Pub Date : 2025-09-20DOI: 10.1016/j.clinbiochem.2025.111016
Siarhei A. Dabravolski , Alexander L. Golovyuk , Olga N. Maltseva , Aleksandra S. Utkina , Alikhan Z. Asoyan , Alexander N. Orekhov
Modified low-density lipoproteins (LDL) play a pivotal role in the pathogenesis of atherosclerosis, contributing to plaque formation and vascular inflammation. This review explores how a diverse array of forms of modified LDL, including carbamylated LDL, nitrated LDL, and desialylated LDL, along with various enzymatic modifications, contribute to disease progression. We discuss how these alterations promote the formation of a heterogeneous, highly atherogenic pool of particles known as electronegative LDL (LDL(−)). Mechanistic insights highlight pathways involving upregulated scavenger receptors, foam cell formation, and chronic inflammatory responses. The diagnostic and prognostic implications of LDL(−), including its association with autoimmune conditions like rheumatoid arthritis and chronic conditions such as type 2 diabetes, underscore its potential as a biomarker for cardiovascular risk. Emerging therapies targeting LDL(−), such as nanoformulations of single-chain fragment variable antibodies, demonstrate promising efficacy in reducing lesion size and inflammation without adverse systemic effects. Despite these advances, a critical barrier to clinical translation is the lack of standardised, high-throughput assays suitable for routine laboratory use. Future research must therefore prioritise the development and validation of robust clinical assays to quantify these atherogenic particles, a crucial step for establishing their role in advanced risk stratification and for guiding novel therapeutic strategies..
{"title":"Modified LDL variants in atherosclerosis: molecular pathways, diagnostic potential, and therapeutic perspectives","authors":"Siarhei A. Dabravolski , Alexander L. Golovyuk , Olga N. Maltseva , Aleksandra S. Utkina , Alikhan Z. Asoyan , Alexander N. Orekhov","doi":"10.1016/j.clinbiochem.2025.111016","DOIUrl":"10.1016/j.clinbiochem.2025.111016","url":null,"abstract":"<div><div>Modified low-density lipoproteins (LDL) play a pivotal role in the pathogenesis of atherosclerosis, contributing to plaque formation and vascular inflammation. This review explores how a diverse array of forms of modified LDL, including carbamylated LDL, nitrated LDL, and desialylated LDL, along with various enzymatic modifications, contribute to disease progression. We discuss how these alterations promote the formation of a heterogeneous, highly atherogenic pool of particles known as electronegative LDL (LDL(−)). Mechanistic insights highlight pathways involving upregulated scavenger receptors, foam cell formation, and chronic inflammatory responses. The diagnostic and prognostic implications of LDL(−), including its association with autoimmune conditions like rheumatoid arthritis and chronic conditions such as type 2 diabetes, underscore its potential as a biomarker for cardiovascular risk. Emerging therapies targeting LDL(−), such as nanoformulations of single-chain fragment variable antibodies, demonstrate promising efficacy in reducing lesion size and inflammation without adverse systemic effects. Despite these advances, a critical barrier to clinical translation is the lack of standardised, high-throughput assays suitable for routine laboratory use. Future research must therefore prioritise the development and validation of robust clinical assays to quantify these atherogenic particles, a crucial step for establishing their role in advanced risk stratification and for guiding novel therapeutic strategies..</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111016"},"PeriodicalIF":2.1,"publicationDate":"2025-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145118121","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-13DOI: 10.1016/j.clinbiochem.2025.111013
Chunyan Zhang , Nan Bai , Yanling Yang , Yuxuan Cheng , Xiyu He , Meng Wang , Honghui Zhou , Yaping Tian
Objectives
Metabolic disorders in newborns can cause significant morbidity and mortality if not diagnosed and managed early. In this study, we developed and validated a reliable LC-MS/MS method to simultaneously quantify methylmalonic acid, methylcitric acid, malonic acid, ethylmalonic acid, and total homocysteine associated with metabolic disorders.
Design and methods
According to Clinical Laboratory Standards Institute (CLSI) guidelines, various analytical performances (i.e., precision, linearity, accuracy and reference intervals) were rigorously evaluated. Meanwhile, sample preparation, chromatographic separation and mass spectrometric detection and incorporating stable isotope-labeled internal standards were optimized. Furthermore, calibrators and quality controls were developed to ensure standardization and traceability.
Results
The coefficient of variations for precision were less than 10.0 %. Meanwhile, the results showed high accuracy (recoveries: 94.57 %–109.60 %) and robust linearity (R2 > 0.9935) over clinically relevant ranges. The limits of detection and limits of quantification for all analytes were established, enabling sensitive detection of pathological elevations. Reference intervals for pediatric populations (ages 0–1 month and 2 months to 18 years) were determined, providing essential baselines for clinical interpretation.
Conclusions
The newly developed method demonstrated good analytical performance and offers a standardized, high-throughput solution for clinical laboratories to improve early diagnosis and personalized management of metabolic diseases.
{"title":"Reliable LC-MS/MS method development and validation for the determination of methylmalonic acid, methylcitric acid, malonic acid, ethylmalonic acid, and total homocysteine in dried blood spots","authors":"Chunyan Zhang , Nan Bai , Yanling Yang , Yuxuan Cheng , Xiyu He , Meng Wang , Honghui Zhou , Yaping Tian","doi":"10.1016/j.clinbiochem.2025.111013","DOIUrl":"10.1016/j.clinbiochem.2025.111013","url":null,"abstract":"<div><h3>Objectives</h3><div>Metabolic disorders in newborns can cause significant morbidity and mortality if not diagnosed and managed early. In this study, we developed and validated a reliable LC-MS/MS method to simultaneously quantify methylmalonic acid, methylcitric acid, malonic acid, ethylmalonic acid, and total homocysteine associated with metabolic disorders.</div></div><div><h3>Design and methods</h3><div>According to Clinical Laboratory Standards Institute (CLSI) guidelines, various analytical performances (i.e., precision, linearity, accuracy and reference intervals) were rigorously evaluated. Meanwhile, sample preparation, chromatographic separation and mass spectrometric detection and incorporating stable isotope-labeled internal standards were optimized. Furthermore, calibrators and quality controls were developed to ensure standardization and traceability.</div></div><div><h3>Results</h3><div>The coefficient of variations for precision were less than 10.0 %. Meanwhile, the results showed high accuracy (recoveries: 94.57 %–109.60 %) and robust linearity (R<sup>2</sup> > 0.9935) over clinically relevant ranges. The limits of detection and limits of quantification for all analytes were established, enabling sensitive detection of pathological elevations. Reference intervals for pediatric populations (ages 0–1 month and 2 months to 18 years) were determined, providing essential baselines for clinical interpretation.</div></div><div><h3>Conclusions</h3><div>The newly developed method demonstrated good analytical performance and offers a standardized, high-throughput solution for clinical laboratories to improve early diagnosis and personalized management of metabolic diseases.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111013"},"PeriodicalIF":2.1,"publicationDate":"2025-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145057325","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Cholesterol metabolism dysregulation is recognized as one of the hallmarks of the cancer with highest mortality rate and the second most common malignancy – lung cancer (LC). LDL and HDL particles, the latter being carriers of the antioxidant paraoxonase-1 (PON1), were already proven to be altered in cancer patients. We have tried to investigate in more depth the cholesterol metabolism perturbances in LC, by analzying content of each of the LDL and HDL subclass and (anti)oxidative activity of each of the HDL subclass separately.
Materials and Methods
LDL and HDL subclasses from blood samples of 89 LC patients and 84 healthy subjects were separated and HDL subclasses PON1 activity assessed using Rainwater method and Gugliucci’s zymogram method, respectively.
Results
LC patients had higher relative proportion of HDL 2 particles, lower proportion of HDL 3 particles, and significantly lower activity of PON1 compared to control group (CG). Relative proportion of PON1 activity was higher on HDL 2b fraction and lower on all HDL 3 fractions of LC patients compared to CG. Relative proportions of LDL I and LDL II particles were increased, while proportions of LDL IV and small dense LDL particles were decreased in LC patients. Relative proportions of HDL and LDL subfractions and PON1 activities on HDL subfractions were found to be dependent on LC type and size, number of comorbidities and sites of progression, and overall response to therapy.
Conclusion
PON1 activity and lipoprotein subfractions distribution seem to be indicators of possible metabolic pathways (disorders) in LC.
{"title":"Assessment of structural and functional characteristics of HDL and LDL in lung cancer patients in order to elucidate mechanisms of cholesterol metabolic pathways disorders","authors":"Milica Belic , Dragana Jovanovic , Milica Miljkovic Trailovic , Miron Sopic , Marina Roksandic-Milenkovic , Vesna Ceriman Krstic , Nemanja Dimic , Jelena Vekic , Aleksandra Zeljkovic , Jelena Kotur-Stevuljevic","doi":"10.1016/j.clinbiochem.2025.111012","DOIUrl":"10.1016/j.clinbiochem.2025.111012","url":null,"abstract":"<div><h3>Introduction</h3><div>Cholesterol metabolism dysregulation is recognized as one of the hallmarks of the cancer with highest mortality rate and the second most common malignancy – lung cancer (LC). LDL and HDL particles, the latter being carriers of the antioxidant paraoxonase-1 (PON1), were already proven to be altered in cancer patients. We have tried to investigate in more depth the cholesterol metabolism perturbances in LC, by analzying content of each of the LDL and HDL subclass and (anti)oxidative activity of each of the HDL subclass separately.</div></div><div><h3>Materials and Methods</h3><div>LDL and HDL subclasses from blood samples of 89 LC patients and 84 healthy subjects were separated and HDL subclasses PON1 activity assessed using Rainwater method and Gugliucci’s zymogram method, respectively.</div></div><div><h3>Results</h3><div>LC patients had higher relative proportion of HDL 2 particles, lower proportion of HDL 3 particles, and significantly lower activity of PON1 compared to control group (CG). Relative proportion of PON1 activity was higher on HDL 2b fraction and lower on all HDL 3 fractions of LC patients compared to CG. Relative proportions of LDL I and LDL II particles were increased, while proportions of LDL IV and small dense LDL particles were decreased in LC patients. Relative proportions of HDL and LDL subfractions and PON1 activities on HDL subfractions were found to be dependent on LC type and size, number of comorbidities and sites of progression, and overall response to therapy.</div></div><div><h3>Conclusion</h3><div>PON1 activity and lipoprotein subfractions distribution seem to be indicators of possible metabolic pathways (disorders) in LC.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111012"},"PeriodicalIF":2.1,"publicationDate":"2025-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145057324","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-10DOI: 10.1016/j.clinbiochem.2025.111011
Xiaozhou Yuan , Donghao Jia , Gefan Wan , Chengbin Wang , Kefu Liu , Yan Meng , Jinyan Duan
Background and aims
Mucopolysaccharidosis (MPS) types I and II are two types of rare lysosomal storage diseases, which lead to the accumulation of glycosaminoglycans due to the lack of the enzyme alpha-L-iduronidase and iduronate 2-sulfatase respectively. There are some similar pathogenic mechanisms and clinical phenotypes but also some specific minute manifestations between these two subtypes.
Materials and methods
We used tandem mass tag mass spectrometry to analyze the differential protein profiles in the urine of MPS I and MPS II patients, and then used parallel reaction monitoring (PRM) to verify our results. We detected the differentially expressed proteins (DEPs) of MPS I and MPS II compared with the control group separately.
Results
We focused on 227 DEPs which showed consistent changes in the urine of both MPS I and MPS II. PRM analysis verified that up-regulated hexosaminidase B and down-regulated hemoglobin alpha-1 showed significant difference in the urine of both subtypes. In addition, we found 391 DEPs by comparative analysis of MPS I and MPS II proteomes and found that DHRS2 contributed to the difference between the two subtypes by PRM verification.
Conclusion
We found that the urine of the two subtypes showed up-regulated HEXB and down regulated HBA1, while DHRS2 was significantly different in the urine of the two subtypes.
{"title":"Urine proteome uncovers common mechanisms between mucopolysaccharidosis types I and II","authors":"Xiaozhou Yuan , Donghao Jia , Gefan Wan , Chengbin Wang , Kefu Liu , Yan Meng , Jinyan Duan","doi":"10.1016/j.clinbiochem.2025.111011","DOIUrl":"10.1016/j.clinbiochem.2025.111011","url":null,"abstract":"<div><h3>Background and aims</h3><div>Mucopolysaccharidosis (MPS) types I and II are two types of rare lysosomal storage diseases, which lead to the accumulation of glycosaminoglycans due to the lack of the enzyme alpha-L-iduronidase and iduronate 2-sulfatase respectively. There are some similar pathogenic mechanisms and clinical phenotypes but also some specific minute manifestations between these two subtypes.</div></div><div><h3>Materials and methods</h3><div>We used tandem mass tag mass spectrometry to analyze the differential protein profiles in the urine of MPS I and MPS II patients, and then used parallel reaction monitoring (PRM) to verify our results. We detected the differentially expressed proteins (DEPs) of MPS I and MPS II compared with the control group separately.</div></div><div><h3>Results</h3><div>We focused on 227 DEPs which showed consistent changes in the urine of both MPS I and MPS II. PRM analysis verified that up-regulated hexosaminidase B and down-regulated hemoglobin alpha-1 showed significant difference in the urine of both subtypes. In addition, we found 391 DEPs by comparative analysis of MPS I and MPS II proteomes and found that DHRS2 contributed to the difference between the two subtypes by PRM verification.</div></div><div><h3>Conclusion</h3><div>We found that the urine of the two subtypes showed up-regulated HEXB and down regulated HBA1, while DHRS2 was significantly different in the urine of the two subtypes.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111011"},"PeriodicalIF":2.1,"publicationDate":"2025-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145045756","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Dihydropyrimidine dehydrogenase (DPD) is the enzyme implicated in the catabolism of the fluoropyrimidines (FP), a class of chemotherapeutics used to treat many cancers. The DPYD gene is known to have a huge number of variants potentially associated with DPD deficiency. Therefore, phenotypic and genotypic characterization of DPD is fundamental for cancer patients before undergoing treatment with FP. The AIOM (Italian Association of Medical Oncology) requires genetic analysis, with the purpose to identify a panel of 5 single nucleotide polymorphisms associated with 5-fluorouracil (5-FU)-induced toxicity, while the biochemical test, which measures uracil levels to predict the residual activity of DPD, is only recommended and not mandatory.
Methods
Single nucleotide polymorphisms were analyzed by real-time polymerase chain reaction (PCR) from peripheral blood. Uracil and dihydrouracil were quantified using an ultra-performance liquid chromatography/tandem mass spectrometry system in plasma obtained from patients before 5-FU treatment. Sanger sequencing was performed for the DPYD gene.
Results
Here, we describe the case of a 70-year-old Caucasian male patient with pancreatic cancer for whom real-time PCR highlighted a heterozygous pathogenic variant c.1905 + 1G > A associated with a 50 % reduction of DPD enzymatic activity. This finding was not confirmed by the biochemical test which revealed a complete absence of DPD activity. By performing Sanger sequencing, we highlighted the concomitant presence of a likely pathogenic variant c.2622 + 1G > A, not compatible with the administration of 5-FU.
Conclusion
Thus far, guidelines provide a limited panel for the identification of pathogenic or likely pathogenic variants of the DPYD gene, therefore we encourage the mandatory use of biochemical tests as a precise diagnostic strategy to detect rare variants and improve drug administration.
{"title":"Phenotyping and genotyping for the dihydropyrimidine dehydrogenase test in Italy: a precise diagnostic strategy to detect rare variants and improve drug administration","authors":"Maria Lucia Tommolini , Mirco Zucchelli , Alberto Frisco , Rossella Ferrante , Beatrice Dufrusine , Claudia Palmarini , Luca Natale , Patrizia Ballerini , Liborio Stuppia , Luca Federici , Damiana Pieragostino , Ilaria Cicalini","doi":"10.1016/j.clinbiochem.2025.111010","DOIUrl":"10.1016/j.clinbiochem.2025.111010","url":null,"abstract":"<div><h3>Introduction</h3><div>Dihydropyrimidine dehydrogenase (DPD) is the enzyme implicated in the catabolism of the fluoropyrimidines (FP), a class of chemotherapeutics used to treat many cancers. The <em>DPYD</em> gene is known to have a huge number of variants potentially associated with DPD deficiency. Therefore, phenotypic and genotypic characterization of DPD is fundamental for cancer patients before undergoing treatment with FP. The AIOM (Italian Association of Medical Oncology) requires genetic analysis, with the purpose to identify a panel of 5 single nucleotide polymorphisms associated with 5-fluorouracil (5-FU)-induced toxicity, while the biochemical test, which measures uracil levels to predict the residual activity of DPD, is only recommended and not mandatory.</div></div><div><h3>Methods</h3><div>Single nucleotide polymorphisms were analyzed by real-time polymerase chain reaction (PCR) from peripheral blood. Uracil and dihydrouracil were quantified using an ultra-performance liquid chromatography/tandem mass spectrometry system in plasma obtained from patients before 5-FU treatment. Sanger sequencing was performed for the <em>DPYD</em> gene.</div></div><div><h3>Results</h3><div>Here, we describe the case of a 70-year-old Caucasian male patient with pancreatic cancer for whom real-time PCR highlighted a heterozygous pathogenic variant c.1905 + 1G > A associated with a 50 % reduction of DPD enzymatic activity. This finding was not confirmed by the biochemical test which revealed a complete absence of DPD activity. By performing Sanger sequencing, we highlighted the concomitant presence of a likely pathogenic variant c.2622 + 1G > A, not compatible with the administration of 5-FU.</div></div><div><h3>Conclusion</h3><div>Thus far, guidelines provide a limited panel for the identification of pathogenic or likely pathogenic variants of the <em>DPYD</em> gene, therefore we encourage the mandatory use of biochemical tests as a precise diagnostic strategy to detect rare variants and improve drug administration.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111010"},"PeriodicalIF":2.1,"publicationDate":"2025-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145052220","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-03DOI: 10.1016/j.clinbiochem.2025.111008
Fang Wu , Song Lu , Dan Zhang , Nelly Abdelfatah , Vijayananda Kundapur , Fergall Magee , Yanwei Xi
Background
5-Fluorouracil (5-FU) and its pro-drug, capecitabine, are widely used to treat solid tumors. Patients with dihydropyrimidine dehydrogenase (DPYD) deficiency are at increased risk for severe treatment-related toxicity. This study reported the implementation of DPYD genotyping in clinical practice and assessed the impact of genotype-guided dosing on clinical outcomes.
Methods
An in-house pharmacogenomic testing using the Elucigene DPYD genotyping kit (Yourgene Health, UK) was established to detect the four most common clinically actionable DPYD alleles, including *2A, *13, HapB3 and c.2846A>T (rs67376798). Six months post-implementation, a retrospective chart review assessed genotype results, chemotherapy regimens, dose modifications, adverse events related to 5-FU or capecitabine, and demographics. Data were de-identified for analysis.
Results
Analytical validation of the DPYD assay showed 100 % sensitivity, specificity, accuracy, reproducibility, and repeatability. The genotyping workflow was successfully integrated into clinical practice, with a rapid turnaround time to meet oncology treatment planning. From July to December 2024, 299 patients underwent DPYD testing; variants were identified in 22 patients, including 20 patients (6.7 %) with clinically significant variants conferring a reduced DPD function and 2 patients with a variant (c.483 + 18G>A; rs56276561) that retains normal DPD function. Among those variants, HapB3 (n = 18) was the most frequent one, characterized by c.1129-5923C>G and c.1236G>A (rs75017182, rs56038477) co-occurring with c.483 + 18G>A (rs56276561). Of 233 patients receiving 5-FU-based chemotherapy, 13 were variant carriers. Genotype-guided dosing allowed early dose optimization, and all carriers completed at least three treatment cycles, with one severe adverse event attributed to oxaliplatin rather than 5-FU.
Conclusions
This study reported the integration of DPYD pharmacogenomic testing into oncology care and evaluated the post-implementation clinical outcomes, highlighting the critical role of pharmacogenomic testing in optimizing cancer treatment and improving patient safety.
{"title":"Clinical implementation and outcome evaluation of dihydropyrimidine dehydrogenase (DPYD) pharmacogenomic testing for fluoropyrimidine dosing in a Canadian Provincial Healthcare center","authors":"Fang Wu , Song Lu , Dan Zhang , Nelly Abdelfatah , Vijayananda Kundapur , Fergall Magee , Yanwei Xi","doi":"10.1016/j.clinbiochem.2025.111008","DOIUrl":"10.1016/j.clinbiochem.2025.111008","url":null,"abstract":"<div><h3>Background</h3><div>5-Fluorouracil (5-FU) and its pro-drug, capecitabine, are widely used to treat solid tumors. Patients with dihydropyrimidine dehydrogenase (<em>DPYD</em>) deficiency are at increased risk for severe treatment-related toxicity. This study reported the implementation of <em>DPYD</em> genotyping in clinical practice and assessed the impact of genotype-guided dosing on clinical outcomes.</div></div><div><h3>Methods</h3><div>An in-house pharmacogenomic testing using the Elucigene <em>DPYD</em> genotyping kit (Yourgene Health, UK) was established to detect the four most common clinically actionable <em>DPYD</em> alleles, including <em>*2A, *13, HapB3</em> and c.2846A>T (rs67376798). Six months post-implementation, a retrospective chart review assessed genotype results, chemotherapy regimens, dose modifications, adverse events related to 5-FU or capecitabine, and demographics. Data were de-identified for analysis.</div></div><div><h3>Results</h3><div>Analytical validation of the <em>DPYD</em> assay showed 100 % sensitivity, specificity, accuracy, reproducibility, and repeatability. The genotyping workflow was successfully integrated into clinical practice, with a rapid turnaround time to meet oncology treatment planning. From July to December 2024, 299 patients underwent <em>DPYD</em> testing; variants were identified in 22 patients, including 20 patients (6.7 %) with clinically significant variants conferring a reduced DPD function and 2 patients with a variant (c.483 + 18G>A; rs56276561) that retains normal DPD function. Among those variants, <em>HapB3</em> (n = 18) was the most frequent one, characterized by c.1129-5923C>G and c.1236G>A <em>(</em>rs75017182, rs56038477) co-occurring with c.483 + 18G>A (rs56276561). Of 233 patients receiving 5-FU-based chemotherapy, 13 were variant carriers. Genotype-guided dosing allowed early dose optimization, and all carriers completed at least three treatment cycles, with one severe adverse event attributed to oxaliplatin rather than 5-FU.</div></div><div><h3>Conclusions</h3><div>This study reported the integration of <em>DPYD</em> pharmacogenomic testing into oncology care and evaluated the post-implementation clinical outcomes, highlighting the critical role of pharmacogenomic testing in optimizing cancer treatment and improving patient safety.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111008"},"PeriodicalIF":2.1,"publicationDate":"2025-09-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145004771","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Breast cancer (BC) remains a leading cause of morbidity and mortality among women globally. This study aims to investigate HER2 mutations in HER2-negative BC and evaluate the diagnostic potential of matrix metalloproteinases (MMP-7, MMP-9) and CYR61 as serum biomarkers.
Material and methods
The study involved 74 women diagnosed with BC (HER2-positive: n = 33; HER2-negative: n = 33; triple negative: n = 8) and 25 healthy controls. HER2 gene mutations were analyzed using the “AmoyDx HER2 Mutation Detection” kit. Serum MMP-7, MMP-9, and CYR61 levels were quantified using Bio-Techne kits (R&D Systems) using the Quantikine ELISA method.
Results
The study found no mutations (A775_G776insYVMA, M774_A775insAYVM, G776 > VC, G776R, G776C, P780_Y781insGSP, V777L, L755P) in exons 18 and 20 of the HER2 gene in HER2 negative BC patients.
MMP-9 serum levels were significantly reduced in BC patients compared to the control group (p < 0.001). However, MMP-7 levels showed no significant variation when compared to healthy women (p = 0.464). CYR61 exhibited high diagnostic accuracy (AUC = 0.95), supporting its potential as a reliable biomarker.
Conclusion
The reduction in serum MMP-9 levels correlates with ongoing tissue processes, primarily due to increased consumption in the extracellular matrix. CYR61 overexpression is observed across all breast cancer subtypes, independent of HER2 activity, suggesting its significant potential for use in BC diagnosis and treatment.
{"title":"HER2 gene mutations and matrix biomarkers in breast cancer","authors":"Leyla Karimova , Gulnara Azizova , Ilaha Shahverdiyeva","doi":"10.1016/j.clinbiochem.2025.111007","DOIUrl":"10.1016/j.clinbiochem.2025.111007","url":null,"abstract":"<div><h3>Objective</h3><div>Breast cancer (BC) remains a leading cause of morbidity and mortality among women globally. This study aims to investigate <em>HER2</em> mutations in HER2-negative BC and evaluate the diagnostic potential of matrix metalloproteinases (MMP-7, MMP-9) and CYR61 as serum biomarkers.</div></div><div><h3>Material and methods</h3><div>The study involved 74 women diagnosed with BC (HER2-positive: n = 33; HER2-negative: n = 33; triple negative: n = 8) and 25 healthy controls. <em>HER2</em> gene mutations were analyzed using the “AmoyDx HER2 Mutation Detection” kit. Serum MMP-7, MMP-9, and CYR61 levels were quantified using Bio-Techne kits (R&D Systems) using the Quantikine ELISA method.</div></div><div><h3>Results</h3><div>The study found no mutations (A775_G776insYVMA, M774_A775insAYVM, G776 > VC, G776R, G776C, P780_Y781insGSP, V777L, L755P) in exons 18 and 20 of the <em>HER2</em> gene in HER2 negative BC patients.</div><div>MMP-9 serum levels were significantly reduced in BC patients compared to the control group (p < 0.001). However, MMP-7 levels showed no significant variation when compared to healthy women (p = 0.464). CYR61 exhibited high diagnostic accuracy (AUC = 0.95), supporting its potential as a reliable biomarker.</div></div><div><h3>Conclusion</h3><div>The reduction in serum MMP-9 levels correlates with ongoing tissue processes, primarily due to increased consumption in the extracellular matrix. CYR61 overexpression is observed across all breast cancer subtypes, independent of HER2 activity, suggesting its significant potential for use in BC diagnosis and treatment.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111007"},"PeriodicalIF":2.1,"publicationDate":"2025-09-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144997110","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-03DOI: 10.1016/j.clinbiochem.2025.111009
Wenqi Wang , Guangxi Wang , Xiaoming Niu , Zhonglan Li , Mingfu Zhang , Zhenchao Xu , Tou Liu
Objective
This present study aimed to measure platelet factor 4 (PF4) protein level in follicular fluid of patients with polycystic ovary syndrome (PCOS) and analyzed the correlation between follicular PF4 level with clinical characteristics.
Methods
Sixty-seven women (36 PCOS patients vs. 31 non-PCOS women) were enrolled in the study. Follicular fluid PF4 level was analyzed by enzyme-linked immunosorbent assay.
Results
The level of PF4 was significantly higher in follicular fluid of patients with PCOS than that of non-PCOS controls (50.5 (95 % CI: 42.8 to 115.86) ng/ml vs. 37.42 (95 % CI: 23.01 to 49.69) ng/ml, p < 0.001). Correlation analysis showed that the level of PF4 was positively related with serum anti-Mullerian hormone (r = 0.3809, p = 0.0015), serum testosterone (r = 0.3629, p = 0.0025), antral follicle count (r = 0.5544, p < 0.0001), and number of oocytes retrieved (r = 0.3799, p = 0.0018) in all patients. The area under the curve of PF4 level in follicular fluid to predict PCOS was 0.723 (95 % CI: 0.600 to 0.847).
Conclusions
Our study demonstrated that the PF4 level was higher in the follicular fluid of patients with PCOS and was associated with key features of PCOS, suggesting that PF4 may play a role in the pathogenesis of PCOS.
目的测定多囊卵巢综合征（PCOS）患者卵泡液中血小板因子4 （PF4）蛋白水平，并分析PF4蛋白水平与临床特征的相关性。方法67例女性（PCOS患者36例，非PCOS患者31例）纳入研究。采用酶联免疫吸附法分析卵泡液PF4水平。结果PCOS患者卵泡液中PF4水平明显高于非PCOS对照组(50.5 （95% CI: 42.8 ~ 115.86) ng/ml vs. 37.42 (95% CI: 23.01 ~ 49.69) ng/ml, p < 0.001）。相关分析显示，PF4水平与所有患者血清抗苗勒管激素（r = 0.3809, p = 0.0015）、血清睾酮（r = 0.3629, p = 0.0025）、窦卵泡计数（r = 0.5544, p < 0.0001）、取卵数（r = 0.3799, p = 0.0018）呈正相关。卵泡液PF4水平预测PCOS的曲线下面积为0.723 （95% CI: 0.600 ~ 0.847）。结论PCOS患者卵泡液中PF4水平升高，与PCOS的关键特征相关，提示PF4可能在PCOS的发病机制中发挥作用。
{"title":"Elevated level of platelet factor 4 in follicular fluid is associated with polycystic ovary syndrome","authors":"Wenqi Wang , Guangxi Wang , Xiaoming Niu , Zhonglan Li , Mingfu Zhang , Zhenchao Xu , Tou Liu","doi":"10.1016/j.clinbiochem.2025.111009","DOIUrl":"10.1016/j.clinbiochem.2025.111009","url":null,"abstract":"<div><h3>Objective</h3><div>This present study aimed to measure platelet factor 4 (PF4) protein level in follicular fluid of patients with polycystic ovary syndrome (PCOS) and analyzed the correlation between follicular PF4 level with clinical characteristics.</div></div><div><h3>Methods</h3><div>Sixty-seven women (36 PCOS patients vs. 31 non-PCOS women) were enrolled in the study. Follicular fluid PF4 level was analyzed by enzyme-linked immunosorbent assay.</div></div><div><h3>Results</h3><div>The level of PF4 was significantly higher in follicular fluid of patients with PCOS than that of non-PCOS controls (50.5 (95 % CI: 42.8 to 115.86) ng/ml vs. 37.42 (95 % CI: 23.01 to 49.69) ng/ml, <em>p</em> < 0.001). Correlation analysis showed that the level of PF4 was positively related with serum anti-Mullerian hormone (r = 0.3809, <em>p</em> = 0.0015), serum testosterone (r = 0.3629, <em>p</em> = 0.0025), antral follicle count (r = 0.5544, <em>p</em> < 0.0001), and number of oocytes retrieved (r = 0.3799, <em>p</em> = 0.0018) in all patients. The area under the curve of PF4 level in follicular fluid to predict PCOS was 0.723 (95 % CI: 0.600 to 0.847).</div></div><div><h3>Conclusions</h3><div>Our study demonstrated that the PF4 level was higher in the follicular fluid of patients with PCOS and was associated with key features of PCOS, suggesting that PF4 may play a role in the pathogenesis of PCOS.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111009"},"PeriodicalIF":2.1,"publicationDate":"2025-09-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144997109","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-09-02DOI: 10.1016/j.clinbiochem.2025.111004
Amir Karin, Catherine Cheng
Objectives
Macrotroponin refers to circulating immunoglobulin-bound cardiac troponin species that may elevate troponin results in patients with or without myocardial injury, causing diagnostic confusion. Clinical laboratories have been recommended to provide a service for troponin interference investigation. We evaluated the applicability of a Protein A/G IgG-depletion procedure as well as polyethylene glycol (PEG) precipitation for detecting macrotroponin interference with the Siemens Atellica troponin I (TnIH) assay.
Methods
Troponin I, IgG, and albumin (internal standard) were measured (Atellica) on the neat and treated plasma to calculate recovery. Reference samples with TnIH ranging from < 1x to > 1000x times the 99th percentile were selected to verify expected recovery. To minimize likelihood of macrotroponin in the reference group, samples with elevated results were only included if recent acute changes in TnIH was documented. 40 samples were used for the IgG-depletion method and 20 for PEG precipitation. 25 samples from patients with unexplained elevation in TnIH were assessed by IgG-depletion.
Results
38 of 40 reference group recoveries exceeded 70 % (median 91 %, IQR 15 %, max 129 %) in the IgG-depletion group consistent with literature on other assays. Specimens from patients with incongruent clinical picture had IgG-depletion recovery median of 11 % (IQR 14 %, max 37 %). PEG-precipitation showed large variation (median 103 %, IQR 89 %, max 227 %).
Conclusions
IgG depletion using Protein A/G can reliably establish IgG-mediated interference with Atellica TnIH. PEG precipitation results are difficult to interpret likely due to matrix effects, especially at values closer to the 99th percentile.
{"title":"Method choice for investigation of macrotroponin interference with the Siemens Atellica high sensitivity troponin I assay","authors":"Amir Karin, Catherine Cheng","doi":"10.1016/j.clinbiochem.2025.111004","DOIUrl":"10.1016/j.clinbiochem.2025.111004","url":null,"abstract":"<div><h3>Objectives</h3><div>Macrotroponin refers to circulating immunoglobulin-bound cardiac troponin species that may elevate troponin results in patients with or without myocardial injury, causing diagnostic confusion. Clinical laboratories have been recommended to provide a service for troponin interference investigation. We evaluated the applicability of a Protein A/G IgG-depletion procedure as well as polyethylene glycol (PEG) precipitation for detecting macrotroponin interference with the Siemens Atellica troponin I (TnIH) assay.</div></div><div><h3>Methods</h3><div>Troponin I, IgG, and albumin (internal standard) were measured (Atellica) on the neat and treated plasma to calculate recovery. Reference samples with TnIH ranging from < 1x to > 1000x times the 99th percentile were selected to verify expected recovery. To minimize likelihood of macrotroponin in the reference group, samples with elevated results were only included if recent acute changes in TnIH was documented. 40 samples were used for the IgG-depletion method and 20 for PEG precipitation. 25 samples from patients with unexplained elevation in TnIH were assessed by IgG-depletion.</div></div><div><h3>Results</h3><div>38 of 40 reference group recoveries exceeded 70 % (median 91 %, IQR 15 %, max 129 %) in the IgG-depletion group consistent with literature on other assays. Specimens from patients with incongruent clinical picture had IgG-depletion recovery median of 11 % (IQR 14 %, max 37 %). PEG-precipitation showed large variation (median 103 %, IQR 89 %, max 227 %).</div></div><div><h3>Conclusions</h3><div>IgG depletion using Protein A/G can reliably establish IgG-mediated interference with Atellica TnIH. PEG precipitation results are difficult to interpret likely due to matrix effects, especially at values closer to the 99th percentile.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111004"},"PeriodicalIF":2.1,"publicationDate":"2025-09-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144999884","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Prostate-specific antigen (PSA), a serine protease primarily expressed in the prostate, has also been detected in hormonally regulated female tissues, including the breast. Some studies suggest a correlation between increased levels of circulating free PSA (fPSA) and breast cancer, but its role remains debated. This study aimed to evaluate this association while minimizing hormonal confounding factors.
Methods
A total of 82 breast cancer patients (aged 35–86 years) and 31 healthy premenopausal women (aged 18–58 years) were enrolled. Patients had a primary breast cancer diagnosis with no other malignancies and had not undergone preoperative chemotherapy or radiotherapy. Participants with hormonal conditions affecting PSA expression were excluded. fPSA levels were measured using an improved VIDAS® fPSA immunoassay with enhanced analytical sensitivity.
Results
Despite the increased sensitivity of the modified assay, fPSA was undetectable in all plasma samples. This may be due to the exclusion of participants with hormonal imbalances who might exhibit higher PSA expression.
Conclusions
The absence of androgen receptor (AR)-positive triple-negative breast cancer (TNBC) patients in this cohort further supports the role of androgens in PSA regulation. These findings suggest that fPSA may not be a reliable circulating biomarker for breast cancer. However, a key limitation is the lack of fPSA assessment within breast cancer tissue. Future studies should investigate its expression in tumors, particularly in AR-positive TNBC, and evaluate circulating fPSA and testosterone levels as potential biomarkers of tumor androgenic activity.
{"title":"Circulating free PSA in breast cancer patients: is it a reliable biomarker?","authors":"Vanessa Susini , Maria Franzini , Silvia Ursino , Maria Ghilardi , Riccardo Morganti , Cristian Scatena , Irene Bianco , Alessandro Mazzoni , Matteo Ghilli","doi":"10.1016/j.clinbiochem.2025.111006","DOIUrl":"10.1016/j.clinbiochem.2025.111006","url":null,"abstract":"<div><h3>Introduction</h3><div>Prostate-specific antigen (PSA), a serine protease primarily expressed in the prostate, has also been detected in hormonally regulated female tissues, including the breast. Some studies suggest a correlation between increased levels of circulating free PSA (fPSA) and breast cancer, but its role remains debated. This study aimed to evaluate this association while minimizing hormonal confounding factors.</div></div><div><h3>Methods</h3><div>A total of 82 breast cancer patients (aged 35–86 years) and 31 healthy premenopausal women (aged 18–58 years) were enrolled. Patients had a primary breast cancer diagnosis with no other malignancies and had not undergone preoperative chemotherapy or radiotherapy. Participants with hormonal conditions affecting PSA expression were excluded. fPSA levels were measured using an improved VIDAS® fPSA immunoassay with enhanced analytical sensitivity.</div></div><div><h3>Results</h3><div>Despite the increased sensitivity of the modified assay, fPSA was undetectable in all plasma samples. This may be due to the exclusion of participants with hormonal imbalances who might exhibit higher PSA expression.</div></div><div><h3>Conclusions</h3><div>The absence of androgen receptor (AR)-positive triple-negative breast cancer (TNBC) patients in this cohort further supports the role of androgens in PSA regulation. These findings suggest that fPSA may not be a reliable circulating biomarker for breast cancer. However, a key limitation is the lack of fPSA assessment within breast cancer tissue. Future studies should investigate its expression in tumors, particularly in AR-positive TNBC, and evaluate circulating fPSA and testosterone levels as potential biomarkers of tumor androgenic activity.</div></div>","PeriodicalId":10172,"journal":{"name":"Clinical biochemistry","volume":"140 ","pages":"Article 111006"},"PeriodicalIF":2.1,"publicationDate":"2025-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144989191","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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