Molecular Biotechnology最新文献

{"title":"Exploring the Targets and Molecular Mechanisms of Curcumin for the Treatment of Bladder Cancer Based on Network Pharmacology, Molecular Docking and Molecular Dynamics.","authors":"Jun Li, Shujie Feng, Xiong Wang, Bingmei Zhang, Qingmin He","doi":"10.1007/s12033-024-01190-x","DOIUrl":"10.1007/s12033-024-01190-x","url":null,"abstract":"<p><p>Curcumin, a phenolic compound derived from turmeric, has demonstrated anti-tumor properties in preclinical models of various cancers. However, the exact mechanism of curcumin in treating bladder cancer remains unclear. This study aimed to elucidate the therapeutic targets and molecular mechanisms of curcumin in the treatment of BC through an integrated approach of network pharmacology, molecular docking, and molecular dynamics simulations. PharmMapper, SuperPred, TargetNet, and SwissTargetPrediction were utilized to acquire targets associated with curcumin, while GeneCards, CTD, DisGeNET, OMIM, and PharmGKB databases were utilized to obtain targets related to bladder cancer. The drug-disease interaction targets were obtained using Venny 2.1.0, and GO and KEGG enrichment analyses were then conducted with the DAVID tool. We constructed a protein-protein interaction (PPI) network and identified tenkey targets. In conclusion, AutoDock Tools 1.5.7 was utilized to conduct molecular docking simulations, followed by additional analysis of the central targets through the GEPIA, HPA, cBioPortal, and TIMER databases. A total of 305 potential anticancer targets of curcumin were obtained. The analysis of GO functional enrichment resulted in a total of 1105 terms, including 786 terms related to biological processes (BP), 105 terms related to cellular components (CC), and 214 terms related to molecular functions (MF). In addition, KEGG pathway enrichment analysis identified 170 relevant signaling pathways. Treating bladder cancer could potentially involve inhibiting pathways like the PI3K-Akt signaling pathway, MAPK signaling pathway, EGFR tyrosine kinase inhibitor resistance, and IL-17 signaling pathway. Activating TNF, ALB, CASP3, and ESR1 while inhibiting AKT1, EGFR, STAT3, BCL2, SRC, and HSP90AA1 can also hinder the proliferation of bladder tumor cells. According to the results of molecular docking, curcumin binds to these central targets in a spontaneous manner, exhibiting binding energies lower than - 1.631 kJ/mol. These findings were further validated at the transcriptional, translational and immune infiltration levels. By utilizing network pharmacology and molecular docking techniques, it was discovered that curcumin possesses diverse effects on multiple targets and pathways for treating bladder cancer. It has the potential to impede the growth of bladder tumor cells by suppressing various pathways including the PI3K-Akt and MAPK signaling pathways, as well as pathways associated with EGFR tyrosine kinase inhibitor resistance and the IL-17 signaling pathway. Curcumin could potentially disrupt the cell cycle advancement in bladder cancer cells by increasing the expression of TNF, ALB, CASP3, and ESR1 while decreasing AKT1, EGFR, STAT3, BCL2, SRC, HSP90AA1, and other targeted genes. These findings reveal the possible molecular pathways through which curcumin exerts its anticancer effects in bladder cancer, and this novel research strategy not only p","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"2138-2159"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141186786","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Increased Immune Infiltration and Improved Prognosis of Head and Neck Squamous Cell Carcinoma Associated with Reduced Ancient Ubiquitous Protein 1 Gene Expression.","authors":"Yi Wang, Qian Wu, Xiao Wei, Gang Huang, Guangyong Feng, Hui Xu, Xiaoxia Gou","doi":"10.1007/s12033-024-01161-2","DOIUrl":"10.1007/s12033-024-01161-2","url":null,"abstract":"<p><p>This study aimed to explore the molecular mechanism underlying the prognostic role of ancient ubiquitous protein 1 (AUP1) in head and neck squamous cell carcinoma (HNSCC) and its relationship with the tumor immune microenvironment. Various web resources were used to analyze the differential expression of AUP1 and its role in the HNSCC pathogenesis. A nomogram aimed at predicting 1-, 3-, and 5-year survival rates was developed based on the patient's clinicopathological characteristics and AUP1 expression pattern. Several algorithms and analytical tools were used to explore the correlation between AUP1 expression and sensitivity to immune checkpoint gene therapy by evaluating infiltrating immune cells in patients with HNSCC. Higher AUP1 mRNA and protein expression levels were observed in most tumors and HNSCC than in the normal tissues. High AUP1 expression was an independent predictive risk factor for the overall survival of patients as it was closely associated with the patients' T, M, clinical, and pathological stages and lymphovascular invasion in HNSCC. In conclusion, AUP1 is involved in the occurrence and progression of HNSCC, may be used as an independent prognostic factor in patients with HNSCC, and could serve as a potential intervention target to improve immunotherapy sensitivity in HNSCC.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"1826-1842"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141306335","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Marine-Derived Cytosine Arabinoside (Ara-C) Inhibits Biofilm Formation by Inhibiting PEL Operon Proteins (Pel A and Pel B) of Pseudomonas aeruginosa: An In Silico Approach.","authors":"Susmita Datta, Vishal Singh, Soma Nag, Dijendra Nath Roy","doi":"10.1007/s12033-024-01169-8","DOIUrl":"10.1007/s12033-024-01169-8","url":null,"abstract":"<p><p>Pseudomonas aeruginosa (P. aeruginosa) is a gram-negative biofilm-forming opportunistic human pathogen whose vital mechanism is biofilm formation for better survival. PelA and PelB proteins of the PEL operon are essential for bacterial-synthesized pellicle polysaccharide (PEL), which is a vital structural component of the biofilm. It helps in adherence of biofilm on the surface and maintenance of cell-to-cell interactions and with other matrix components. Here, in-silico molecular docking and simulation studies were performed against PelA and PelB using ten natural bioactive compounds, individually [podocarpic acids, ferruginol, scopadulcic acid B, pisiferic acid, metachromin A, Cytarabine (cytosine arabinoside; Ara-C), ursolic acid, oleanolic acid, maslinic acid, and betulinic acid], those have already been established as anti-infectious compounds. The results obtained from AutoDock and Glide-Schordinger stated that a marine-derived cytosine arabinoside (Ara-C) among the ten compounds binds active sites of PelA and PelB, exhibiting strong binding affinity [Trp224 (hydrogen), Ser219 (polar), Val234 (hydrophobic) for PelA; Leu365 and Glu389 (hydrogen), Gln366 (polar) for PelB] with high negative binding energy - 5.518 kcal/mol and - 6.056 kcal/mol, respectively. The molecular dynamic and simulation studies for 100 ns showed the MMGBSA binding energy scores are - 16.4 kcal/mol (Ara-C with PelA), and - 22.25 kcal/mol (Ara-C with PelB). Further, ADME/T studies indicate the IC₅₀ values of AraC are 6.10 mM for PelA and 18.78 mM for PelB, which is a comparatively very low dose. The zero violation of Lipinski's Rule of Five further established that Ara-C is a good candidate for drug development. Thus, Ara-C could be considered a potent anti-biofilm compound against PEL operon-dependent biofilm formation of P. aeruginosa.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"1924-1938"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140912566","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"METTL3 Promotes OSCC Progression by Down-Regulating WEE1 in a m6A-YTHDF2-Dependent Manner.","authors":"Yongxu Su, Yanjia Hu, Binbin Qu, Rongchang Lei, Ge Guo","doi":"10.1007/s12033-024-01165-y","DOIUrl":"10.1007/s12033-024-01165-y","url":null,"abstract":"<p><p>Oral squamous cell carcinoma (OSCC) is a common and highly lethal epithelial cancer. This study aimed to confirm the role of METTL3 in promoting OSCC and investigate its specific underlying mechanisms. Expression of the METTL3, YTH domain-containing family 2 (YTHDF2), and WEE1 were examined in normal oral epithelial cells and OSCC cells. Cell functions were examined after overexpressing WEE1 in OSCC cells. MeRIP-qPCR analysis was used to detect WEE1 m6A levels in HOK, SCC25, and CAL27 cells. WEE1 and its m6A levels were evaluated in OSCC cells by knocking down METTL3/YTHDF2, assessing the interaction between METTL3/YTHDF2 and WEE1. The impact of METTL3 and YTHDF2 downregulation on WEE1 mRNA stability was also investigated. The tumor weight and volume in a nude mouse model of OSCC after overexpression of WEE1 and YTHDF2 were measured. Expression of Ki-67 and WEE1 in OSCC tissue was detected using immunohistochemistry. Compared to normal oral epithelial cells, METTL3 and YTHDF2 were upregulated in OSCC cells, while WEE1 was downregulated, and there was a negative correlation between WEE1 and METTL3/YTHDF2 expression. WEE1 overexpression inhibited proliferation, invasion, and migration while promoting apoptosis in OSCC cells. METTL3 and YTHDF2 bound to WEE1 mRNA. METTL3/YTHDF2 knockdown increased WEE1 levels and WEE1 mRNA stability. METTL3 inhibition reduced WEE1 m6A levels. Inhibition of METTL3 weakened the interaction between YTHDF2 and WEE1 mRNA. In vivo, overexpression of WEE1 suppressed OSCC development, which was reversed by overexpression of YTHDF2. METTL3 facilitates the progression of OSCC through m6A-YTHDF2-dependent downregulation of WEE1.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"1867-1879"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140922476","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Nucleoporin 93 Regulates Cancer Cell Growth and Stemness in Bladder Cancer via Wnt/β-Catenin Signaling.","authors":"Zhe Wang, Jing Zhang, Lina Luo, Chao Zhang, Xiaomeng Huang, Shuo Liu, Huaian Chen, Wenlong Miao","doi":"10.1007/s12033-024-01184-9","DOIUrl":"10.1007/s12033-024-01184-9","url":null,"abstract":"<p><p>Bladder cancer (BLCA) is a prevalent cancer type with an unmet need for new therapeutic strategies. Nucleoporin 93 (Nup93) is implicated in the pathophysiology of several cancers, but its relationship with bladder cancer remains unclear. Nup93 expression was analyzed in TCGA datasets and 88 BLCA patient samples. Survival analysis and Cox regression models evaluated the association between Nup93 levels and patient prognosis. BLCA cells were used to investigate the effects of Nup93 overexpression or knockdown on cell growth, invasion, stemness (sphere formation and ALDH2 + cancer stem cell marker), and Wnt/β-catenin signaling in vitro. The Wnt activator BML-284 was used to confirm the involvement of Wnt/β-catenin signaling pathway. A xenograft mouse model validated the in vitro findings. Nup93 was highly expressed in BLCA tissues and cell lines, and high Nup93 expression correlated with poor prognosis in BLCA patients. Nup93 silencing inhibited BLCA cell proliferation, Wnt/β-catenin activation, and cancer cell stemness. Conversely, Nup93 overexpression promoted these effects. BML-284 partially rescued the reduction in cell growth and stemness markers caused by Nup93 knockdown. Nup93 knockdown also suppressed the tumor formation of BLCA cells in vivo. Nup93 regulates BLCA cell growth and stemness via the Wnt/β-catenin pathway, suggesting its potential as a prognostic marker and therapeutic target in BLCA.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"2072-2084"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140922478","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A Novel Neutrophil Extracellular Trap Signature Predicts Patient Chemotherapy Resistance and Prognosis in Lung Adenocarcinoma.","authors":"Long Xing, Shuangli Wu, Shiyue Xue, Xingya Li","doi":"10.1007/s12033-024-01170-1","DOIUrl":"10.1007/s12033-024-01170-1","url":null,"abstract":"<p><p>Chemoresistance is a key obstacle in the long-term survival of patients with locally and advanced lung adenocarcinoma (LUAD). This study used bioinformatic analysis to reveal the chemoresistance of gene-neutrophil extracellular traps (NETs) associated with LUAD. RNA sequencing data and LUAD expression patterns were obtained from the Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases, respectively. The GeneCards database was used to identify NETosis-related genes (NRGs). To identify hub genes with significant and consistent expression, differential analysis was performed using the TCGA-LUAD and GEO datasets. LUAD subtypes were determined based on these hub genes, followed by prognostic analysis. Immunological scoring and infiltration analysis were conducted using NETosis scores (N-scores) derived from the TCGA-LUAD dataset. A clinical prognostic model was established and analyzed, and its clinical applications explored. Twenty-two hub genes were identified, and consensus clustering was used to identify two subgroups based on their expression levels. The Kaplan-Meier (KM) curves demonstrated statistically significant differences in prognosis between the two LUAD subtypes. Based on the median score, patients were further divided into high and low N-score groups, and KM curves showed that the N-scores were more precise at predicting the prognosis of patients with LUAD for overall survival (OS). Immunological infiltration analysis revealed significant differences in the abundances of 10 immune cell infiltrates between the high and low N-score groups. Risk scores indicated significant differences in prognosis between the two extreme score groups. The risk scores for the prognostic model also indicated significant differences between the two groups. The results provide new insights into NETosis-related differentially expressed genes (NRDEGs) associated with chemotherapy resistance in patients with LUAD. The established prognostic model is promising and could help with clinical applications to evaluate patient survival and therapeutic efficiency.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"1939-1957"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140907612","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CEBPA Restrains the Malignant Progression of Breast Cancer by Prompting the Transcription of SOCS2.","authors":"Jin-Li Wang, Wei-Wei Ji, Ao-Li Huang, Zhen Liu, Deng-Feng Chen","doi":"10.1007/s12033-024-01189-4","DOIUrl":"10.1007/s12033-024-01189-4","url":null,"abstract":"<p><p>The suppressor of cytokine signaling 2 (SOCS2) has been identified to act as a tumor suppressor in breast cancer (BC) progression. However, the action of SOCS2 in macrophage polarization in BC cells has not been reported yet. The qRT-PCR and western blotting were adopted for detecting the levels of mRNAs and proteins. The macrophage M2 polarization was analyzed by flow cytometry. Analyses of cell oncogenic phenotypes and tumor growth were conducted using 5-ethynyl-2'-deoxyuridine (EdU), flow cytometry, scratch, Transwell, tube formation assays in vitro, and tumor xenograft assay in vivo, respectively. The interaction between CEBPA (CCAAT Enhancer Binding Protein Alpha) and SOCS2 was confirmed using bioinformatics analysis and dual-luciferase reporter assay. SOCS2 was lowly expressed in BC tissues and cells. Functionally, overexpression of SOCS2 inhibited macrophage M2 polarization, and impaired BC cell proliferation, angiogenesis, and metastasis. Mechanistically, CEBPA bound to the promoter region of SOCS2, and promoted its transcription. A low CEBPA expression was observed in BC tissues and cells. Forced expression of CEBPA also suppressed macrophage M2 polarization, BC cell proliferation, angiogenesis, and metastasis. Moreover, the anticancer effects mediated by CEBPA were abolished by SOCS2 knockdown. In addition, CEBPA overexpression impeded BC growth in nude mice by regulating SOCS2. CEBPA suppressed macrophage M2 polarization, BC cell proliferation, angiogenesis, and metastasis by promoting SOCS2 transcription in a targeted manner.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"2127-2137"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141076209","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Strategies of Bladder Reconstruction after Partial or Radical Cystectomy for Bladder Cancer.","authors":"Xiao Xue Zeng, Yuyan Wu","doi":"10.1007/s12033-024-01163-0","DOIUrl":"10.1007/s12033-024-01163-0","url":null,"abstract":"<p><p>The standard strategy is to reconstruct bladder by use of bowel segments as material in bladder cancer with radical cystectomy clinically. Both natural derived and non natural derived materials are investigated in bladder reconstruction. Studies on mechanical bladder, bladder transplantation and bladder xenotransplantation are currently limited although heart and kidney transplantation or xenotransplantation are successful to a certain extent, and bone prostheses are applied in clinical contexts. Earlier limited number of studies associated with bladder xenograft from animals to humans were not particular promising in results. Although there have been investigations on pig to human cardiac xenotransplantation with CRISPR Cas9 gene editing, the CRISPR Cas technique is not yet widely researched in porcine bladder related gene editing for the potential of human bladder replacement for bladder cancer. The advancement of technologies such as gene editing, bioprinting and induced pluripotent stem cells allow further research into partial or whole bladder replacement strategies. Porcine bladder is suggested as a potential source material for bladder reconstruction due to its alikeness to human bladder. Challenges that exist with all these approaches need to be overcome. This paper aims to review gene editing technology such as the CRISPR Cas systems as tools in bladder reconstruction, bladder xenotransplantation and hybrid bladder with technologies of induced pluripotent stem cells and genome editing, bioprinting for bladder replacement for bladder reconstruction and to restore normal bladder control function after cystectomy for bladder cancer.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"1735-1751"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140958318","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Guardians Turned Culprits: NETosis and Its Influence on Pulmonary Fibrosis Development.","authors":"Aleena Varughese, Akarsha Balnadupete, Poornima Ramesh, Thottethodi Subrahmanya Keshava Prasad, Ayshath Burhana Nidha, Yashodhar Bhandary","doi":"10.1007/s12033-024-01171-0","DOIUrl":"10.1007/s12033-024-01171-0","url":null,"abstract":"<p><p>Idiopathic pulmonary fibrosis (IPF) is a debilitating, life-threatening irreversible lung disease characterized by the excessive accumulation of fibrotic tissue in the lungs, impairing their function. The exact mechanisms underlying Pulmonary fibrosis (PF) are multifaceted and not yet fully understood. Reports show that during COVID-19 pandemic, PF was dramatically increased due to the hyperactivation of the immune system. Neutrophils and macrophages are the patrolling immune cells that keep the microenvironment balanced. Neutrophil extracellular traps (NETs) are a normal protective mechanism of neutrophils. The chief components of the NETs include DNA, citrullinated histones, and anti-microbial peptides which are released by the activated neutrophils. However, it is becoming increasingly evident that hyperactivation of immune cells can also turn into criminals when it comes to pathological state. Dysregulated NETosis may contribute to sustained inflammation, overactivation of fibroblasts, and ultimately promoting collagen deposition which is the characteristic feature of PF. The role of NETs along with inflammation is attaining greater attention. However, seldom researches are related to the relationship between NETs causing PF. This review highlights the cellular mechanism of NETs-induced pulmonary fibrosis, which could give a better understanding of molecular targets which may be helpful for treating NETs-induced PF.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"1752-1764"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140876840","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Design and Production of a Novel Anti-PD-1 Nanobody by CDR Grafting and Site-Directed Mutagenesis Approach.","authors":"Mahsa Mirzaei, Shima Mirhoseini, Mohammad Mehdi Heidari, Mehri Khatami","doi":"10.1007/s12033-024-01162-1","DOIUrl":"10.1007/s12033-024-01162-1","url":null,"abstract":"<p><p>Programmed cell death protein-1 (PD-1) is a membrane protein expressed on the surface of activated T-cells, B-cells, natural killer cells, dendritic cells, macrophages, and monocytes. Inhibition of the PD-1/PD-L1 interaction by monoclonal antibodies (mAbs) has many therapeutic benefits and has led to a major advance in the treatment of various types of tumors. Due to the large size and immunogenicity of the antibodies (Abs), using small molecules such as nanobodies (nanobodies or VHH) is more appropriate for this purpose. In this research, the complementarity determining regions (CDR) grafting method was used to produce anti-PD-1 nanobody. For producing the grafted anti-PD-1 nanobody, CDRs from the tislelizumab mAb were grafted into the frameworks of a nanobody whose sequence is similar to the tislelizumab mAb. Also, the site-directed mutagenesis method was used to produce two mutated anti-PD-1 nanobodies which increased the affinity of grafted anti-PD-1 nanobodies. Two amino acid substitutions (Tyr97Arg and Tyr102Arg) in the VHH-CDR3 were used to improve grafted nanobody affinity and the binding capacity of the mutated nanobodies. The binding of the anti-PD-1 nanobodies and PD-1 antigen (Ag) was confirmed by Dot blot, western blot, and indirect ELISA analysis. According to the results of these in silico and in vitro studies, the binding between grafted and mutated nanobodies with PD-1 was confirmed. Also, our findings show that site-directed mutagenesis can increase the affinity of nanobodies.</p>","PeriodicalId":18865,"journal":{"name":"Molecular Biotechnology","volume":" ","pages":"1843-1851"},"PeriodicalIF":2.4,"publicationDate":"2025-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140912564","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}