Association Between ABCB1 Genetic Variants and Persistent Chemotherapy-Induced Alopecia in Women With Breast Cancer.

Importance: Persistent chemotherapy-induced alopecia (pCIA) has been recently described in patients with breast cancer and in its most severe form occurs in up to 10% of these patients. Genetic risk factors associated with pCIA have not been adequately explored. Objective: To identify genetic variants associated with pCIA. Design, Setting, and Participants: In this genetic association study, 215 women with breast cancer treated with docetaxel-based chemotherapy with a follow-up of 1.5 to 10 years after the end of the treatment were recruited retrospectively through 3 hospital oncology units across Spain between 2005 and 2018. Severe pCIA was defined as lack of scalp hair recovery (Common Terminology Criteria for Adverse Events, version 3.0, grade 2) 18 months or more after the end of treatment. Patients with grade 2 pCIA were selected as cases, and those with no sign of residual alopecia 12 months after the end of docetaxel treatment were selected as controls. A genome-wide association study in a discovery phase was conducted, and logistic regression was used to identify variants associated with the risk to develop this adverse effect. The validity of the association was addressed through a replication phase. Exposures: Docetaxel-based chemotherapy. Main Outcomes and Measures: Genotypes of single-nucleotide variants associated with pCIA. Results: In total, 215 women with breast cancer (median age, 51.6 years; interquartile range, 44-60 years) were recruited (173 patients for the discovery phase and 42 patients for the replication phase). In the discovery phase, ABCB1 genetic variants were associated with risk to develop pCIA. In particular, single-nucleotide variation rs1202179, a regulatory variant located in an enhancer element that interacts with the ABCB1 promoter, was associated with the occurrence of pCIA. This finding was validated in the replication cohort (combined odds ratio, 4.05; 95% CI, 2.46-6.67; P = 3.946 × 10-8). This variant is associated with ABCB1 mRNA expression, and the risk allele was associated with decreased ABCB1 expression levels (P = 1.64 × 10-20). Conclusions and Relevance: This is the first study, to our knowledge, that identifies an association between a regulatory variant in the ABCB1 gene and the occurrence of pCIA in patients with breast cancer who were treated with docetaxel-based therapies. This finding suggests an important insight into the biological mechanisms underlying pCIA and opens the opportunity to explore personalized treatment of these patients.

In search of an evidence-based strategy for quality assessment of human tissue samples: report of the tissue Biospecimen Research Working Group of the Spanish Biobank Network.

The purpose of the present work is to underline the importance of obtaining a standardized procedure to ensure and evaluate both clinical and research usability of human tissue samples. The study, which was carried out by the Biospecimen Science Working Group of the Spanish Biobank Network, is based on a general overview of the current situation about quality assurance in human tissue biospecimens. It was conducted an exhaustive review of the analytical techniques used to evaluate the quality of human tissue samples over the past 30 years, as well as their reference values if they were published, and classified them according to the biomolecules evaluated: (i) DNA, (ii) RNA, and (iii) soluble or/and fixed proteins for immunochemistry. More than 130 publications released between 1989 and 2019 were analysed, most of them reporting results focused on the analysis of tumour and biopsy samples. A quality assessment proposal with an algorithm has been developed for both frozen tissue samples and formalin-fixed paraffin-embedded (FFPE) samples, according to the expected quality of sample based on the available pre-analytical information and the experience of the participants in the Working Group. The high heterogeneity of human tissue samples and the wide number of pre-analytic factors associated to quality of samples makes it very difficult to harmonize the quality criteria. However, the proposed method to assess human tissue sample integrity and antigenicity will not only help to evaluate whether stored human tissue samples fit for the purpose of biomarker development, but will also allow to perform further studies, such as assessing the impact of different pre-analytical factors on very well characterized samples or evaluating the readjustment of tissue sample collection, processing and storing procedures. By ensuring the quality of the samples used on research, the reproducibility of scientific results will be guaranteed.

Role of glutathione in the regulation of epigenetic mechanisms in disease.

Epigenetics is a rapidly growing field that studies gene expression modifications not involving changes in the DNA sequence. Histone H3, one of the basic proteins in the nucleosomes that make up chromatin, is S-glutathionylated in mammalian cells and tissues, making Gamma-L-glutamyl-L-cysteinylglycine, glutathione (GSH), a physiological antioxidant and second messenger in cells, a new post-translational modifier of the histone code that alters the structure of the nucleosome. However, the role of GSH in the epigenetic mechanisms likely goes beyond a mere structural function. Evidence supports the hypothesis that there is a link between GSH metabolism and the control of epigenetic mechanisms at different levels (i.e., substrate availability, enzymatic activity for DNA methylation, changes in the expression of microRNAs, and participation in the histone code). However, little is known about the molecular pathways by which GSH can control epigenetic events. Studying mutations in enzymes involved in GSH metabolism and the alterations of the levels of cofactors affecting epigenetic mechanisms appears challenging. However, the number of diseases induced by aberrant epigenetic regulation is growing, so elucidating the intricate network between GSH metabolism, oxidative stress and epigenetics could shed light on how their deregulation contributes to the development of neurodegeneration, cancer, metabolic pathologies and many other types of diseases.

MicroRNA profile in very young women with breast cancer.

BACKGROUND: Breast cancer is rarely diagnosed in very young women (35 years old or younger), and it often presents with distinct clinical-pathological features related to a more aggressive phenotype and worse prognosis when diagnosed at this early age. A pending question is whether breast cancer in very young women arises from the deregulation of different underlying mechanisms, something that will make this disease an entity differentiated from breast cancer diagnosed in older patients. METHODS: We performed a comprehensive study of miRNA expression using miRNA Affymetrix2.0 array on paraffin-embedded tumour tissue of 42 breast cancer patients 35 years old or younger, 17 patients between 45 and 65 years old and 29 older than 65 years. Data were statistically analyzed by t-test and a hierarchical clustering via average linkage method was conducted. Results were validated by qRT-PCR. Putative targeted pathways were obtained using DIANA miRPath online software. RESULTS: The results show a differential and unique miRNA expression profile of 121 miRNAs (p-value <0.05), 96 of those with a FDR-value <0.05. Hierarchical clustering grouped the samples according to their age, but not by subtype nor by tumour characteristics. We were able to validate by qRT-PCR differences in the expression of 6 miRNAs: miR-1228*, miR-3196, miR-1275, miR-92b, miR-139 and miR-1207. Moreover, all of the miRNAs maintained the expression trend. The validated miRNAs pointed out pathways related to cell motility, invasion and proliferation. CONCLUSIONS: The study suggests that breast cancer in very young women appears as a distinct molecular signature. To our knowledge, this is the first time that a validated microRNA profile, distinctive to breast cancer in very young women, has been presented. The miRNA signature may be relevant to open an important field of research in order to elucidate the underlying mechanism in this particular disease, which in a more clinical setting, could potentially help to identify therapeutic targets in this particular set of patients.

High stability of microRNAs in tissue samples of compromised quality.

Degradation of tissue samples limits performing RNA-based molecular studies, but little is known about the potential usefulness of samples of compromised quality for studies focused on miRNAs. In this work we analyze a series of cryopreserved tissue samples (n = 14), frozen samples that underwent a severe thawing process (n = 10), and their paired formalin-fixed paraffin-embedded (FFPE) tissue samples (n = 24) from patients with breast cancer obtained during primary surgical resection and collected in 2011. Quality and integrity analyses of the total and small fraction of RNA were carried out. Recovery of specific RNA molecules (miRNAs hsa-miR-21, hsa-miR-125b, and hsa-miR-191; snoRNA RNU6B; and mRNAs GAPDH and HPRT1) was also analyzed by quantitative RT-PCR. Our results suggest that visualisation of the small RNA electrophoretic profiles obtained using the Agilent 2100 bioanalyzer makes it possible to differentiate between the three groups of samples (optimally frozen, thawed, and FFPE). We demonstrate that specific miRNA molecules can be similarly recovered from different tissue sample sources, which supports their high degree of stability. We conclude that miRNAs are robustly detected irrespective of the quality of the tissue sample. In this regard, a word of caution should be raised before degraded samples are discarded: although prior quality assessment of the biological material to be analyzed is recommended, our work demonstrates that degraded tissue samples are also suitable for miRNA studies.

Epigenetic biomarkers in laboratory diagnostics: emerging approaches and opportunities.

Epigenetics has emerged as a new and promising field in recent years. Lifestyle, stress, drugs, physiopathological situations and pharmacological interventions have a great impact on the epigenetic code of the cells by altering the methylome, miRNA expression and the covalent histone modifications. Since there exists a need to find new biomarkers and improve diagnosis for several diseases, the research on epigenetic biomarkers for molecular diagnostics encourages the translation of this field from the bench to clinical practice. In this context, deciphering intricate epigenetic modifications involved in several molecular processes is a challenge that will be solved in the near future. In this review, the authors present an overview of the high-throughput technologies and laboratory techniques available for epigenetic studies, and also discuss which of them are more reliable to be used in a clinical diagnostic laboratory. In addition, the authors describe the most promising epigenetic biomarkers in lung, colorectal and prostate cancer, in which most advances have been achieved. Finally, the authors describe epigenetic biomarkers in some rare diseases; these rare syndromes are paradigms for a specific impaired molecular pathway, thus providing valuable information on the discovery of new epigenetic biomarkers.

Specific defects in different transcription complexes compensate for the requirement of the negative cofactor 2 repressor in Saccharomyces cerevisiae.

Negative cofactor 2 (NC2) has been described as an essential and evolutionarily conserved transcriptional repressor, although in vitro and in vivo experiments suggest that it can function as both a positive and a negative effector of transcription. NC2 operates by interacting with the core promoter and components of the basal transcription machinery, like the TATA-binding protein (TBP). In this work, we have isolated mutants that suppress the growth defect caused by the depletion of NC2. We have identified mutations affecting components of three different complexes involved in the control of basal transcription: the mediator, TFIIH, and RNA pol II itself. Mutations in RNA pol II include both overexpression of truncated forms of the two largest subunits (Rpb1 and Rpb2) and reduced levels of these proteins. Suppression of NC2 depletion was also observed by reducing the amounts of the mediator essential components Nut2 and Med7, as well as by deleting any of the nonessential mediator components, except Med2, Med3, and Gal11 subunits. Interestingly, the Med2/Med3/Gal11 triad forms a submodule within the mediator tail. Our results support the existence of different components within the basic transcription complexes that antagonistically interact with the NC2 repressor and suggest that the correct balance between the activities of specific positive and negative components is essential for cell growth.