Molecular Diagnosis & Therapy最新文献

ESR1 gene mutations represent one of the main mechanisms of acquired resistance to endocrine therapy (ET) in estrogen receptor-positive (ER+) breast cancer. The introduction of liquid biopsy as a minimally invasive technique for analyzing circulating tumor DNA (ctDNA) has opened new avenues for real-time mutation monitoring and personalized treatment strategies. This review explores the clinical relevance of ESR1 mutations in endocrine resistance, the potential of liquid biopsy for early detection and monitoring, and the integration of advanced sequencing technologies and artificial intelligence to improve diagnostic accuracy. Preclinical and clinical studies on key mutations (D538G, Y537S) were analyzed, emerging technologies [(next-generation sequencing (NGS), digital droplet PCR (ddPCR), Cancer Personalized Profiling by deep Sequencing (CAPP-Seq), Targeted Digital Sequencing (TARDIS)] were compared, and survival data from seven major studies were summarized to assess the impact of ESR1 mutations on progression-free survival (PFS) and overall survival (OS). The results show that these mutations, particularly those affecting the ligand-binding domain, are associated with reduced efficacy of aromatase inhibitors and increased tumor aggressiveness. Liquid biopsy proves useful for early detection of resistance mutations and dynamic disease monitoring, but its clinical implementation is limited by low ctDNA levels, technological variability, and the lack of standardized clinical cut-offs. Integration with tissue biopsy, radiomics, and artificial intelligence (AI)-based platforms enhances its clinical utility and prognostic value. In conclusion, liquid biopsy, when combined with advanced technologies and predictive tools, represents an innovative resource for the personalized management of ER+ breast cancer, with the potential to guide timely therapeutic interventions and improve long-term survival.

Background and objective: Human papillomavirus (HPV) is the cause of most cervical cancers and is released as circulating cell-free tumour HPV DNA (ctHPV DNA) into circulation. Earlier studies have indicated that ctHPV DNA is a promising biomarker for analysing treatment response and for recurrence surveillance. However, factors influencing the release of ctHPV DNA, including HPV type and HPV viral load, have not been extensively studied and additional biomarkers for prognosis are needed. Therefore, here we analysed ctHPV DNA, HPV type and viral load in relation to each other and to progression-free survival in patients with locally advanced or advanced cervical cancer.

Methods: Pre-treatment biopsies and blood samples were collected from patients diagnosed with cervical cancer (Federation of Gynecology and Obstetrics [FIGO] stage IB-IV). One hundred and seventeen patients with HPV-positive tumours were included. Human papillomavirus type-specific, droplet digital polymerase chain reaction (ddPCR) assays were used to analyse previously genotyped biopsies for the viral load. Pre-treatment plasma from 92/117 patients were available and analysed for ctHPV DNA and total cell-free DNA levels. Results were related to patient and tumour characteristics and progression-free survival. Patients were grouped based on HPV species where alpha-9-species (including HPV16) and alpha-7-species (including HPV 18) constituted the majority of cases.

Results: Cell-free tumour HPV DNA was found in 83/92 (90.2%) of pre-treatment plasma samples. Higher biopsy viral load was significantly related to a higher ctHPV DNA level. Higher stage and larger primary tumour size were also associated with higher ctHPV DNA level. Alpha-9 species, including HPV16, had a significantly higher viral load (16×), a higher ctHPV DNA level (17×), and a higher detection rate in plasma than alpha-7 species, including HPV18. Alpha-9 species also had significantly better progression-free survival than alpha-7 species. Additional factors leading to better progression-free survival included a lower stage, a lower total cell-free DNA level, a viral load in the 90th percentile and, in the high-risk cervical cancer group, a higher pre-treatment ctHPV DNA level.

Conclusions: Cell-free tumour HPV DNA, HPV type and viral load are promising biomarkers in cervical cancer. The lower sensitivity for ctHPV DNA detection for alpha-7 species, including HPV18, needs to be considered in future studies on ctHPV DNA, especially if used as a marker for relapse during surveillance when ctHPV DNA levels are very low.

Introduction: Trinucleotide repeat expansion in CTG18.1, in intron 2 of TCF4 (MIM *602272, #613267), is the main cause of Fuchs endothelial corneal dystrophy (FECD), accounting for around 75% of cases in Caucasians. CTG18.1 repeat expansion has typically been detected in peripheral blood genomic DNA by Southern blotting or short tandem repeat polymerase chain reaction (STR-PCR) combined with triplet-repeat primed PCR (TP-PCR) if needed. However both methods estimate the size of the expanded repeat relative to a size standard, and the former requires microgram amounts of DNA. To support the development of therapies, a high-throughput screening approach for repeat expansions in FECD is required. Here, we present a sensitive assay using long-range PCR and nanopore sequencing of genomic DNA to accurately resolve the CTG18.1 repeat.

Methods: The CTG18.1 locus was analysed in genomic DNA from peripheral blood leukocytes by two different methods, and results were compared. The first approach used STR-PCR and capillary electrophoresis, followed by confirmatory testing of apparent homozygotes by TP-PCR. The second used long-range PCR, library preparation and long-read sequencing on an Oxford Nanopore Technologies MinION, with resolution of repeat length using the STRique algorithm.

Results: CTG18.1 expansion was screened for in 119 patients with FECD and 83 controls, by STR/TP-PCR genotyping and, independently, by long-range PCR/long-read nanopore sequencing. Both methods gave comparable results, but the latter was also able to measure repeat length. A total of 73.1% of FECD cases (87/119) and 1.2% of age-matched controls (1/83) had at least one CTG18.1 expansion that was ≥ 50 repeats. The expanded CTG18.1 allele was inherited across multiple generations in four larger families, in a manner consistent with causing a dominant phenotype, revealing that some younger family members may be at risk. The G allele of SNP rs599550, ~1kb away from the expansion, is linked (in cis) with expanded alleles in 80.8% of FECD alleles with an expansion, compared with 12.5% in FECD alleles in cases without an expansion and 14.6% in Europeans.

Discussion: We demonstrate that long-range PCR and long-read nanopore sequencing is a sensitive method requiring only nanograms of DNA, which can be scaled up for high-throughput detection and accurate sizing of CTG18.1 in peripheral blood DNA. The SNP, rs599550, is in linkage disequilibrium with the expansion and physically closer than rs613872, previously used in FECD association studies, making it better for use in diagnostic or association studies.

Purpose: Epilepsy is highly prevalent among children, making it one of the most common neurological diseases in the pediatric population. Its diagnosis is problematic, largely depending on clinical judgment and inaccurate investigative studies. Thus, a more objective investigation is warranted. MicroRNAs (miRNAs) are small molecules found in various body fluids and tissues that prove to have potential, as they play an important role in the pathogenesis of epilepsy. This review evaluates the use of miRNAs in the diagnosis, prediction, and prognosis of pediatric epilepsy. Furthermore, it discusses the use of miRNAs as therapeutic agents and the relationship between miRNAs and antiepileptic drugs (AEDs).

Methods: From inception until 7 July 2024, a thorough search of PubMed, Europe PMC, PubMed Central, and Google Scholar was conducted.

Results: Our review is based on 18 studies on pediatric patients with epilepsy according to the selection criteria. A total of 33 different miRNAs for diagnosis, 13 for prediction, and 2 for prognosis of pediatric epilepsy in a total sample size of 663, 111, and 163 pediatric patients, respectively, in addition to 20 miRNAs for the diagnosis of focal epilepsy, and a further 4 for generalized epilepsy in the same population were interpreted in our systematic review.

Conclusions: These studies suggest that miRNA usage as a diagnostic, predictive, and prognostic biomarker for pediatric patients with epilepsy is promising. However, further research on that population is needed, as the number of studies is limited, with several bias concerns and heterogeneity, with the need for clinical trials to assess the use of miRNAs as drug agents. PROSPERO registration no. ID: PROSPERO 2024 CRD42024578258.

Long coronavirus disease (COVID) is a multisystem condition that affects a significant proportion of individuals following severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, with persistent symptoms ranging from fatigue and cognitive dysfunction to cardiovascular disorders. It is estimated that 30-60% of infected individuals experience symptoms lasting more than 12 weeks. Despite advances in understanding acute infection, the pathophysiological mechanisms underlying long COVID remain unclear. Current hypotheses suggest that viral persistence, immune dysfunction, and metabolic alterations play central roles. Omics approaches, including metabolomics, proteomics, and lipidomics, have played a crucial role in investigating molecular changes, identifying biomarkers, and refining therapeutic strategies. This review discusses recent advances in understanding long COVID, addressing its mechanisms, risk factors, the impact of viral variants, and the role of vaccination, with an emphasis on the importance of omics technologies in elucidating this condition.

Early and accurate diagnosis of Alzheimer's disease is crucial for enabling timely intervention and improving patient outcomes. Recent advancements in molecular and imaging methodologies have significantly enhanced the detection of Alzheimer's disease at its early stages and have improved the quality of research in the field. Key molecular approaches include the identification of biomarkers such as amyloid-beta plaques and tau protein tangles, which are central to Alzheimer's disease pathology. These biomarkers can be detected through biofluid analysis or imaging methods, offering high sensitivity, however with disadvantages, which are discussed here. Despite the transition of some of these methods from research settings to clinical practice, several challenges persist, including the need for standardisation across diagnostic platforms and ensuring the accessibility of these advanced technologies within diverse healthcare systems. Additionally, the high cost and requirement for specialised expertise remain significant barriers. Here, we discuss the need to improve the effectiveness of early AD diagnosis, the ongoing research that is being conducted to refine biomarker detection, enhance imaging techniques and develop more cost-effective non-invasive diagnostic approaches. These advancements are essential to overcoming current limitations and ensuring a broader application in clinical settings.

Background: Molecular profiling is quickly becoming standard for patients with advanced cancer, with an increasing number of biomarker-directed therapies and innovative precision diagnostics available. However, with the expansion of relevant biomarkers, clinicians often face challenges obtaining optimal detection from limited tumor tissue. Here, we present biomarker detection rates from comprehensive genomic and immune profiling (CGIP) performed as a component of routine clinical care using a multi-modal testing strategy.

Methods: CGIP was performed on 20,645 solid tumor specimens in a CAP/CLIA and NYS CLEP-certified reference laboratory, including DNA- and RNA-based next-generation sequencing (NGS), RNA gene expression profiling, and PD-L1 immunohistochemistry (IHC). RNA and DNA were co-extracted to optimize tissue usage. Clinical significance of detected biomarkers was classified in accordance with the joint consensus recommendations of the Association for Molecular Pathology (AMP), American Society of Clinical Oncology (ASCO), and the College of American Pathologists (CAP).

Results: Adequacy of specimens for analysis with each test component varied from 99.8% (20,612) for PD-L1 IHC to 87.7% (18,113) for RNA-based NGS. DNA-based NGS had a > 96.0% success rate across all result components (short variants, copy number alterations, and genomic signatures), while RNA-based NGS and gene expression profiling were successful for 92.1% (16,689) and 90.2% (17,275) of cases, respectively. Median turnaround time from specimen receipt in the testing laboratory to report delivery was 8 days (range 1-35). Within our cohort of 15,815 cases with complete results available, 61.0% (9650) had at least one tier 1 biomarker with known clinical significance, 88.8% (14,039) had at least one tier 2 biomarker with potential clinical significance, 57.5% (9,090) had both tier 1 and 2 biomarkers, and 7.7% (1216) had no clinically significant biomarkers detected. Biomarker detection rates varied across tumor types, increasing with the addition of testing modalities.

Conclusions: Utilization of a multi-modal CGIP testing strategy resulted in a high rate of test success and detection of clinically relevant biomarkers while optimizing tissue usage.

Prademagene zamikeracel (ZEVASKYN™) is an autologous cell sheet-based gene therapy developed by Abeona Therapeutics Inc. for the treatment of wounds in patients with recessive dystrophic epidermolysis bullosa. Prademagene zamikeracel contains the patient's own genetically modified cells with functional copies of the COL7A1 gene, as patients lacking functional copies of this gene may develop chronic open wounds caused by the separation of dermal layers. This article summarizes the milestones in the development of prademagene zamikeracel leading to this first approval for treatment of wounds in adult and paediatric patients with recessive dystrophic epidermolysis bullosa.

Fatty acid desaturases are key enzymes in lipid metabolism, crucial for converting saturated fatty acids into monounsaturated fatty acids. This review examines the roles of stearoyl-CoA desaturase, fatty acid desaturase 1, fatty acid desaturase 2 and fatty acid desaturase 3 in health and disease, highlighting their impact on cellular functions such as membrane fluidity, signalling and energy homeostasis. Dysregulation of desaturase activity can have significant implications for human health, contributing to the development of various pathological conditions, including cardiovascular disease, metabolic disorders and cancer. Fatty acid desaturase 1 and fatty acid desaturase 2 overexpression in cancer correlates with tumour growth and chemoresistance, marking them as potential therapeutic targets. The development of inhibitors targeting stearoyl-CoA desaturase and fatty acid desaturase 1 offers promising therapeutic avenues for metabolic diseases and cancer. This review underscores the critical roles of desaturases in various diseases. It emphasises the need for ongoing research to develop effective diagnostic, prognostic and therapeutic strategies, ultimately improving patient outcomes in altered fatty acid metabolism conditions.