Study On Chemotherapy Biomarkers Of Multiple Myeloma Based On Drug Exposure And Metabonomics

Objective: Multiple myeloma is a malignant tumor,and chemotherapy is currently one of the main treatment methods in clinical practice,which can bring varying degrees of adverse reactions and therapeutic effects to patients.This article aims to study the exposure concentrations,adverse reactions,and efficacy of drugs such as lenalidomide,cyclophosphamide,bortezomib,and dexamethasone in patients with multiple myeloma through drug metabolism and metabolomics methods.The aim is to identify biomarkers for clinical efficacy in multiple myeloma through metabolomics techniques,and ultimately investigate the metabolic pathways associated with these biomarkers.Methods: 1.Explore a method that can simultaneously and quickly detect thalidomide,lenalidomide,and pomadolide in human plasma samples using liquid chromatography-mass spectrometry,and validate its clinical application.This method uses thalidomide-d4 as the internal standard and uses ZORBAX SB-C18 column(2.1×150 mm,3.5 μm)Separate the chromatographic column using aqueous phase A(0.1%FA aqueous solution)and organic phase B(ACN solution)as mobile phases to complete the gradient elution procedure.The protein precipitation method using methanol as a protein precipitant for plasma sample pretreatment,while referring to the guidance requirements for the validation of quantitative analysis methods for biological samples in the Chinese Pharmacopoeia(2020 edition)for methodological validation;Finally,combined with the relevant compound analysis method established in our laboratory in the early stage,the exposure concentrations of lenalidomide,cyclophosphamide,bortezomib,and dexamethasone in plasma samples of patients with multiple myeloma were detected,and the correlation between the drug’s exposure concentration in vivo and adverse reactions and efficacy was analyzed.2.Non targeted metabolomics research: Using the Agilent 1290 ultra high performance liquid chromatography to detect plasma samples collected from patients and explore the metabolic spectrum differences of compounds in them;Statistical analysis of the above results and identification of compounds were completed through software such as Progenesis QI and EZinfor.Evaluate the efficacy of patients after taking medication,group them according to their quality,and screen for compounds with significant differences;Using component databases such as HMDB and Chem Spider,search and identify endogenous metabolites,and then use the Metabo Analyst 5.0 metabolic pathway analysis database to obtain the metabolic pathways involved in metabolites.3.Targeted metabolomics research: Quantitative analysis of plasma samples collected from patients using the Agilent 6460 A triple quadrupole mass spectrometer.Using SIMCA and Graph Pad Prism 8.0.2 software to process data,analyze changes in the patient’s metabolic spectrum,identify relevant small molecule differential metabolites,and perform correlation analysis between differential metabolites and multiple myeloma;Obtain possible metabolic pathways involved through the Metabo Analyst 5.0 database;Finally,use the selected biomarkers to establish an early warning model and evaluate its diagnostic effectiveness.Results:1.The analytical method established in this study can complete the chromatographic separation and detection of thalidomide,lenalidomide,and pomalidomide within 4 min.The peak shape is good,and the stability of thalidomide,lenalidomide,and pomalidomide is good.The extraction recovery rate is83.90%-119.07%,and the matrix effect is 73.68%-116.75%.The intra day/inter day accuracy and precision,specificity,dilution effect,and residual effect all meet the relevant requirements of biological sample analysis.Based on the method established in the laboratory in the early stage,sample measurements were conducted.The concentrations of lenalidomide were 1.11 ng/mL-143.08 ng/mL,cyclophosphamide was 0.27 ng/mL-608.98 ng/mL,bortezomib was 1.34 ng/mL-61.66 ng/mL,and dexamethasone was 0.97 ng/mL-106.02 ng/mL.The individual differences in drug exposure concentrations in patients were significant,resulting in adverse reactions such as numbness of hands and feet and liver damage.2.Non targeted metabolomics research results: The collected 58 patients with multiple myeloma were classified based on the efficacy after treatment,and two groups were obtained: the CR-VGPR group(36 cases)and the SD-PD group(22 cases).After data analysis,10 differential metabolites were obtained,including alanine,arginine,tyrosine,hippuric acid,proline,methionine,homocysteine,etc.,mainly involving the enrichment of pathways such as cysteine and methionine metabolism,aminoacyl t RNA biosynthesis,arginine biosynthesis,primary bile acid biosynthesis,alanine,aspartic acid,and glutamate metabolism.3.Targeted metabolomics research results: Targeted detection of 32 amino acids was performed on the plasma of 58 patients,and 13 possible biomarkers were obtained,including hippuric acid,glutamic acid,cystine,asymmetric dimethylarginine,aspartic acid,cysteine,L-ornithine ammonia,L-methionine,etc.,mainly involving amino acid t RNA biosynthesis,alanine,aspartic acid and glutamate metabolism,glycine,glutamate metabolism,etc.Enrichment of pathways such as serine and threonine metabolism,arginine biosynthesis,and histidine metabolism.Using logistic regression modeling and ROC curve for the diagnosis of multiple myeloma,the AUC of the regression model is 0.9319,which has certain diagnostic value for it;Through the ROC curve,it was found that L-pyroglutamic acid(AUC=0.6750,P=0.0312)and glycine(AUC=0.6819,P=0.0251),with P<0.05,have certain predictive value.Conclusion: This article establishes a mass spectrometry method for rapid determination of three immune modulators,combined with previous laboratory research,to determine the correlation between drug exposure concentration,adverse reactions,and efficacy in patients with multiple myeloma.Drug exposure concentration can diagnose the occurrence of adverse reactions,but cannot distinguish the quality of efficacy.Therefore,non targeted metabolomics was used to screen for metabolic foreign bodies,and alanine,arginine,tyrosine,hippuric acid,proline,L-cysteine,methionine,homocysteine,glutamic acid,and lysine were discovered.At the same time,metabolic pathway enrichment analysis mainly involves the metabolism and synthesis of various amino acids;Through targeted amino acid metabolomics research,it was found that there is a metabolic spectrum disorder of amino acid content differences in plasma samples,and the results of the two metabolomics studies are almost consistent.We established a ROC curve evaluation model for multiple myeloma and amino acid monofactor,and found that L-pyroglutamine and glycine were the main endogenous biomarkers with good diagnostic value.