedness coefficient (in parentheses) for the reference populations were 2.0% (6.5%), 1.9% (4.9%) and 2.1% (9.3%), respectively. The results showed that the effective population size derived from the rate of inbreeding ranged from 46 to 108 and if derived from the rate of coancestry it ranged from 35 to 74. With regard to these results, the analysed breeds showed a high probability of allele loss and consequent loss of genetic diversity.

Insights into BRCA Cancer Predisposition from Integrated Germline and Somatic Analyses in 7632 Cancers

Background It is often assumed any cancer in a germline BRCA1 or BRCA2 (collectively termed BRCA) mutation carrier was caused by that mutation. It is also often assumed the occurrence of breast or ovarian cancer in an individual with a variant of uncertain significance (VUS) suggests the VUS is pathogenic. These assumptions have profound management implications for cancer patients and healthy individuals. Methods We compared the frequency of BRCA mutations, allele loss, and Signature 3 in 7632 individuals with 28 cancers and 1000 population controls. Because only increased frequency was the focus of the study, all statistical tests were one-sided. Results Individuals with breast or ovarian cancer had increased germline BRCA pathogenic mutation frequencies compared to controls (P = 1.0x10−10 and P = 1.4x10−34, respectively). There was no increase in other cancer types. Wild-type allele loss and Signature 3 were statistically significantly higher in breast and ovarian cancers with BRCA mutations compared with other cancers with BRCA mutations (P = 5.1x10−10 and P = 3.7x10−9) and cancers without BRCA mutations (P = 2.8x10−53 and P = 1.0x10−134). There was no difference between non-breast and non-ovarian cancers with BRCA mutations and cancers without BRCA mutations. Allele loss and Signature 3 were statistically significantly higher in breast and ovarian cancers in individuals with BRCA pathogenic mutations compared to those with VUS (P = 3.8x10−17 and P = 1.6x10−8) or benign variants (P = 1.2x10−28 and P = 2.2x10−10). There was no difference between individuals with BRCA VUS and those with benign variants. Conclusions These data show that non-breast and non-ovarian cancers in individuals with germline BRCA pathogenic mutations are often not causally related to the mutation and that BRCA VUS are highly unlikely to be pathogenic. These results should reduce inappropriate management of germline BRCA information.

Combination of Complement-Dependent Cytotoxicity and Relative Fluorescent Quantification of HLA Length Polymorphisms Facilitates the Detection of a Loss of Heterozygosity

Loss of heterozygosity (LOH) is a common event in malignant cells. In this work we introduce a new approach to identify patients with loss of heterozygosity in the HLA region either at first diagnosis or after HLA mismatched allogeneic HSCT. Diagnosis of LOH requires a high purity of recipient target cells. FACS is time consuming and also frequently prevented by rather nonspecific or unknown immune phenotype. The approach for recipient cell enrichment is based on HLA targeted complement-dependent cytotoxicity (CDC). Relative fluorescent quantification (RFQ) analysis of HLA intron length polymorphisms then allows analysis of HLA heterozygosity. The approach is exemplified in recent clinical cases illustrating the detection of an acquired allele loss. As illustrated in one case with DPB1, distinct HLA loci in donor and patient were sufficient for both proof of donor cell removal and evaluation of allele loss in the patient's leukemic cells. Results were confirmed using HLA-B RFQ analysis and leukemia-associated aberrant immunophenotype (LAIP) based cell sort. Both results confirmed suspected loss of HLA heterozygosity. Our approach complements or substitutes for FACS-based cell enrichment; hence it may be further developed as novel routine diagnostic tool. This allows rapid recipient cell purification and testing for loss of HLA heterozygosity before and after allogeneic HSCT in easily accessible peripheral blood samples.