Molecular Diagnosis & Therapy

Interindividual variability in drug response is a major clinical problem. Polymedication and genetic polymorphisms modulating drug-metabolising enzyme activities (cytochromes P450, CYP) are identified sources of variability in drug responses. We present here the relevant data on the clinical impact of the major CYP polymorphisms (CYP2D6, CYP2C19 and CYP2C9) on drug therapy where genotyping and phenotyping may be considered, and the guidelines developed when available. CYP2D6 is responsible for the oxidative metabolism of up to 25% of commonly prescribed drugs such as antidepressants, antipsychotics, opioids, antiarrythmics and tamoxifen. The ultrarapid metaboliser (UM) phenotype is recognised as a cause of therapeutic inefficacy of antidepressant, whereas an increased risk of toxicity has been reported in poor metabolisers (PMs) with several psychotropics (desipramine, venlafaxine, amitriptyline, haloperidol). CYP2D6 polymorphism influences the analgesic response to prodrug opioids (codeine, tramadol and oxycodone). In PMs for CYP2D6, reduced analgesic effects have been observed, whereas in UMs cases of life-threatening toxicity have been reported with tramadol and codeine. CYP2D6 PM phenotype has been associated with an increased risk of toxicity of metoprolol, timolol, carvedilol and propafenone. Although conflicting results have been reported regarding the association between CYP2D6 genotype and tamoxifen effects, CYP2D6 genotyping may be useful in selecting adjuvant hormonal therapy in postmenopausal women. CYP2C19 is responsible for metabolising clopidogrel, proton pump inhibitors (PPIs) and some antidepressants. Carriers of CYP2C19 variant alleles exhibit a reduced capacity to produce the active metabolite of clopidogrel, and are at increased risk of adverse cardiovascular events. For PPIs, it has been shown that the mean intragastric pH values and the Helicobacter pylori eradication rates were higher in carriers of CYP2C19 variant alleles. CYP2C19 is involved in the metabolism of several antidepressants. As a result of an increased risk of adverse effects in CYP2C19 PMs, dose reductions are recommended for some agents (imipramine, sertraline). CYP2C9 is responsible for metabolising vitamin K antagonists (VKAs), non-steroidal anti-inflammatory drugs (NSAIDs), sulfonylureas, angiotensin II receptor antagonists and phenytoin. For VKAs, CYP2C9 polymorphism has been associated with lower doses, longer time to reach treatment stability and higher frequencies of supratherapeutic international normalised ratios (INRs). Prescribing algorithms are available in order to adapt dosing to genotype. Although the existing data are controversial, some studies have suggested an increased risk of NSAID-associated gastrointestinal bleeding in carriers of CYP2C9 variant alleles. A relationship between CYP2C9 polymorphisms and the pharmacokinetics of sulfonylureas and angiotensin II receptor antagonists has also been observed. The clinical impact in terms of hypoglycaemia and blood pressure was, however, modest. Finally, homozygous and heterozygous carriers of CYP2C9 variant alleles require lower doses of phenytoin to reach therapeutic plasma concentrations, and are at increased risk of toxicity. New diagnostic techniques made safer and easier should allow quicker diagnosis of metabolic variations. Genotyping and phenotyping may therefore be considered where dosing guidelines according to CYP genotype have been published, and help identify the right molecule for the right patient.

BACKGROUND: Identification of variable epidermal growth factor receptor (EGFR) gene mutations in non-small cell lung cancer (NSCLC) is important for the selection of appropriate targeted therapies. This meta-analysis was conducted to provide a worldwide overview of EGFR mutation and submutation (specifically exon 19 deletions, exon 21 L858R substitutions, and others) prevalence, and identify important covariates that influence EGFR mutation status in patients with advanced NSCLC to address this clinical data gap. METHODS: in Ovid were searched for studies published between 2004 and 2019 with cohorts of ≥ 50 adults with EGFR mutations, focusing on stage III/IV NSCLC (≤ 20% of patients with stage I/II NSCLC). Linear mixed-effects models were fitted to EGFR mutation endpoints using logistic transformation (logit), assuming a binomial distribution. The model included terms for an intercept reflecting European studies and further additive terms for other continents. EGFR submutations examined were exon 19 deletions, exon 21 L858R substitutions, and others. RESULTS: Of 3969 abstracts screened, 57 studies were included in the overall EGFR mutation analysis and 74 were included in the submutation analysis relative to the overall EGFR mutation population (Europe, n = 12; Asia, n = 51; North America, n = 5; Central America, n = 1; South America, n = 1; Oceania, n = 1; Global, n = 3). The final overall EGFR mutations model estimated Asian and European prevalence of 49.1% and 12.8%, respectively, and included an additive covariate for the proportion of male patients in a study. There were no significant covariates in the submutation analyses. Most submutations were actionable: exon 19 deletions (49.2% [Asia]; 48.4% [Europe]); exon 21 L858R substitutions (41.1% [Asia]; 29.9% [Europe]). CONCLUSIONS: Although EGFR mutation prevalence was higher in Asian than Western countries, data support worldwide testing for EGFR overall and submutations to inform appropriate targeted treatment decisions.

Autism spectrum disorder (ASD) affects ~ 2% of children in the United States. The etiology of ASD likely involves environmental factors triggering physiological abnormalities in genetically sensitive individuals. One of these major physiological abnormalities is mitochondrial dysfunction, which may affect a significant subset of children with ASD. Here we systematically review the literature on human studies of mitochondrial dysfunction related to ASD. Clinical aspects of mitochondrial dysfunction in ASD include unusual neurodevelopmental regression, especially if triggered by an inflammatory event, gastrointestinal symptoms, seizures, motor delays, fatigue and lethargy. Traditional biomarkers of mitochondrial disease are widely reported to be abnormal in ASD, but appear non-specific. Newer biomarkers include buccal cell enzymology, biomarkers of fatty acid metabolism, non-mitochondrial enzyme function, apoptosis markers and mitochondrial antibodies. Many genetic abnormalities are associated with mitochondrial dysfunction in ASD, including chromosomal abnormalities, mitochondrial DNA mutations and large-scale deletions, and mutations in both mitochondrial and non-mitochondrial nuclear genes. Mitochondrial dysfunction has been described in immune and buccal cells, fibroblasts, muscle and gastrointestinal tissue and the brains of individuals with ASD. Several environmental factors, including toxicants, microbiome metabolites and an oxidized microenvironment are shown to modulate mitochondrial function in ASD tissues. Investigations of treatments for mitochondrial dysfunction in ASD are promising but preliminary. The etiology of mitochondrial dysfunction and how to define it in ASD is currently unclear. However, preliminary evidence suggests that the mitochondria may be a fruitful target for treatment and prevention of ASD. Further research is needed to better understand the role of mitochondrial dysfunction in the pathophysiology of ASD.

Oncology indispensably leads us to personalized medicine, which allows an individual approach to be taken with each patient. Personalized oncology is based on pharmacogenomics and the effect of genetic differences in individuals (germline and somatic) on the way cancer patients respond to chemotherapeutics. Biomarkers detected using molecular biology tools allow the molecular characterization of cancer signatures and provide information relevant for personalized treatment. Biomarkers can be divided into two main subgroups: prognostic and predictive. The aim of the application of prognostic biomarkers, which provide information on the overall cancer outcome in patients, is to facilitate cancer diagnosis, usually with no need for putting invasive methods into use. Predictive biomarkers help to optimize therapy decisions, as they provide information on the likelihood of response to a given chemotherapeutic. Among the prognostic factors that identify patients with different outcome risks (e.g., recurrence of the disease), the following factors can be distinguished: somatic and germline mutations, changes in DNA methylation that lead to the enhancement or suppression of gene expression, the occurrence of elevated levels of microRNA (miRNA) capable of binding specific messenger RNA (mRNA) molecules, which affects gene expression, as well as the presence of circulating tumor cells (CTCs) in blood, which leads to a poor prognosis for the patient. Biomarkers for personalized oncology are used mainly in molecular diagnostics of chronic myeloid leukemia, colon, breast and lung cancer, and recently in melanoma. They are successfully used in the evaluation of the benefits that can be achieved through targeted therapy or in the evaluation of toxic effects of the chemotherapeutic used in the therapy.

BACKGROUND: Genomic profiling of solid tumors by next-generation sequencing (NGS) supports personalized therapy selection, but routine clinical adoption requires robust analytical and clinical validation. The Oncomine™ Dx Express Test (ODxET) is a companion diagnostic and tumor profiling in vitro diagnostic NGS assay for detecting targeted single- and multinucleotide variants (SNVs/MNVs), insertions, deletions, copy number variations (CNVs), and RNA fusions or splice variants in formalin-fixed, paraffin-embedded (FFPE) solid tumor samples. METHODS: In a retrospective, blinded analytical study, 462 FFPE samples representing 15 tumor types and including 76 wild-type samples, were analyzed with ODxET and compared with validated orthogonal methods. Reproducibility and repeatability were evaluated across three sites, each using two operators, two sequencers, three reagent lots, and two replicates. Clinical performance for epidermal growth factor receptor (EGFR) exon 20 insertion detection was assessed using non-small cell lung cancer (NSCLC) samples from the WU-KONG1B sunvozertinib clinical trial and commercially sourced biomarker-negative specimens, with clinical effectiveness evaluated in patients treated with the approved sunvozertinib dose. RESULTS: The ODxET assay showed high analytical concordance for EGFR exon 20 insertions, with a positive percent agreement (PPA) of 100% and a negative percent agreement (NPA) of 95.5%. Across the reportable range of profiling variant classes, the ODxET assay also showed high analytical concordance with orthogonal methods for SNVs (PPA = 99.5%, NPA = 100%), insertions and deletions (PPA and NPA = 100%), CNVs (PPA = 94.2%, NPA = 99.3%), and RNA variants (PPA = 95.9%, NPA = 100%). DNA variant repeatability was 100%, except for one BRAF p.V600E instance (NPA = 97.1%). RNA repeatability ranged from 88.9% (ALK fusion) to 100%. Similarly, the ODxET assay showed high clinical concordance with the clinical trial assays for EGFR exon 20 insertions, with a PPA of 97.9%, a NPA of 100%, and an overall percent agreement (OPA) of 98.9%. In the ODxET EGFR exon 20 insertion-positive/clinical trial assay-positive subgroup, response rates were consistent with those observed in the overall sunvozertinib trial population. CONCLUSIONS: These findings support the analytical concordance, reproducibility, and clinical utility of the ODxET assay for molecular profiling of FFPE solid tumor samples.

Manganese-enhanced magnetic resonance imaging (MEMRI) is a functional imaging technique with several potential applications in various research fields. Manganese-enhanced magnetic resonance imaging uses paramagnetic manganese (II) ions (Mn 2+ ) as the contrast enhancer, as they accumulate mainly in the cytosol and mitochondria, shorten the longitudinal relaxation time (T₁) and thereby increase signal intensity on T₁-weighted sequences. Most often used in preclinical neuroimaging, MEMRI is also gaining attention in cancer research, providing potentially new insights into tumour biology and treatment response monitoring. The use of MEMRI faces challenges such as the potential toxicity of free manganese and the need for chelation (mangafodipir trisodium, MnPyC3A), careful dosing below ≈25 µmol Mn 2+ /kg and development of new imaging protocols. Ongoing translational research aims to improve the safety profile of manganese agents and optimise imaging strategies. Early clinical studies in neurology and cardiology are exploring MEMRI; clinical oncology use remains investigational and limited. We review preclinical advances and the limited early human experience, outlining safety considerations and the potential, yet unproven, effectiveness of MEMRI in oncology. The present review also contextualises MEMRI within the broader landscape of metal-based precision imaging and neutron capture therapy approaches, including boron neutron capture therapy and gadolinium neutron capture therapy, which represent complementary strategies exploiting the tumour-selective accumulation of specific metal ions. Recent clinical and translational evidence supports the maturation of these approaches: a phase II trial of accelerator-based boron neutron capture therapy with borofalan in head and neck cancer reported acceptable safety, and a systematic review across malignant tumours confirmed favourable efficacy and safety signals for boron neutron capture therapy. Manganese-based nanotheranostics are concurrently emerging as a route to combine magnetic resonance imaging with image-guided therapy.

Schistosomiasis remains a significant public health issue globally. This study aimed to compare the performance of two molecular diagnostic systems for the detection of low-intensity Schistosoma japonicum infections. A Maxwell kit-based copro-DNA- SjTR1 -qPCR workflow ( SjTR1 -qPCR-S1) and a NaOH-based copro-DNA- SjTR1 -qPCR workflow ( SjTR1 -qPCR-S2) were compared as integrated systems using faecal samples ( n = 554) collected from an S. japonicum -endemic area in Leyte Province, the Philippines. The duplicated Kato–Katz (KK) was performed on three stool samples collected from different days for each participant. The diagnostic performance of KK and the SjTR1 -qPCR systems were comprehensively compared with a composite reference where a participant was classified as positive if the KK was positive or any qPCR assay was positive. The KK, SjTR1 -qPCR-S1 and SjTR1 -qPCR-S2 recorded positivity rates of 25.1% (139/554), 25.5% (141/554) and 29.8% (165/554), respectively. The overall S. japonicum positivity rate was 39.2% (217/554) as determined from the composite reference. Both SjTR1 -qPCR systems exhibited excellent sensitivity in individuals with moderate-heavy (eggs per gram ≥ 100) and light (eggs per gram 10–99) infection intensity, with reduced sensitivity observed only in individuals with extremely low egg burden (eggs per gram 1–9). Globally, SjTR1 -qPCR-S2 demonstrated significantly higher sensitivity (76.0%) compared with the KK (64.1%) and SjTR1 -qPCR-S1 (65.0%) [ p < 0.0161 and p = 0.0021, respectively, χ 2 m ]. For samples positive in both SjTR1 -qPCR systems ( n = 125), SjTR1 -qPCR-S1 showed a significantly higher median Ct value than SjTR1 -qPCR-S2 ( p < 0.0001, Wilcoxon signed-rank test). Both SjTR1 -qPCR systems showed a significant negative correlation between Ct values and faecal egg counts ( ρ = – 0.5947 for SjTR1 -qPCR-S1, and ρ = – 0.4844 for SjTR1 -qPCR-S2, both p < 0.0001). Moderate agreement was observed between both SjTR1 -qPCR systems and the KK ( κ = 0.560 and κ = 0.512, respectively). However, substantial agreement was found between SjTR1 -qPCR-S1 and SjTR1 -qPCR-S2 ( κ = 0.748). The combined qPCR systems showed a sensitivity of 83.4%. SjTR1 -qPCR-S2 is cost effective and less reliant on specialised equipment compared with SjTR1 -qPCR-S1. With further optimisation, the SjTR1 -qPCR-S2 system can be a powerful and game-changing option for integration into the current surveillance network to support the eradication of schistosomiasis japonica.

INTRODUCTION: Achieving clear resection margins (> 5 mm) is critical during laryngeal and hypopharyngeal tumor resection, as positive margins are associated with reduced disease-specific survival. Near-infrared (NIR) fluorescence imaging has emerged as a valuable tool for intraoperative margin assessment by enabling real-time visualization of residual tumor tissue. However, current fluorescent contrast agents rely on intravenous administration, often requiring dosing hours to days before surgery. Also, targeting tumor-specific biomarkers with heterogeneous expression limits applicability. To address these limitations, we evaluated the feasibility of a topically applied pH-activatable fluorophore (PH10), which selectively fluoresces under acidic conditions. This ex vivo proof-of-concept study investigates whether topical PH10 can visualize laryngeal and hypopharyngeal tumors. METHODS: Freshly resected laryngeal and hypopharyngeal squamous cell carcinoma (SCC) specimens were obtained from ten patients and topically sprayed with PH10 immediately after surgery. Following a 2-min incubation period, NIR fluorescence images were acquired before and after three sequential saline washes. Fluorescence intensity was quantified as the tumor-to-background ratio (TBR); tumor localization was confirmed on histopathology. RESULTS: Laryngeal tumors demonstrated tumor-specific fluorescence upon PH10 application, with significantly higher signal intensity in tumor tissue compared to adjacent normal mucosa (P < 0.05). The median TBR after the first wash was 2.3 (interquartile range 1.7-3.0), exceeding the clinically relevant threshold of 1.5 in seven of seven laryngeal SCC cases; however, this threshold was not reached in either of the two hypopharyngeal carcinoma cases included. CONCLUSIONS: This proof-of-concept study suggests that PH10, a sprayable pH-activatable fluorophore, can enable rapid and high-contrast visualization of laryngeal tumors, although this was not observed in the small number of hypopharyngeal cases evaluated. Given its topical application and rapid activation, PH10 may hold promise for intraoperative margin assessment, yet these preliminary findings, based on a small and heterogeneous sample, require validation in larger cohorts, including direct correlation with deep resection margins, before clinical utility can be established.

BACKGROUND: The estrogen receptor-positive, human epidermal growth factor receptor 2-negative metastatic breast cancer (MBC) treatment landscape continues to evolve with targeted therapy development. Estrogen receptor 1 (ESR1) testing is recommended at recurrence or each progression event to identify patients for effective therapies. However, there is limited information on testing and mutation patterns in the real world that can inform patient identification in the clinic. METHODS: A retrospective observational study among patients with estrogen receptor-positive, human epidermal growth factor receptor 2-negative MBC who initiated first line (1L) therapy with aromatase inhibitors or selective estrogen receptor degraders ± CDK4/6 inhibitors from 1 January, 2020 to 31 March, 2025, using the Flatiron Health Research Database was conducted. ESR1 testing and mutation positivity rates at 1L to third line initiation, stratified by endocrine-resistant, 1L endocrine-sensitive, and endocrine-sensitive/de novo MBC were descriptively reported. RESULTS: Overall testing rates varied across subgroups, with one or more ESR1 test per patient in 61.2% of patients with endocrine-resistant MBC (n = 2152), 50.5% in patients with 1L endocrine-sensitive MBC (n = 827), and 43.1% in patients with endocrine-sensitive/de novo MBC (n = 5772). Testing rates at 1L initiation were 36.2% in endocrine-resistant MBC and 19.6% in endocrine-sensitive/de novo MBC. Testing rates were approximately 32% at second line and 25% at third line across subgroups. ESR1 mutation positivity (ESR1m+) rates varied by subgroup at 1L initiation and were highest among patients with endocrine-resistant MBC (24.7%), followed by patients with 1L endocrine-sensitive MBC (6.9%) and patients with endocrine-sensitive/de novo MBC (2.1%). All groups had ESR1m+ rates around 30% at second line initiation and 40% at third line initiation. In an exploratory analysis, ~ 60% of ESR1 tests during 1L were conducted in tissue and ~ 40% in blood. CONCLUSIONS: In this population, real-world ESR1 testing rates were low. One in four patients with endocrine-resistant MBC were ESR1m+ at 1L initiation, ~ 30% were ESR1m+ at second line across subgroups, and ~ 40% were ESR1m+ at third line across subgroups. More blood-based testing at recurrence and progression is recommended in the real world to align with guideline recommendations as ESR1m detection is more sensitive in blood than in tissue.