inter and intra tumoral heterogeneity in dna damage evaluated by comet assay in early breast cancer patients

Abstract There are no clinical tools to functionally assess degree of DNA damage in breast cancer. The comet assay is an accepted research tool for assessing DNA damage, however, most cancer studies have assessed lymphocytes as surrogate cells. The aim of this pilot study was to use the comet assay in early breast cancer directly in tumor tissue to compare DNA damage between and within traditionally defined subgroups, and to explore intra-tumoral heterogeneity. Scrapings of tumor and healthy breast tissue were obtained at primary surgery from 104 women. Comet assay was applied to quantitatively assess DNA damage, revealing substantial inter- and intra-subgroup variation. Marked intra-tumoral heterogeneity was evident across all subgroups. The degree of DNA damage for an individual could not be predicted by breast cancer subgroup. Comet assay warrants further study as a potential clinical tool for identification of tumoral DNA damage and ultimately, individualised use of DNA damaging therapy

Sequence variation in the human transcription factor gene POU5F1

<p>Abstract</p> <p>Background</p> <p>POU5F1 expression is required to maintain stem cell pluripotency and for primordial germ cells to retain proliferative capability in embryonic development. Recent evidence suggests that <it>POU5F1 </it>may also be a testicular germ cell carcinoma (TGCC) oncogene, and <it>POU5F1 </it>variation may influence TGCC risk. As an important first step to a genetic association study, we sought to identify all common sequence variants in an 11.3 kb region containing <it>POU5F1</it>, and to describe the linkage disequilibrium patterns, using DNA from individuals of African-descent (AD) and European-descent (ED).</p> <p>Results</p> <p>A higher number of polymorphisms was observed in the AD (n = 102) versus ED (n = 82) population. Among the 41 observed haplotypes, 21 (51%) and 12 (29%) were unique to the AD and ED populations, respectively, while 8 (20%) were observed in both. The number of tagging polymorphisms necessary to explain at least 80% of common variation (minor allele frequency ≥ 0.10) due to the remaining untyped polymorphisms was 17 for an AD and 10 for an ED population, providing a 4.0- and 7.0-fold gain in genotyping efficiency for characterizing nucleotide variation, respectively.</p> <p>Conclusion</p> <p><it>POU5F1 </it>is highly polymorphic, however a smaller subset of polymorphisms can tag the observed genetic variation with little loss of information.</p

Perinatal and 2-year neurodevelopmental outcome in late preterm fetal compromise: the TRUFFLE 2 randomised trial protocol

Introduction: Following the detection of fetal growth restriction, there is no consensus about the criteria that should trigger delivery in the late preterm period. The consequences of inappropriate early or late delivery are potentially important yet practice varies widely around the world, with abnormal findings from fetal heart rate monitoring invariably leading to delivery. Indices derived from fetal cerebral Doppler examination may guide such decisions although there are few studies in this area. We propose a randomised, controlled trial to establish the optimum method of timing delivery between 32 weeks and 36 weeks 6 days of gestation. We hypothesise that delivery on evidence of cerebral blood flow redistribution reduces a composite of perinatal poor outcome, death and short-term hypoxia-related morbidity, with no worsening of neurodevelopmental outcome at 2 years. Methods and analysis: Women with non-anomalous singleton pregnancies 32+0 to 36+6 weeks of gestation in whom the estimated fetal weight or abdominal circumference is &lt;10th percentile or has decreased by 50 percentiles since 18-32 weeks will be included for observational data collection. Participants will be randomised if cerebral blood flow redistribution is identified, based on umbilical to middle cerebral artery pulsatility index ratio values. Computerised cardiotocography (cCTG) must show normal fetal heart rate short term variation (≥4.5 msec) and absence of decelerations at randomisation. Randomisation will be 1:1 to immediate delivery or delayed delivery (based on cCTG abnormalities or other worsening fetal condition). The primary outcome is poor condition at birth and/or fetal or neonatal death and/or major neonatal morbidity, the secondary non-inferiority outcome is 2-year infant general health and neurodevelopmental outcome based on the Parent Report of Children's Abilities-Revised questionnaire. Ethics and dissemination: The Study Coordination Centre has obtained approval from London-Riverside Research Ethics Committee (REC) and Health Regulatory Authority (HRA). Publication will be in line with NIHR Open Access policy. Trial registration number: Main sponsor: Imperial College London, Reference: 19QC5491. Funders: NIHR HTA, Reference: 127 976. Study coordination centre: Imperial College Healthcare NHS Trust, Du Cane Road, London, W12 0HS with Centre for Trials Research, College of Biomedical &amp; Life Sciences, Cardiff University. IRAS Project ID: 266 400. REC reference: 20/LO/0031. ISRCTN registry: 76 016 200

The paradox of Bcl-2: How does paclitaxel convert Bcl-2 function from antiapoptotic to proapoptotic?

Bcl-2 is an antiapoptotic protein often overexpressed in human cancer and exerts an antiapoptotic function at mitochondrial level, through the opening suppression of the permeability transition pore complex (PTPC). This process prevents the consequent leakage of mitochondrial transmembrane potential and effluxing from mitochondria of proapoptotic factors such as Cytochrome C, AIF and APAF-1. Recently, in ovarian cancer cells paclitaxel (PTX)-resistance has been associated to downregulation of Bcl-2. In order to investigate this paradox, human Bcl-2 was stably transfected in PTX-resistant ovarian cancer cells and mitochondria were isolated from these cells. Mitochondria prepared from untransfected control cells showed undetectable levels of Bcl-2 and PTX was unable to modulate the opening of PTPC, as demonstrated by the uptake of the cationic fluorochrome Rhodamine 123. In the same cells transfected with Bcl-2, sensitivity to paclitaxel was restored and the drug was able to induce the opening of PTPC and the leakage of mitochondrial transmembrane potential. Among Bcl-2 family members, the peculiar difference of Bcl-2 consists in the disordered loop domain. Therefore, we prepared stable Bcl-2 transformed cells with a construct devoid of the loop domain (Bcl-2-Δ). In mitochondria prepared from these cells, PTX was unable to modulate the opening of PTPC. These findings suggest that the disordered loop of Bcl-2 is involved in the PTX binding, but did not clarify if this occurs in a direct or indirect way, with the involvement of other proteins. In order to address this issue, we used plasmon resonance-based optical biosensor technology. In vitro Bcl-2 and Bcl-2-Δ translated proteins were immobilised on a optical biochip to permit a real time monitoring of the potential binding of paclitaxel to Bcl-2. The experimental output pointed out that PTX directly binds to Bcl-2, and not to Bcl-2-Δ, this signaling that the PTX binding site is located within the disordered loop domain of the protein. With the aim of better understanding the binding mechanism, we performed molecular modelling investigations, taking as template the experimentally determined structure of class I beta-tubulin in complex with PTX. Results indicated that the putative binding site of PTX in the disordered loop domain is indeed very similar to that present in beta tubulin. In beta-tubulin PTX binds to a pocket with strong interactions with His229, Pro360-Pro359 and the sequence Arg278-Ser277-Thr276-Leu275-Pro274. Similar interactions were found in our paclitaxel/Bcl-2 docked complex: His58, Pro39-Pro40 and the sequence Thr69-Arg68-Ala-67-Val66-Pro65. Due to the similarity in the binding, it is likely that PTX is a peptide-mimicking structure with a structure similar to endogenous agonist(s) able to bind to both beta-tubulin and Bcl-2, and consequently transferring a death signal from microtubules to mitochondria

Paclitaxel Directly Binds to Bcl-2 and Functionally Mimics Activity of Nur77.

We reported previously that Bcl-2 is paradoxically downregulated in paclitaxel-resistant cancer cells. We reveal here that paclitaxel directly targets Bcl-2 in the loop domain, thereby facilitating the initiation of apoptosis. Molecular modeling revealed an extraordinary similarity between the paclitaxel binding sites in Bcl-2 and β-tubulin, leading us to speculate that paclitaxel could be mimetic of an endogenous peptide ligand, which binds both proteins. We tested the hypothesis that paclitaxel mimics Nur77, which, like paclitaxel, changes the function of Bcl-2. This premise was confirmed by Nur77 interacting with both paclitaxel targets (Bcl-2 and β-tubulin) and a peptide sequence mimicking the Nur77 structural region, thus reproducing the paclitaxel-like effects of tubulin polymerization and opening the permeability transition pore channel in mitochondria. This discovery could help in the development of novel anticancer agents with nontaxane skeleton as well as in identifying the clinical subsets responsive to paclitaxel-based therap

Axin1 Protects Colon Carcinogenesis by an Immune-Mediated EffectSummary

Background &amp; Aims: Axin1 is a negative regulator of wingless-type MMTV integration site family, member 1 (Wnt)/β-catenin signaling with tumor-suppressor function. The Wnt pathway has a critical role in the intestine, both during homeostasis and cancer, but the role of Axin1 remains elusive. Methods: We assessed the role of Axin1 in normal intestinal homeostasis, with control, epithelial-specific, Axin1-knockout mice (Axin1ΔIEC) and Axin2-knockout mice. We evaluated the tumor-suppressor function of Axin1 during chemically induced colorectal tumorigenesis and dextran sulfate sodium–induced colitis, and performed comparative gene expression profiling by whole-genome RNA sequencing. The clinical relevance of the Axin1-dependent gene expression signature then was tested in a database of 2239 clinical colorectal cancer (CRC) samples. Results: We found that Axin1 was dispensable for normal intestinal homeostasis and redundant with Axin2 for Wnt pathway down-regulation. Axin1 deficiency in intestinal epithelial cells rendered mice more susceptible to chemically induced colon carcinogenesis, but reduced dextran sulfate sodium–induced colitis by attenuating the induction of a proinflammatory program. RNA-seq analyses identified an interferon γ/T-helper1 immune program controlled by Axin1 that enhances the inflammatory response and protects against CRC. The Axin1-dependent gene expression signature was applied to human CRC samples and identified a group of patients with potential vulnerability to immune checkpoint blockade therapies. Conclusions: Our study establishes, in vivo, that Axin1 has redundant function with Axin2 for Wnt down-regulation and infers a new role for Axin1. Physiologically, Axin1 stimulates gut inflammation via an interferon γ/Th1 program that prevents tumor growth. Linked to its T-cell–mediated effect, the colonic Axin1 signature offers therapeutic perspectives for CRC

Characterization of Stromal Tumor-infiltrating Lymphocytes and Genomic Alterations in Metastatic Lobular Breast Cancer.

Invasive lobular carcinoma (ILC) represents the second most common histologic breast cancer subtype after invasive ductal carcinoma (IDC). While primary ILC has been extensively studied, metastatic ILC has been poorly characterized at the genomic and immune level.info:eu-repo/semantics/publishe