Proteomic Profiling of Endothelial Cells Exposed to Mitomycin C: Key Proteins and Pathways Underlying Genotoxic Stress-Induced Endothelial Dysfunction.

Mitomycin C (MMC)-induced genotoxic stress can be considered to be a novel trigger of endothelial dysfunction and atherosclerosis-a leading cause of cardiovascular morbidity and mortality worldwide. Given the increasing genotoxic load on the human organism, the decryption of the molecular pathways underlying genotoxic stress-induced endothelial dysfunction could improve our understanding of the role of genotoxic stress in atherogenesis. Here, we performed a proteomic profiling of human coronary artery endothelial cells (HCAECs) and human internal thoracic endothelial cells (HITAECs) in vitro that were exposed to MMC to identify the biochemical pathways and proteins underlying genotoxic stress-induced endothelial dysfunction. We denoted 198 and 71 unique, differentially expressed proteins (DEPs) in the MMC-treated HCAECs and HITAECs, respectively; only 4 DEPs were identified in both the HCAECs and HITAECs. In the MMC-treated HCAECs, 44.5% of the DEPs were upregulated and 55.5% of the DEPs were downregulated, while in HITAECs, these percentages were 72% and 28%, respectively. The denoted DEPs are involved in the processes of nucleotides and RNA metabolism, vesicle-mediated transport, post-translation protein modification, cell cycle control, the transport of small molecules, transcription and signal transduction. The obtained results could improve our understanding of the fundamental basis of atherogenesis and help in the justification of genotoxic stress as a risk factor for atherosclerosis.

45 years of tetracycline post exposure prophylaxis for STIs and the risk of tetracycline resistance: a systematic review and meta-analysis.

There is considerable interest in the use of doxycycline post exposure prophylaxis (PEP) to reduce the incidence of bacterial sexually transmitted infections (STIs). An important concern is that this could select for tetracycline resistance in these STIs and other species. We searched PubMed and Google Scholar, (1948-2023) for randomized controlled trials comparing tetracycline PEP with non-tetracycline controls. The primary outcome was antimicrobial resistance (AMR) to tetracyclines in all bacterial species with available data. Our search yielded 140 studies, of which three met the inclusion criteria. Tetracycline PEP was associated with an increasedprevalence of tetracycline resistance in Neisseria gonorrhoeae, but this effect was not statistically significant (Pooled OR 2.3, 95% CI 0.9-3.4). PEP had a marked effect on the N. gonorrhoeae tetracycline MIC distribution in the one study where this was assessed. Prophylactic efficacy was 100% at low MICs and 0% at high MICs. In the one study where this was assessed, PEP resulted in a significant increase in tetracycline resistance in commensal Neisseria species compared to the control group (OR 2.9, 95% CI 1.5-5.5) but no significant effect on the prevalence of tetracycline resistance in Staphylococcus aureus. The available evidence suggests that PEP with tetracyclines could be associated with selecting tetracycline resistance in N. gonorrhoeae and commensal Neisseria species.

Designing Molecularly Imprinted Polymer-Modified Boron-Doped Diamond Electrodes for Highly Selective Electrochemical Drug Sensors.

Drug detection in biological solutions is essential in studying the pharmacokinetics of the body. Electrochemical detection is an accurate and rapid method, but measuring multiple drugs that react at similar potentials is challenging. Herein, we developed an electrochemical sensor using a boron-doped diamond (BDD) electrode modified with a molecularly imprinted polymer (MIP) to provide specificity in drug sensing. The MIP is a polymer material designed to recognize and capture template molecules, enabling the selective detection of target molecules. In this study, we selected the anticancer drug doxorubicin (DOX) as the template molecule. In the electrochemical measurements using an unmodified BDD, the DOX reduction was observed at approximately -0.5 V (vs Ag/AgCl). Other drugs, i.e., mitomycin C or clonazepam (CZP), also underwent a reduction reaction at a similar potential to that of DOX, when using the unmodified BDD, which rendered the accurate quantification of DOX in a mixture challenging. Similar measurements conducted in PBS using the MIP-BDD only resulted in a DOX reduction current, with no reduction reaction observed in the presence of mitomycin C and CZP. These results suggest that the MIP, whose template molecule is DOX, inhibits the reduction of other drugs on the electrode surface. Selective DOX measurement using the MIP-BDD was also possible in human plasma, and the respective limits of detection of DOX in PBS and human plasma were 32.10 and 16.61 nM. The MIP-BDD was durable for use in six repeated measurements, and MIP-BDD may be applicable as an electrochemical sensor for application in therapeutic drug monitoring.

Capecitabine/cisplatin combined with concurrent intensity-modulated radiation therapy: a feasible therapeutic strategy for anal squamous cell carcinoma

Purpose To evaluate the efficacy and safety of capecitabine/cisplatin (XP) combined with intensity-modulated radiation therapy (IMRT) in patients with non-metastatic anal squamous cell carcinoma (ASCC). Method and materials All patients with ASCC who received radical concurrent chemoradiotherapy in the past 8 years were screened. Patients who received XP or mitomycin/5-fluorouracil (MF) were selected and analyzed retrospectively. Results ASCC is an uncommon cancer, there were 36 patients were included in our study. The XP group and MF group included 18 patients each. The clinical complete response (cCR) rates in the XP group and the MF group were 94.4% and 88.9%, respectively (P = 1). The 2-year local control (LC), disease-free survival (DFS), and colostomy-free survival (CFS) rates were higher in the XP group than in the MF group (100% vs 93.3%, P = 0.32). Hematologic toxicities, especially grade ≥ 3 leukopenia (11.1% vs 44.4%, P = 0.06) and neutropenia (5.6% vs 61.1%, P = 0.001), were lower in the XP group than MF group. As a result of fewer side effects, fewer patients in the XP group demanded the dose reduction of chemotherapy (11.1% vs 50%, P = 0.03) and radiation interruption (55.6% vs 77.8%, P = 0.289). Delayed radiotherapy was shorter in the XP group (2.5 vs 6.5 days, P = 0.042) than in the MF group. Conclusion The XP regimen was as effective as the MF regimen in non-metastatic ASCC. Compared with the standard MF regimen, XP combined with IMRT showed higher treatment completion and lower toxicities. It could be considered a feasible alternative for patients with non-metastatic ASCC (AU)

Co-Assembly of Cancer Drugs with Cyclo-HH Peptides: Insights from Simulations and Experiments.

Peptide-based nanomaterials can serve as promising drug delivery agents, facilitating the release of active pharmaceutical ingredients while reducing the risk of adverse reactions. We previously demonstrated that Cyclo-Histidine-Histidine (Cyclo-HH), co-assembled with cancer drug Epirubicin, zinc, and nitrate ions, can constitute an attractive drug delivery system, combining drug self-encapsulation, enhanced fluorescence, and the ability to transport the drug into cells. Here, we investigated both computationally and experimentally whether Cyclo-HH could co-assemble, in the presence of zinc and nitrate ions, with other cancer drugs with different physicochemical properties. Our studies indicated that Methotrexate, in addition to Epirubicin and its epimer Doxorubicin, and to a lesser extent Mitomycin-C and 5-Fluorouracil, have the capacity to co-assemble with Cyclo-HH, zinc, and nitrate ions, while a significantly lower propensity was observed for Cisplatin. Epirubicin, Doxorubicin, and Methorexate showed improved drug encapsulation and drug release properties, compared to Mitomycin-C and 5-Fluorouracil. We demonstrated the biocompatibility of the co-assembled systems, as well as their ability to intracellularly release the drugs, particularly for Epirubicin, Doxorubicin, and Methorexate. Zinc and nitrate were shown to be important in the co-assembly, coordinating with drugs and/or Cyclo-HH, thereby enabling drug-peptide as well as drug-drug interactions in successfully formed nanocarriers. The insights could be used in the future design of advanced cancer therapeutic systems with improved properties.

Patterns of treatment of high-risk BCG-unresponsive non-muscle invasive bladder cancer (NMIBC) patients among Arab urologists.

PURPOSE: To understand the treatment plans suggested for BCG-unresponsive non-muscle invasive disease (NMIBC) patients in the Arab countries and therapeutic decisions applied for BCG-naive patients during BCG shortage time. METHODS: A 10-minute online survey was distributed through the Arab Association of Urology (AAU) office to urologists in the Arab countries who treat patients with NMIBC. RESULTS: One hundred six urologists responded to the survey. The majority of urologists had treated, in the past 6 months, > 10 patients with NMIBC who were considered BCG-unresponsive (55% of respondents). Radical cystectomy (RC) was the most popular treatment option (recommended by 50%) for these patients. This was followed by intravesical chemotherapy (30%), repeat BCG therapy (12%), resection with ongoing surveillance (8%). Clinical trials and intravenous checkpoint inhibitors were never selected. The most preferred intravesical chemotherapy was by ranking: 60% gemcitabine, 19% mitomycin C, 8% docetaxel, 8% gemcitabine/docetaxel, 4% sequential gemcitabine/mitomycin C, and 1% valrubicin. The use of intravesical chemotherapy appears limited by Arab urologists due to concerns regarding clinical efficacy (fear of progression) and the lack of clear recommendations by urology societies. Given the BCG shortage, which may vary per Arab country, Arab urologists have adjusted by prioritizing BCG for T1 and carcinoma in situ (CIS) patients over Ta, adapting intravesical chemotherapy, and reducing the dose/strength of BCG administered. Most physicians report an eagerness to utilize novel therapies to address the BCG deficit, especially to try intravesical chemotherapy. CONCLUSIONS: Even though Arab urologists are in the majority of cases selecting RC for BCG-unresponsive cases, one-third of them are most recently initiating intravesical chemotherapy as an alternative option. To further assist Arab urologists in the appropriate selection of BCG unresponsive high risk NMIBC patient treatments, enhanced education and pathway protocols are needed.

A hypoxia biomarker does not predict benefit from giving chemotherapy with radiotherapy in the BC2001 randomised controlled trial.

BACKGROUND: BC2001 showed combining chemotherapy (5-FU + mitomycin-C) with radiotherapy improves loco-regional disease-free survival in patients with muscle-invasive bladder cancer (MIBC). We previously showed a 24-gene hypoxia-associated signature predicted benefit from hypoxia-modifying radiosensitisation in BCON and hypothesised that only patients with low hypoxia scores (HSs) would benefit from chemotherapy in BC2001. BC2001 allowed conventional (64Gy/32 fractions) or hypofractionated (55Gy/20 fractions) radiotherapy. An exploratory analysis tested an additional hypothesis that hypofractionation reduces reoxygenation and would be detrimental for patients with hypoxic tumours. METHODS: RNA was extracted from pre-treatment biopsies (298 BC2001 patients), transcriptomic data generated (Affymetrix Clariom-S arrays), HSs calculated (median expression of 24-signature genes) and patients stratified as hypoxia-high or -low (cut-off: cohort median). PRIMARY ENDPOINT: invasive loco-regional control (ILRC); secondary overall survival. FINDINGS: Hypoxia affected overall survival (HR = 1.30; 95% CI 0.99-1.70; p = 0.062): more uncertainty for ILRC (HR = 1.29; 95% CI 0.82-2.03; p = 0.264). Benefit from chemotherapy was similar for patients with high or low HSs, with no interaction between HS and treatment arm. High HS associated with poor ILRC following hypofractionated (n = 90, HR 1.69; 95% CI 0.99-2.89 p = 0.057) but not conventional (n = 207, HR 0.70; 95% CI 0.28-1.80, p = 0.461) radiotherapy. The finding was confirmed in an independent cohort (BCON) where hypoxia associated with a poor prognosis for patients receiving hypofractionated (n = 51; HR 14.2; 95% CI 1.7-119; p = 0.015) but not conventional (n = 24, HR 1.04; 95% CI 0.07-15.5, p = 0.978) radiotherapy. INTERPRETATION: Tumour hypoxia status does not affect benefit from BC2001 chemotherapy. Hypoxia appears to affect fractionation sensitivity. Use of HSs to personalise treatment needs testing in a biomarker-stratified trial. FUNDING: Cancer Research UK, NIHR, MRC.

Two-year outcomes of anterior versus posterior scleral application of mitomycin C-soaked sponge in trabeculectomy.

PURPOSE: To compare the safety and efficacy of two different application methods of mitomycin C (MMC)-soaked sponge in trabeculectomy. STUDY DESIGN: Retrospective study. METHODS: We included 71 eyes of 71 patients that had undergone trabeculectomy. In the anterior scleral application group, 36 eyes were treated using the long side of the MMC-soaked sponge placed parallel to the limbus. The efficacy and safety in these eyes were compared with eyes treated with the posterior scleral application group, consisting of 35 eyes treated with the long side of the MMC-soaked sponge placed perpendicular to the limbus. The follow-up period was 2 years. The safety of the procedure, including bleb morphology and complications, was the primary outcome, while the success rate was the secondary outcome. RESULTS: The cumulative success rate at 2 years postoperatively was 94.4% in the anterior and and 94.3% in the posterior scleral application group (P = 1.000). However, with the posterior scleral application of the MMC-soaked sponge, blebs were more low-lying (P = 0.048), less in extent (P < 0.001), more normally vascularized (P = 0.027) and more posteriorly directed (P < 0.001). Furthermore, the incidence of thin-walled cystic bleb (P = 0.028) and bleb leakage (P = 0.025) was significantly lower in the posterior scleral application group than in the anterior group. CONCLUSION: Although there were similar success rates, the posterior scleral application of MMC-soaked sponge with trabeculectomy was safer with a better bleb morphology than the anterior scleral application.

A Secreted BBE-Like Enzyme Acting as a Drug-Binding Efflux Carrier Confers Microbial Self-Resistance to Mitomycin C.

The berberine bridge enzyme (BBE)-like flavoproteins have attracted continuous attention for their capability to catalyze various oxidative reactions. Here we demonstrate that MitR, a secreted BBE-like enzyme, functions as a special drug-binding efflux protein evolved from quinone reductase. Moreover, this protein provides self-resistance to its hosts toward the DNA-alkylating agent mitomycin C with a distinctive strategy, featured by independently performing drug binding and efflux.

BCG and Alternative Therapies to BCG Therapy for Non-Muscle-Invasive Bladder Cancer.

Bladder cancer is a heterogeneous disease. Treatment decisions are mostly decided based on disease stage (non-muscle invasive or muscle invasive). Patients with muscle-invasive disease will be offered a radical treatment combined with systemic therapy, while in those with non-muscle-invasive disease, an attempt to resect the tumor endoscopically will usually be followed by different intravesical instillations. The goal of intravesical therapy is to decrease the recurrence and/or progression of the tumor. In the current landscape of bladder cancer treatment, BCG is given intravesically to induce an inflammatory response and recruit immune cells to attack the malignant cells and induce immune memory. While the response to BCG treatment has changed the course of bladder cancer management and spared many "bladders", some patients may develop BCG-unresponsive disease, leaving radical surgery as the best choice of curative treatment. As a result, a lot of effort has been put into identifying novel therapies like systemic pembrolizumab and Nadofaragene-Firadenovac to continue sparing bladders if BCG is ineffective. Moreover, recent logistic issues with BCG production caused a worldwide BCG shortage, re-sparking interest in alternative BCG treatments including mitomycin C, sequential gemcitabine with docetaxel, and others. This review encompasses both the historic and current role of BCG in the treatment of non-muscle-invasive bladder cancer, revisiting BCG alternative therapies and reviewing the novel therapeutics that were approved for the BCG-unresponsive stage or are under active investigation.

Self-Reinforced Bimetallic Mito-Jammer for Ca2+ Overload-Mediated Cascade Mitochondrial Damage for Cancer Cuproptosis Sensitization.

Overproduction of reactive oxygen species (ROS), metal ion accumulation, and tricarboxylic acid cycle collapse are crucial factors in mitochondria-mediated cell death. However, the highly adaptive nature and damage-repair capabilities of malignant tumors strongly limit the efficacy of treatments based on a single treatment mode. To address this challenge, a self-reinforced bimetallic Mito-Jammer is developed by incorporating doxorubicin (DOX) and calcium peroxide (CaO2) into hyaluronic acid (HA) -modified metal-organic frameworks (MOF). After cellular, Mito-Jammer dissociates into CaO2 and Cu2+ in the tumor microenvironment. The exposed CaO2 further yields hydrogen peroxide (H2O2) and Ca2+ in a weakly acidic environment to strengthen the Cu2+-based Fenton-like reaction. Furthermore, the combination of chemodynamic therapy and Ca2+ overload exacerbates ROS storms and mitochondrial damage, resulting in the downregulation of intracellular adenosine triphosphate (ATP) levels and blocking of Cu-ATPase to sensitize cuproptosis. This multilevel interaction strategy also activates robust immunogenic cell death and suppresses tumor metastasis simultaneously. This study presents a multivariate model for revolutionizing mitochondria damage, relying on the continuous retention of bimetallic ions to boost cuproptosis/immunotherapy in cancer.

Sensing chemical-induced DNA damage using CRISPR/Cas9-mediated gene-deletion yeast-reporter strains.

Microorganism-based genotoxicity assessments are vital for evaluating potential chemical-induced DNA damage. In this study, we developed both chromosomally integrated and single-copy plasmid-based reporter assays in budding yeast using a RNR3 promoter-driven luciferase gene. These assays were designed to compare the response to genotoxic chemicals with a pre-established multicopy plasmid-based assay. Despite exhibiting the lowest luciferase activity, the chromosomally integrated reporter assay showed the highest fold induction (i.e., the ratio of luciferase activity in the presence and absence of the chemical) compared with the established plasmid-based assay. Using CRISPR/Cas9 technology, we generated mutants with single- or double-gene deletions, affecting major DNA repair pathways or cell permeability. This enabled us to evaluate reporter gene responses to genotoxicants in a single-copy plasmid-based assay. Elevated background activities were observed in several mutants, such as mag1Δ cells, even without exposure to chemicals. However, substantial luciferase induction was detected in single-deletion mutants following exposure to specific chemicals, including mag1Δ, mms2Δ, and rad59Δ cells treated with methyl methanesulfonate; rad59Δ cells exposed to camptothecin; and mms2Δ and rad10Δ cells treated with mitomycin C (MMC) and cisplatin (CDDP). Notably, mms2Δ/rad10Δ cells treated with MMC or CDDP exhibited significantly enhanced luciferase induction compared with the parent single-deletion mutants, suggesting that postreplication and for nucleotide excision repair processes predominantly contribute to repairing DNA crosslinks. Overall, our findings demonstrate the utility of yeast-based reporter assays employing strains with multiple-deletion mutations in DNA repair genes. These assays serve as valuable tools for investigating DNA repair mechanisms and assessing chemical-induced DNA damage. KEY POINTS: • Responses to genotoxic chemicals were investigated in three types of reporter yeast. • Yeast strains with single- and double-deletions of DNA repair genes were tested. • Two DNA repair pathways predominantly contributed to DNA crosslink repair in yeast.

Use of In Vivo Corneal Confocal Microscopy to Guide Excimer Laser With Adjunctive Mitomycin C for Treatment of Recalcitrant Acanthamoeba Keratitis.

PURPOSE: The aim of this study was to describe 3 cases of recalcitrant Acanthamoeba keratitis (AK) that were successfully treated using in vivo corneal confocal microscopy (IVCM) to guide excimer laser ablation depth with adjunctive mitomycin C 0.02%. METHODS: Three patients diagnosed with AK did not respond to several weeks of intensive topical therapy with antiamoebic agents. The patient underwent phototherapeutic keratectomy with topical mitomycin C 0.02% application. The maximum stromal depth of cysts measured by IVCM was 80 µm, 100 µm, and 240 µm, and the stromal ablation depths were 80 µm, 100 µm, and 100 µm, respectively. RESULTS: In all 3 eyes, AK resolved after a single excimer laser application, and topical treatment was gradually discontinued within 6 weeks afterward. In 1 eye, penetrating corneal transplantation was performed 6 weeks after phototherapeutic keratectomy because of ongoing severe corneal pain. IVCM and histology of the corneal transplant did not reveal any Acanthamoeba cysts within the excised corneal button. No recurrence was observed during the follow-up period of 19 to 34 months. CONCLUSIONS: IVCM-guided phototherapeutic keratectomy with mitomycin C 0.02% seems to be a safe and successful approach for the treatment of AK, especially in cases of resistance to topical treatment. Corneal IVCM should be performed before laser application to measure cyst depth, determine ablation depth, and assess postoperative treatment success.

Mitochondrial and Cellular Function in Fibroblasts, Induced Neurons, and Astrocytes Derived from Case Study Patients: Insights into Major Depression as a Mitochondria-Associated Disease.

The link between mitochondria and major depressive disorder (MDD) is increasingly evident, underscored both by mitochondria's involvement in many mechanisms identified in depression and the high prevalence of MDD in individuals with mitochondrial disorders. Mitochondrial functions and energy metabolism are increasingly considered to be involved in MDD's pathogenesis. This study focused on cellular and mitochondrial (dys)function in two atypical cases: an antidepressant non-responding MDD patient ("Non-R") and another with an unexplained mitochondrial disorder ("Mito"). Skin biopsies from these patients and controls were used to generate various cell types, including astrocytes and neurons, and cellular and mitochondrial functions were analyzed. Similarities were observed between the Mito patient and a broader MDD cohort, including decreased respiration and mitochondrial function. Conversely, the Non-R patient exhibited increased respiratory rates, mitochondrial calcium, and resting membrane potential. In conclusion, the Non-R patient's data offered a new perspective on MDD, suggesting a detrimental imbalance in mitochondrial and cellular processes, rather than simply reduced functions. Meanwhile, the Mito patient's data revealed the extensive effects of mitochondrial dysfunctions on cellular functions, potentially highlighting new MDD-associated impairments. Together, these case studies enhance our comprehension of MDD.

STIM1 translocation to the nucleus protects cells from DNA damage.

DNA damage represents a challenge for cells, as this damage must be eliminated to preserve cell viability and the transmission of genetic information. To reduce or eliminate unscheduled chemical modifications in genomic DNA, an extensive signaling network, known as the DNA damage response (DDR) pathway, ensures this repair. In this work, and by means of a proteomic analysis aimed at studying the STIM1 protein interactome, we have found that STIM1 is closely related to the protection from endogenous DNA damage, replicative stress, as well as to the response to interstrand crosslinks (ICLs). Here we show that STIM1 has a nuclear localization signal that mediates its translocation to the nucleus, and that this translocation and the association of STIM1 to chromatin increases in response to mitomycin-C (MMC), an ICL-inducing agent. Consequently, STIM1-deficient cell lines show higher levels of basal DNA damage, replicative stress, and increased sensitivity to MMC. We show that STIM1 normalizes FANCD2 protein levels in the nucleus, which explains the increased sensitivity of STIM1-KO cells to MMC. This study not only unveils a previously unknown nuclear function for the endoplasmic reticulum protein STIM1 but also expands our understanding of the genes involved in DNA repair.

SN-38, an active metabolite of irinotecan, inhibits transcription of nuclear factor erythroid 2-related factor 2 and enhances drug sensitivity of colorectal cancer cells.

Nuclear factor erythroid 2-related factor 2 (Nrf2) significantly contributes to drug resistance of cancer cells, and Nrf2 inhibitors have been vigorously pursued. Repurposing of existing drugs, especially anticancer drugs, is a straightforward and promising strategy to find clinically available Nrf2 inhibitors and effective drug combinations. Topoisomerase inhibitors SN-38 (an active metabolite of irinotecan), topotecan, mitoxantrone, and epirubicin were found to significantly suppress Nrf2 transcriptional activity in cancer cells. SN-38, the most potent one among them, significantly inhibited the transcription of Nrf2, as indicated by decreased mRNA level and binding of RNA polymerase II to NFE2L2 gene, while no impact on Nrf2 protein or mRNA degradation was observed. SN-38 synergized with Nrf2-sensitive anticancer drugs such as mitomycin C in killing colorectal cancer cells, and irinotecan and mitomycin C synergistically inhibited the growth of SW480 xenografts in nude mice. Our study identified SN-38 and three other topoisomerase inhibitors as Nrf2 inhibitors, revealed the Nrf2-inhibitory mechanism of SN-38, and indicate that clinically feasible drug combinations could be designed based on their interactions with Nrf2 signaling.

Male with an apparently normal phenotype carrying a BRCA1 exon 20 duplication in trans to a BRCA1 frameshift variant.

BACKGROUND: Reports of dual carriers of pathogenic BRCA1 variants in trans are extremely rare, and so far, most individuals have been associated with a Fanconi Anemia-like phenotype. METHODS: We identified two families with a BRCA1 in-frame exon 20 duplication (Ex20dup). In one male individual, the variant was in trans with the BRCA1 frameshift variant c.2475delC p.(Asp825Glufs*21). We performed splicing analysis and used a transcription activation domain (TAD) assay to assess the functional impact of Ex20dup. We collected pedigrees and mapped the breakpoints of the duplication by long- and short-read genome sequencing. In addition, we performed a mitomycin C (MMC) assay from the dual carrier using cultured lymphoblastoid cells. RESULTS: Genome sequencing and RNA analysis revealed the BRCA1 exon 20 duplication to be in tandem. The duplication was expressed without skipping any one of the two exon 20 copies, resulting in a lack of wild-type transcripts from this allele. TAD assay indicated that the Ex20dup variant has a functional level similar to the well-known moderate penetrant pathogenic BRCA1 variant c.5096G > A p.(Arg1699Gln). MMC assay of the dual carrier indicated a slightly impaired chromosomal repair ability. CONCLUSIONS: This is the first reported case where two BRCA1 variants with demonstrated functional impact are identified in trans in a male patient with an apparently normal clinical phenotype and no BRCA1-associated cancer. The results pinpoint a minimum necessary BRCA1 protein activity to avoid a Fanconi Anemia-like phenotype in compound heterozygous status and yet still predispose carriers to hormone-related cancers. These findings urge caution when counseling families regarding potential Fanconi Anemia risk. Furthermore, prudence should be taken when classifying individual variants as benign based on co-occurrence in trans with well-established pathogenic variants.

Intermethod comparability analyses of gepotidacin antimicrobial susceptibility tests using a large collection of globally collected Escherichia coli and Staphylococcus saprophyticus clinical isolates.

Gepotidacin (GSK2140944) is a novel, bactericidal, first in class triazaacenaphthylene bacterial type II topoisomerase inhibitor in development for the treatment of uncomplicated urinary tract infections and gonorrhea. The performance of several antimicrobial susceptibility methods (broth microdilution, gradient diffusion, and disk diffusion) for gepotidacin were evaluated using over 5800 recent Escherichia coli and Staphylococcus saprophyticus clinical isolates. Reference broth microdilution gepotidacin MICs showed an essential agreement of 95.9 % and 98.1 % with MICs by gradient diffusion for E. coli and S. saprophyticus isolates, respectively. Gepotidacin susceptibility using disks produced by 2 manufacturers had good agreement with an R2 values of 0.95 and 99.2 % of overall zone diameters agreeing within 3 mm. A correlation with an overall R2 value of 0.72 between MICs by broth microdilution and zone diameters by disk diffusion was observed. This data should assist in the clinical development of gepotidacin and provide reliable susceptibility methods to evaluate its activity.

Combination intravesical chemotherapy for non-muscle invasive bladder cancer (NMIBC) as first-line or rescue therapy: where do we stand now?

INTRODUCTION: The combination of intravesical gemcitabine (Gem) with docetaxel (Doce) or with mitomycin C (MMC) has been used in the primary setting as an alternative to Bacillus Calmette-Guerin (BCG) to treat high-risk (HR) and intermediate-risk (IR) non-muscle invasive bladder cancer (NMIBC), as well in the rescue setting for patients in whom BCG has failed. AREA COVERED: Efficacy and safety of Gem/Doce and Gem/MMC to treat NMIBC in BCG-naive and failure settings. EXPERT OPINION: In the BCG-naive setting, Gem/Doce was the primary alternative combination therapy reported, with a weighted mean of 12- and 24-month recurrence-free survival (RFS) of 79% and 77% for HR disease and 84% and 76% for IR disease, respectively. In the HR BCG-failure setting, the weighted mean of 12- and 24-month RFS was 60% and 42% for Gem/Doce and 63% and 40% for Gem/MMC. While patients without BCG exposure and papillary disease only benefit the most from Gem/Doce, there is also reasonable efficacy in BCG refractory disease and CIS. Combination therapy is well tolerated, with grade III toxicity reported in less than 1% of patients. Unlike single-agent chemotherapy, intravesical Gem/Doce is considered effective and safe regardless of risk-stratification.

The antifungal metabolites from coculture of Aspergillus fumigatus and Alternaria alternata associated with Coffea arabica.

One new cyclohexenone derivative, asperfumtone A (1) along with six known compounds were obtained from the coculture of Aspergillus fumigatus and Alternaria alternata associated with Coffea arabica. The configuration of 2 was first reported in the research. The structures were determined by extensive spectroscopic analyses, and ECD calculation. Compounds 3, 4 and 7 showed significant antifungal activities against coffee phytopathogens A. alternata and Fusarium incarnatum with MICs of 1 µg/mL. Compounds 1 and 2 showed weak antifungal activities against A. alternata and F. incarnatum with MICs of 32-64 µg/mL.