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The Journal of International Medical... Jun 2024Breast cancer, particularly triple-negative breast cancer (TNBC), poses a significant global health burden. Chemotherapy was the mainstay treatment for TNBC patients...
BACKGROUND
Breast cancer, particularly triple-negative breast cancer (TNBC), poses a significant global health burden. Chemotherapy was the mainstay treatment for TNBC patients until immunotherapy was introduced. Studies indicate a noteworthy prevalence (0.2% to 18.6%) of mismatch repair protein (MMRP) deficiency in TNBC, with recent research highlighting the potential of immunotherapy for MMRP-deficient metastatic breast cancer. This study aims to identify MMRP deficiency in TNBC patients using immunohistochemistry.
METHODS
A retrospective cohort study design was used and included TNBC patients treated between 2015 and 2021 at King Hussein Cancer Center. Immunohistochemistry was conducted to assess MMRP expression.
RESULTS
Among 152 patients, 14 (9.2%) exhibited deficient MMR (dMMR). Loss of PMS2 expression was observed in 13 patients, 5 of whom showed loss of MLH1 expression. Loss of MSH6 and MSH2 expression was observed in one patient. The median follow-up duration was 44 (3-102) months. Despite the higher survival rate (80.8%, 5 years) of dMMR patients than of proficient MMR patients (62.3%), overall survival did not significantly differ between the two groups.
CONCLUSION
Approximately 9% of TNBC patients exhibit dMMR. dMMR could be used to predict outcomes and identify patients with TNBC who may benefit from immunotherapy.
Topics: Humans; Female; Triple Negative Breast Neoplasms; Middle Aged; Adult; Retrospective Studies; Mismatch Repair Endonuclease PMS2; Aged; DNA-Binding Proteins; DNA Mismatch Repair; MutL Protein Homolog 1; MutS Homolog 2 Protein; Biomarkers, Tumor; Survival Rate; Immunohistochemistry; Aged, 80 and over; Prognosis
PubMed: 38902203
DOI: 10.1177/03000605241259747 -
International Journal of Molecular... May 2024Pathogenic variants in the gene lead to a systemic disease with karyomegalic interstitial nephritis (KIN) at the forefront clinically. The phenotypic-genotypic features...
Pathogenic variants in the gene lead to a systemic disease with karyomegalic interstitial nephritis (KIN) at the forefront clinically. The phenotypic-genotypic features of a mutation-related disease involving five members of a Hungarian Caucasian family are presented. Each had adult-onset chronic kidney disease of unknown cause treated with renal replacement therapy and elevated liver enzymes. Short stature, emaciation, latte-colored skin, freckles, and a hawk-like nose in four patients, a limited intellect in two patients, and chronic restrictive lung disease in one patient completed the phenotype. Severe infections occurred in four patients. All five patients had ceased. Four patients underwent autopsy. KIN and extrarenal karyomegaly were observed histologically; the livers showed no specific abnormality. The genotyping using formalin-fixed tissue samples detected a hitherto undescribed homozygous mutation (c.1673_1674insT/p.Met558lfs*4; exon 5) in three of these patients and a heterozygous mutation in one patient. The reason for the heterozygosity is discussed. In addition, 56 family members consented to the screening for mutation from which 17 individuals proved to be heterozygous carriers; a blood chemistry evaluation of their kidney and liver function did not find any abnormality. The clinical presentation of FAN1-related disease was multifaceted, and not yet described manifestations were observed besides kidney and liver disease. Mutation in this gene should be suspected in adults with small kidneys of unknown cause, elevated liver enzymes, and recurrent infections, even without a family history.
Topics: Humans; Male; Female; Hungary; Mutation; Adult; Phenotype; Pedigree; Middle Aged; Exodeoxyribonucleases; Multifunctional Enzymes; Endodeoxyribonucleases; Genotype; Renal Insufficiency, Chronic
PubMed: 38892095
DOI: 10.3390/ijms25115907 -
Journal of Translational Medicine Jun 2024Inherited variations in DNA double-strand break (DSB) repair pathway are known to influence ovarian cancer occurrence, progression and treatment response. Despite its...
BACKGROUND
Inherited variations in DNA double-strand break (DSB) repair pathway are known to influence ovarian cancer occurrence, progression and treatment response. Despite its significance, survival-associated genetic variants within the DSB pathway remain underexplored.
METHODS
In the present study, we performed a two-phase analysis of 19,290 single-nucleotide polymorphisms (SNPs) in 199 genes in the DSB repair pathway from a genome-wide association study (GWAS) dataset and explored their associations with overall survival (OS) in 1039 Han Chinese epithelial ovarian carcinoma (EOC) patients. After utilizing multivariate Cox regression analysis with bayesian false-discovery probability for multiple test correction, significant genetic variations were identified and subsequently underwent functional prediction and validation.
RESULTS
We discovered a significant association between poor overall survival and the functional variant GEN1 rs56070363 C > T (CT + TT vs. TT, adjusted hazard ratio (HR) = 2.50, P < 0.001). And the impact of GEN1 rs56070363 C > T on survival was attributed to its reduced binding affinity to hsa-miR-1287-5p and the resultant upregulation of GEN1 mRNA expression. Overexpression of GEN1 aggregated EOC cell proliferation, invasion and migration presumably by influencing the expression of immune inhibitory factors, thereby elevating the proportion of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) and then constructing an immunosuppressive tumor microenvironment.
CONCLUSIONS
In conclusion, GEN1 rs56070363 variant could serve as a potential predictive biomarker and chemotherapeutic target for improving the survival of EOC patients.
Topics: Female; Humans; Middle Aged; Carcinoma, Ovarian Epithelial; Cell Line, Tumor; Cell Movement; Cell Proliferation; China; East Asian People; Gene Expression Regulation, Neoplastic; Genome-Wide Association Study; Kaplan-Meier Estimate; MicroRNAs; Neoplasm Invasiveness; Ovarian Neoplasms; Polymorphism, Single Nucleotide; Prognosis; Survival Analysis; Holliday Junction Resolvases
PubMed: 38890669
DOI: 10.1186/s12967-024-05236-1 -
Nature Communications Jun 2024Holliday junction resolution is a crucial process in homologous recombination and DNA double-strand break repair. Complete Holliday junction resolution requires two...
Holliday junction resolution is a crucial process in homologous recombination and DNA double-strand break repair. Complete Holliday junction resolution requires two stepwise incisions across the center of the junction, but the precise mechanism of metal ion-catalyzed Holliday junction cleavage remains elusive. Here, we perform a metal ion-triggered catalysis in crystals to investigate the mechanism of Holliday junction cleavage by MOC1. We capture the structures of MOC1 in complex with a nicked Holliday junction at various catalytic states, including the ground state, the one-metal ion binding state, and the two-metal ion binding state. Moreover, we also identify a third metal ion that may aid in the nucleophilic attack on the scissile phosphate. Further structural and biochemical analyses reveal a metal ion-mediated allosteric regulation between the two active sites, contributing to the enhancement of the second strand cleavage following the first strand cleavage, as well as the precise symmetric cleavage across the Holliday junction. Our work provides insights into the mechanism of metal ion-catalyzed Holliday junction resolution by MOC1, with implications for understanding how cells preserve genome integrity during the Holliday junction resolution phase.
Topics: DNA, Cruciform; Metals; Holliday Junction Resolvases; Catalytic Domain; Saccharomyces cerevisiae Proteins; Crystallography, X-Ray; Ions; DNA Breaks, Double-Stranded; Models, Molecular; Allosteric Regulation
PubMed: 38886375
DOI: 10.1038/s41467-024-49490-9 -
Medicine Jun 2024In Algeria, the issue of antibiotic resistance is on the rise, being the Staphylococcus aureus infection as a significant concern of hospital-acquired infections. The... (Observational Study)
Observational Study
In Algeria, the issue of antibiotic resistance is on the rise, being the Staphylococcus aureus infection as a significant concern of hospital-acquired infections. The emergence of antibiotic resistance in this bacterium poses a worldwide challenge. The aim of this study aims to establish the incidence of S aureus strains in Algeria as well as identify phenotypic and genotypic resistance based on the "mecA" and "nuc" genes. From 2014 to 2017, a total of 185 S aureus strains were isolated from patients at a hospital in the city of Rouïba, Algiers the number of isolates was slightly higher in males at 58.06% compared to females at 41.94%, resulting in a sex ratio of 1.38. the Oxacillin and Cefoxitin DD test (1 μg oxacillin disk and 30 μg cefoxitin disk) identified 42 strains as resistant. The results indicated high resistance to lactam antibiotics, with penicillin having a 100% resistance rate. There was also significant resistance to oxacillin (51.25%) and cefoxitin (50%). This resistance was frequently associated with resistance to other antibiotic classes, such as aminoglycosides (50%) and Macrolides (28.29%). To confirm methicillin-resistant characteristics, a polymerase chain reaction (PCR) multiplex was conducted on 10 isolates (6 SARM; 4 MSSA) on a phenotypic level. Three isolates tested positive for "mecA," while 7 were negative. All strains carry the nuc gene, which is specific to S aureus. In Algeria, the incidence of S aureus resistance is slightly lower compared to other countries, but it is increasing over time. It is now more crucial than ever to restrict the proliferation of multidrug-resistant strains and reduce undue antibiotic prescriptions. To achieve this, it is vital to keep updated on the epidemiology of this bacterium and its antibiotic susceptibility. This will enable the formulation of appropriate preventive control measures to manage its progression.
Topics: Humans; Anti-Bacterial Agents; Female; Male; Staphylococcus aureus; Staphylococcal Infections; Algeria; Prevalence; Microbial Sensitivity Tests; Bacterial Proteins; Oxacillin; Adult; Penicillin-Binding Proteins; Cefoxitin; Middle Aged; Micrococcal Nuclease; Drug Resistance, Bacterial; Drug Resistance, Multiple, Bacterial; Methicillin-Resistant Staphylococcus aureus
PubMed: 38875387
DOI: 10.1097/MD.0000000000038562 -
Open Forum Infectious Diseases Jun 2024A future (Strep A) vaccine will ideally prevent a significant burden of lower limb cellulitis; however, natural immune responses to proposed vaccine antigens following...
BACKGROUND
A future (Strep A) vaccine will ideally prevent a significant burden of lower limb cellulitis; however, natural immune responses to proposed vaccine antigens following an episode of cellulitis remain uncharacterized.
METHODS
We enrolled 63 patients with cellulitis and 26 with invasive beta hemolytic streptococci infection, using a multiplexed assay to measure immunoglobulin G against Strep A vaccine candidate antigens, including: streptolysin O (SLO), deoxyribonuclease B (DNB), group A carbohydrate (GAC), C5a peptidase (ScpA), cell envelope proteinase (SpyCEP), and adhesion and division protein (SpyAD). Responses in the invasive cohort were used to predict the infecting etiology in the cellulitis cohort.
RESULTS
Of 41 patients with cellulitis and paired serological samples, 68.3% had evidence of beta hemolytic streptococci infection by conventional anti-SLO and/or anti-DNB criteria. A positive serological response to at least 1 of the tested antigens was seen in 78.0% of the cellulitis cohort. Individually, anti-SLO (58.5%), anti-SpyAD (46.3%), and anti-ScpA (39.0%) were the most common. Based on principal component analysis, increases in these 3 antibodies, without responses to DNB, GAC, and SpyCEP characterized (SDSE) infection.
CONCLUSIONS
SDSE appears to be the predominant cause of lower limb cellulitis. Effective Strep A vaccines incorporating antigens that provide additional cross protection against SDSE may prevent a significant burden of lower limb cellulitis.
PubMed: 38872850
DOI: 10.1093/ofid/ofae272 -
Nature Communications Jun 2024Genetic testing is crucial for precision cancer medicine. However, detecting multiple same-site insertions or deletions (indels) is challenging. Here, we introduce CoHIT...
Genetic testing is crucial for precision cancer medicine. However, detecting multiple same-site insertions or deletions (indels) is challenging. Here, we introduce CoHIT (Cas12a-based One-for-all High-speed Isothermal Test), a one-pot CRISPR-based assay for indel detection. Leveraging an engineered AsCas12a protein variant with high mismatch tolerance and broad PAM scope, CoHIT can use a single crRNA to detect multiple NPM1 gene c.863_864 4-bp insertions in acute myeloid leukemia (AML). After optimizing multiple parameters, CoHIT achieves a detection limit of 0.01% and rapid results within 30 minutes, without wild-type cross-reactivity. It successfully identifies NPM1 mutations in 30 out of 108 AML patients and demonstrates potential in monitoring minimal residual disease (MRD) through continuous sample analysis from three patients. The CoHIT method is also competent for detecting indels of KIT, BRAF, and EGFR genes. Integration with lateral flow test strips and microfluidic chips highlights CoHIT's adaptability and multiplexing capability, promising significant advancements in clinical cancer diagnostics.
Topics: Humans; Leukemia, Myeloid, Acute; INDEL Mutation; CRISPR-Cas Systems; Nucleophosmin; Neoplasm, Residual; Nuclear Proteins; Proto-Oncogene Proteins B-raf; Genetic Testing; ErbB Receptors; Bacterial Proteins; Endodeoxyribonucleases; CRISPR-Associated Proteins
PubMed: 38866774
DOI: 10.1038/s41467-024-49414-7 -
PLoS Biology Jun 2024CRISPR-Cas12a, often regarded as a precise genome editor, still requires improvements in specificity. In this study, we used a GFP-activation assay to screen 14 new...
CRISPR-Cas12a, often regarded as a precise genome editor, still requires improvements in specificity. In this study, we used a GFP-activation assay to screen 14 new Cas12a nucleases for mammalian genome editing, successfully identifying 9 active ones. Notably, these Cas12a nucleases prefer pyrimidine-rich PAMs. Among these nucleases, we extensively characterized Mb4Cas12a obtained from Moraxella bovis CCUG 2133, which recognizes a YYN PAM (Y = C or T). Our biochemical analysis demonstrates that Mb4Cas12a can cleave double-strand DNA across a wide temperature range. To improve specificity, we constructed a SWISS-MODEL of Mb4Cas12a based on the FnCas12a crystal structure and identified 8 amino acids potentially forming hydrogen bonds at the target DNA-crRNA interface. By replacing these amino acids with alanine to disrupt the hydrogen bond, we tested the influence of each mutation on Mb4Cas12a specificity. Interestingly, the F370A mutation improved specificity with minimal influence on activity. Further study showed that Mb4Cas12a-F370A is capable of discriminating single-nucleotide polymorphisms. These new Cas12a orthologs and high-fidelity variants hold substantial promise for therapeutic applications.
Topics: Gene Editing; CRISPR-Cas Systems; CRISPR-Associated Proteins; Alleles; Humans; Endodeoxyribonucleases; Animals; Protein Engineering; Bacterial Proteins; Polymorphism, Single Nucleotide; Mutation; DNA; HEK293 Cells
PubMed: 38865309
DOI: 10.1371/journal.pbio.3002680 -
Nature Communications Jun 2024Sickle cell disease is a devastating blood disorder that originates from a single point mutation in the HBB gene coding for hemoglobin. Here, we develop a GMP-compatible...
Sickle cell disease is a devastating blood disorder that originates from a single point mutation in the HBB gene coding for hemoglobin. Here, we develop a GMP-compatible TALEN-mediated gene editing process enabling efficient HBB correction via a DNA repair template while minimizing risks associated with HBB inactivation. Comparing viral versus non-viral DNA repair template delivery in hematopoietic stem and progenitor cells in vitro, both strategies achieve comparable HBB correction and result in over 50% expression of normal adult hemoglobin in red blood cells without inducing β-thalassemic phenotype. In an immunodeficient female mouse model, transplanted cells edited with the non-viral strategy exhibit higher engraftment and gene correction levels compared to those edited with the viral strategy. Transcriptomic analysis reveals that non-viral DNA repair template delivery mitigates P53-mediated toxicity and preserves high levels of long-term hematopoietic stem cells. This work paves the way for TALEN-based autologous gene therapy for sickle cell disease.
Topics: Anemia, Sickle Cell; Gene Editing; Animals; Hematopoietic Stem Cells; Humans; Female; Mice; Genetic Therapy; Transcription Activator-Like Effector Nucleases; Hematopoietic Stem Cell Transplantation; beta-Globins; Tumor Suppressor Protein p53; DNA Repair; Mutation; beta-Thalassemia; Disease Models, Animal; Gene Transfer Techniques
PubMed: 38862518
DOI: 10.1038/s41467-024-49353-3 -
Biosensors & Bioelectronics Sep 2024Developing highly sensitive and specific on-site tests is imperative to strengthen preparedness against future emerging infectious diseases. Here, we describe the...
Developing highly sensitive and specific on-site tests is imperative to strengthen preparedness against future emerging infectious diseases. Here, we describe the construction of a Cas12a-mediated DNAzyme actuator capable of converting the recognition of a specific DNA sequence into an amplified colorimetric signal. To address viral RNA extraction challenges for on-site applications, we developed a rapid and efficient method capable of lysing the viral particles, preserving the released viral RNA, and concentrating the viral RNA. Integration of the DNAzyme actuator with the viral RNA extraction method and loop-mediated isothermal amplification enables a streamlined colorimetric assay for highly sensitive colorimetric detection of respiratory RNA viruses in gargle and saliva. This assay can detect as few as 83 viral particles/100 μL in gargle and 166 viral particles/100 μL in saliva. The entire assay, from sample processing to visual detection, was completed within 1 h at a single controlled temperature. We validated the assay by detecting SARS-CoV-2 in 207 gargle and saliva samples, achieving a clinical sensitivity of 96.3 % and specificity of 100%. The assay is adaptable for detecting specific nucleic acid sequences in other pathogens and is suitable for resource-limited settings.
Topics: Colorimetry; RNA, Viral; SARS-CoV-2; DNA, Catalytic; Biosensing Techniques; Saliva; Humans; Nucleic Acid Amplification Techniques; COVID-19; CRISPR-Associated Proteins; Endodeoxyribonucleases; Limit of Detection; Feces; Bacterial Proteins; Molecular Diagnostic Techniques
PubMed: 38838573
DOI: 10.1016/j.bios.2024.116429