ABSTRACT
The development of drug resistance in malignant tumors leads to disease progression, creating a bottleneck in treatment. Bevacizumab is widely used clinically, and acts by inhibiting angiogenesis to "starve" tumors. Continuous treatment can readily induce rebound proliferation of tumor blood vessels, leading to drug resistance. Previously, we found that the fragment crystallizable (Fc) region of bevacizumab cooperates with the Toll-like receptor-4 (TLR4) ligand to induce M2b polarization in macrophages and secrete tumor necrosis factor-α (TNFα), which promotes immunosuppression, tumor metastasis, and angiogenesis. However, the downstream mechanism underlying TNFα-mediated bevacizumab resistance requires further investigation. Our RNA-Seq analysis results revealed that the expression of endothelial cell specific molecule-1 (ESM1) increased significantly in drug-resistant tumors and promoted metastasis and angiogenesis in vitro and in vivo. Furthermore, TNFα induced the upregulation of ESM1, which promotes metastasis and angiogenesis and regulates matrix metalloprotease-9 (MMP9), vascular endothelial growth factor (VEGF), and delta-like ligand-4 molecules (DLL4). Accordingly, the curative effect of bevacizumab improved by neutralizing ESM1 with high-affinity anti-ESM1 monoclonal antibody 1-2B7 in bevacizumab-resistant mice. This study provides important insights regarding the molecular mechanism by which TNFα-induced ESM1 expression promotes angiogenesis, which is significant for elucidating the mechanism of bevacizumab drug resistance and possibly identifying appropriate biosimilar molecules.
ABSTRACT
Adoptive immunotherapies based on the transfer of functional immune cells hold great promise in treating a wide range of malignant diseases, especially cancers, autoimmune diseases, and infectious diseases. However, manufacturing issues and biological barriers lead to the insufficient population of target-selective effector cells at diseased sites after adoptive transfer, hindering effective clinical translation. The convergence of immunology, cellular biology, and materials science lays a foundation for developing biomaterial-based engineering platforms to overcome these challenges. Biomaterials can be rationally designed to improve ex vivo immune cell expansion, expedite functional engineering, facilitate protective delivery of immune cells in situ, and navigate the infused cells in vivo. Herein, this review presents a comprehensive summary of the latest progress in biomaterial-based strategies to enhance the efficacy of adoptive cell therapy, focusing on function-specific biomaterial design, and also discusses the challenges and prospects of this field.
Subject(s)
Biocompatible Materials , Neoplasms , Cell- and Tissue-Based Therapy , Humans , Immunotherapy, Adoptive , Neoplasms/therapyABSTRACT
Bevacizumab in neoadjuvant therapy provides a new hope of improved survival for patients with triple-negative breast cancer (TNBC) by targeting vascular endothelial growth factor in combination with chemotherapy, but curative effect is limited by bevacizumab's continuous use while mechanisms remain incompletely understood. More and more researches reported that tumor-associated macrophages mediate resistance to chemotherapy and radiotherapy in various tumors. Here we developed a TNBC model resistant to bevacizumab under bevacizumab continuous administration. It was found that proportion of a specific subset of tumor-associated macrophages characterized as M2b (CD11b+ CD86high IL10high) increased and responsible for acquired resistance to bevacizumab. Then, we showed that RAW264.7 macrophages could be polarized to M2b subtype on simultaneous exposure to bevacizumab and TLR4 ligands as occurs in the context of continuous bevacizumab treatment. Concordantly, in TLR4-deleted C57BL/10ScNJNju (TLR4lps-del) mut/mut mice with bevacizumab treatment model, it was verified that the M2b macrophage could be induced by Fc gamma receptor-TLR4 cross-talk. In MDA-MB-231-resistant tumor-bearing mice, the content of TNFα in serum kept going up consistent with CCL1, a chemokine of M2b macrophage. In vitro neutralizing tumor necrosis factor α (TNFα) could inhibit the tumor progression caused by M2b culture medium and tumor IDO1 expression. Therefore, we thought that TNFα is a key tumor-promoting effector molecule secreted by M2b macrophage. Accordingly, the curative effect of bevacizumab was proved to be significantly improved by neutralizing TNFα with anti-TNFα nanobody. This study is expected to provide theoretical and clinical evidence elucidating the drug resistance in patients receiving bevacizumab.
Subject(s)
Antineoplastic Agents, Immunological/therapeutic use , Bevacizumab/therapeutic use , Triple Negative Breast Neoplasms/drug therapy , Tumor Necrosis Factor-alpha/metabolism , Animals , Antineoplastic Agents, Immunological/pharmacology , Bevacizumab/pharmacology , Female , Humans , MiceABSTRACT
BACKGROUND: Genome-wide association studies (GWAS) have proven successful in predicting genetic risk of disease using single-locus models; however, identifying single nucleotide polymorphism (SNP) interactions at the genome-wide scale is limited due to computational and statistical challenges. We addressed the computational burden encountered when detecting SNP interactions for survival analysis, such as age of disease-onset. To confront this problem, we developed a novel algorithm, called the Efficient Survival Multifactor Dimensionality Reduction (ES-MDR) method, which used Martingale Residuals as the outcome parameter to estimate survival outcomes, and implemented the Quantitative Multifactor Dimensionality Reduction method to identify significant interactions associated with age of disease-onset. METHODS: To demonstrate efficacy, we evaluated this method on two simulation data sets to estimate the type I error rate and power. Simulations showed that ES-MDR identified interactions using less computational workload and allowed for adjustment of covariates. We applied ES-MDR on the OncoArray-TRICL Consortium data with 14,935 cases and 12,787 controls for lung cancer (SNPs = 108,254) to search over all two-way interactions to identify genetic interactions associated with lung cancer age-of-onset. We tested the best model in an independent data set from the OncoArray-TRICL data. RESULTS: Our experiment on the OncoArray-TRICL data identified many one-way and two-way models with a single-base deletion in the noncoding region of BRCA1 (HR 1.24, P = 3.15 × 10-15), as the top marker to predict age of lung cancer onset. CONCLUSIONS: From the results of our extensive simulations and analysis of a large GWAS study, we demonstrated that our method is an efficient algorithm that identified genetic interactions to include in our models to predict survival outcomes.
Subject(s)
Algorithms , Biomarkers, Tumor/genetics , Computational Biology/methods , Gene Expression Regulation, Neoplastic , Genome-Wide Association Study , Lung Neoplasms/mortality , Polymorphism, Single Nucleotide , Adolescent , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/metabolism , Case-Control Studies , Female , Genetic Predisposition to Disease , Genotype , Humans , Lung Neoplasms/genetics , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Male , Middle Aged , Multifactor Dimensionality Reduction , Prognosis , Survival Rate , Young AdultABSTRACT
AIMS: Triple-negative breast cancer (TNBC) is not sensitive to current endocrine treatments, so new treatment strategies need to be explored. Based on previous antitumour studies on anti-TNFα nanobody, we designed a novel fusion nanobody to enhance antitumour activity of the anti-TNFα nanobody in TNBC. MAIN METHODS: The RGD4C contains RGD sequence, which is the smallest recognition unit binding to the αvß3 receptor on tumour cell membranes and involved in tumour cell adhesion, proliferation, and metastasis. RGD4C was fused to anti-TNFα nanobody to investigate the antitumour activity in vitro and in vivo. KEY FINDINGS: The antitumour effects of fusion nanobody V-L-R-H could effectively bind to αvß3 and inhibit cell migration and proliferation of MDA-MB-231, which had satisfying purification efficiency and approving antigen or receptor binding activity. V-L-R-H could inhibit the TNFα-mediated PI3K/AKT/NF-κB signal pathway and integrin αvß3 correlative FAK focal adhesion signal pathway. Mouse xenograft tumour experiments showed that the V-L-R-H could inhibit tumour proliferation and metastasis; reduce the TNFα, HIFα, Ki67, and CD31 concentrations in tumour; and inhibit the process of epithelial-mesenchymal transition. SIGNIFICANCE: The fusion nanobody enhanced antitumour activity of the anti-TNFα nanobody on TNBC. It provided a reference for the design of dual functional fusion proteins and development of tumour treatment strategies of antagonistic TNFα and αvß3, and a new therapeutic strategy and research direction for the treatment of TNBC.
Subject(s)
Antineoplastic Agents, Immunological/therapeutic use , Oligopeptides/metabolism , Recombinant Fusion Proteins/therapeutic use , Single-Domain Antibodies/therapeutic use , Triple Negative Breast Neoplasms/drug therapy , Tumor Necrosis Factor-alpha/immunology , Animals , Cell Line , Cell Line, Tumor , Cell Proliferation/drug effects , Female , Fibroblasts/drug effects , Gene Expression , Humans , Integrin alphaVbeta3/metabolism , Mice , Mice, Inbred BALB C , Neoplasm Metastasis/prevention & control , Oligopeptides/chemistry , Oligopeptides/genetics , Pichia/metabolism , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/chemistry , Single-Domain Antibodies/genetics , Triple Negative Breast Neoplasms/pathology , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Tumor Necrosis Factor-alpha/pharmacology , Xenograft Model Antitumor AssaysABSTRACT
Few germline mutations are known to affect lung cancer risk. We performed analyses of rare variants from 39,146 individuals of European ancestry and investigated gene expression levels in 7,773 samples. We find a large-effect association with an ATM L2307F (rs56009889) mutation in adenocarcinoma for discovery (adjusted Odds Ratio = 8.82, P = 1.18 × 10-15) and replication (adjusted OR = 2.93, P = 2.22 × 10-3) that is more pronounced in females (adjusted OR = 6.81 and 3.19 and for discovery and replication). We observe an excess loss of heterozygosity in lung tumors among ATM L2307F allele carriers. L2307F is more frequent (4%) among Ashkenazi Jewish populations. We also observe an association in discovery (adjusted OR = 2.61, P = 7.98 × 10-22) and replication datasets (adjusted OR = 1.55, P = 0.06) with a loss-of-function mutation, Q4X (rs150665432) of an uncharacterized gene, KIAA0930. Our findings implicate germline genetic variants in ATM with lung cancer susceptibility and suggest KIAA0930 as a novel candidate gene for lung cancer risk.
Subject(s)
Adenocarcinoma/genetics , Ataxia Telangiectasia Mutated Proteins/genetics , Lung Neoplasms/genetics , Aged , Alleles , Databases, Genetic , Female , Genetic Predisposition to Disease , Genotyping Techniques , Germ-Line Mutation , Heterozygote , Humans , Jews/genetics , Male , Middle Aged , Mutation, Missense , Odds Ratio , Oligonucleotide Array Sequence Analysis , Pedigree , RNA-Seq , Risk Factors , White People/geneticsABSTRACT
Colorectal cancer (CRC) is a type of malignant cancer that has become particularly prevalent worldwide. It is of crucial importance to CRC treatment that the underlying molecular mechanism of CRC progression is determined. The NRAS gene is an important small G protein that is involved in various biological processes, including cancers. NRAS is an oncogene in many neoplasms but its function and regulation in CRC have seldom been investigated. In this study, it was uncovered that the NRAS protein was significantly upregulated in CRC tissues. According to a bioinformatics prediction, we identified that miR-144 may target NRAS to suppress its expression. In vitro experiments indicated that miR-144 decreased NRAS expression in different CRC cell lines (SW480, LoVo, and Caco2). By inhibiting NRAS, miR-144 repress SW480 cell proliferation and migration. Moreover, miR-144 decelerated the growth of SW480 xenograft tumors in vivo by targeting NRAS. In summary, our results identified a novel miR-144-NRAS axis in CRC that could promote the research and treatment of CRC.
ABSTRACT
In 2009, the FDA approved bevacizumab for the treatment of adult patients diagnosed with recurrent glioblastoma. However, the poor permeability of the macromolecules across the blood-brain barrier, determined by multifactorial anatomical and physiological milieu, restricts the clinical therapeutic effect of bevacizumab. The low-density lipoprotein receptor related protein 1 (LRP1) is highly expressed in the endothelial cells of the brain capillary and the glioma cells. Angiopep-2 (ANG) is a 19-aa oligopeptide that can bind to LRP1 and penetrate the blood-brain barrier by receptor-mediated transport. Therefore, ANG can be used as a dual-targeting drug delivery carrier into the brain and the glioma sites. In this study, ANG gene was fused with the C-terminal domain of single-chain antigen binding fragment (scFab) of the anti-VEGF antibody and recombinant scFab-ANG protein was expressed and purified using Rosatte (DE3) strain. We confirmed that ANG could carry anti-VEGF-scFab, penetrate a three-dimensional model of the brain tumor, and cross the hCMEC/D3 monolayer in the in vitro blood-brain barrier model. The animal experiments demonstrated that 3 h after the tail intravenous protein injection, the fluorescent signals in the brains of the mice in the scFab-ANG group were stronger than that in the scFab group. Furthermore, the study of the in situ rat glioma model shows that scFab-ANG could target glioma while anti-VEGF-scFab could not. These findings indicate that scFab-ANG had stronger transepithelial permeability and glioma targeting capacity. Thus, it can be a potential candidate drug for glioblastoma therapy.
ABSTRACT
INTRODUCTION: Inherited susceptibility to lung cancer risk in never-smokers is poorly understood. The major reason for this gap in knowledge is that this disease is relatively uncommon (except in Asians), making it difficult to assemble an adequate study sample. In this study we conducted a genome-wide association study on the largest, to date, set of European-descent never-smokers with lung cancer. METHODS: We conducted a two-phase (discovery and replication) genome-wide association study in never-smokers of European descent. We further augmented the sample by performing a meta-analysis with never-smokers from the recent OncoArray study, which resulted in a total of 3636 cases and 6295 controls. We also compare our findings with those in smokers with lung cancer. RESULTS: We detected three genome-wide statistically significant single nucleotide polymorphisms rs31490 (odds ratio [OR]: 0.769, 95% confidence interval [CI]: 0.722-0.820; p value 5.31 × 10-16), rs380286 (OR: 0.770, 95% CI: 0.723-0.820; p value 4.32 × 10-16), and rs4975616 (OR: 0.778, 95% CI: 0.730-0.829; p value 1.04 × 10-14). All three mapped to Chromosome 5 CLPTM1L-TERT region, previously shown to be associated with lung cancer risk in smokers and in never-smoker Asian women, and risk of other cancers including breast, ovarian, colorectal, and prostate. CONCLUSIONS: We found that genetic susceptibility to lung cancer in never-smokers is associated to genetic variants with pan-cancer risk effects. The comparison with smokers shows that top variants previously shown to be associated with lung cancer risk only confer risk in the presence of tobacco exposure, underscoring the importance of gene-environment interactions in the etiology of this disease.
Subject(s)
Chromosomes, Human, Pair 5 , Lung Neoplasms/epidemiology , Lung Neoplasms/genetics , Membrane Proteins/genetics , Telomerase/genetics , Case-Control Studies , Europe/epidemiology , Female , Genetic Predisposition to Disease , Genetic Variation , Genome-Wide Association Study/methods , Genotyping Techniques/methods , Humans , Middle Aged , Polymorphism, Single Nucleotide , Risk FactorsABSTRACT
Increasing researches have focused on cancer metastasis and development. The ectonucleotidase CD73 is one of the most common cell surface enzymes that are involved in immunosuppression. In this study, the recombinant plasmid pET28a-CD73 was constructed and the CD73 protein was overexpressed in E. coli as an inclusion body that was then subjected to refolding. The anti-CD73 monoclonal antibody (3F7) was obtained by hybridoma technology. The antibody subtype was identified as IgG2a with an affinity constant of 5.75 nM. This antibody could be applied to immunofluorescence and flow cytometry. The results showed that the CD73 protein was not only located in the cytoplasm but also distributed on the surface of triple-negative breast cancer cells MDA-MB-231 and MDA-MB-468. Moreover, the level of CD73 protein was associated with the survival rate. Although the anti-CD73 antibody was not able to inhibit tumor cell growth, it could enhance the cytotoxic effect of Doxorubicin to triple-negative breast cancer cells. In vitro function assay results indicated that anti-CD73 mAb could inhibit cell migration and invasion in both human triple-negative breast cancer and mouse 4T1 cell lines. In this process, both the LC3I/LC3II ratio and p62 protein levels increased, which indicated that the blockage of CD73 could inhibit cell autophagy, and cell migration and invasion were restored by rapamycin. In vivo, anti-CD73 mAb could significantly inhibit lung metastasis of 4T1 cells in a mouse xenograft model. Taken together, this novel anti-CD73 antibody could be developed as an adjuvant drug for triple-negative breast cancer therapy and can be useful in tumor diagnosis.
Subject(s)
5'-Nucleotidase/immunology , Antibodies, Monoclonal/therapeutic use , Autophagy , Cell Movement/drug effects , Triple Negative Breast Neoplasms/metabolism , 5'-Nucleotidase/antagonists & inhibitors , Animals , Antibodies, Monoclonal/pharmacology , Antibodies, Neutralizing/pharmacology , Antibodies, Neutralizing/therapeutic use , Female , Humans , MCF-7 Cells , Mice , Mice, Inbred BALB C , Triple Negative Breast Neoplasms/drug therapyABSTRACT
Human MutT homolog 1 (MTH1) hydrolyses oxidised nucleotide triphosphates, thereby preventing them from being incorporated into DNA; MTH1 has been found to be elevated in many types of cancers, including lung, stomach cancer, melanoma and breast cancer. Thus, tumourtargeted hMTH1 may be valuable for developing novel anticancer therapies. In the present study, we prepared human MTH1 protein and its monoclonal antibody (mAb). The hMTH1 gene was cloned into the prokaryotic expression vector pET28a and optimally expressed in the E. coli Transetta (DE3) strain. Using an NiNTA column and a G50 gel filtration column, 20.1 mg of active hMTH1 was obtained from 1,000 ml of bacterial culture, and the purity was over 98%, as detected by highperformance liquid chromatography (HPLC). The half maximal inhibitory concentration (IC50) of TH287 (hMTH1 inhibitor) was determined to be 3.53±0.47 nM using the recombinant hMTH1 protein (rhMTH1). The enzyme activity assay showed the Michaelis constant (Km) and the catalytic constant (kcat) of the protein were 106.13±48.83 µM and 3.64±0.58 sec1, respectively. The antihMTH1 mAb was obtained via the hybridoma technique and validated by western blot analysis. In addition, an immunofluorescence assay (IFA) and ELISA determined that the mAb could efficiently bind to natural hMTH1 expressed on the human breast cancer cell line MCF7. Taken together, the results showed the rhMTH1 is an active protein and has practical applications for inhibitor selection, and our prepared hMTH1 mAb will provide a valuable tool for the further characterisation of hMTH1 and antitumour medicinal development in future.
Subject(s)
Antibodies, Monoclonal/immunology , DNA Repair Enzymes/immunology , DNA Repair Enzymes/metabolism , Drug Development/methods , Phosphoric Monoester Hydrolases/immunology , Phosphoric Monoester Hydrolases/metabolism , Antibodies, Monoclonal/isolation & purification , Cell Line, Tumor , Cloning, Molecular , DNA Repair Enzymes/antagonists & inhibitors , DNA Repair Enzymes/isolation & purification , Drug Screening Assays, Antitumor/methods , Enzyme Assays/methods , Humans , Oxidation-Reduction , Phosphoric Monoester Hydrolases/antagonists & inhibitors , Phosphoric Monoester Hydrolases/isolation & purification , Recombinant Proteins/immunology , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolismABSTRACT
Bispecific antibodies, which can bind to two different epitopes on the same or different antigens simultaneously, have recently emerged as attractive candidates for study in various diseases. Our present study successfully constructs and expresses a fully human, bispecific, single-chain diabody (BsDb) that can bind to vascular endothelial growth factor 165 (VEGF165) and programmed death-1 (PD-1) in Pichia pastoris. Under the optimal expression conditions (methanol concentration, 1%; pH, 4.0; inoculum density, OD600 = 4, and the induction time, 96 h), the maximum production level of this BsDb is achieved at approximately 20 mg/L. The recombinant BsDb is purified in one step using nickel-nitrilotriacetic acid (Ni-NTA) column chromatography with a purity of more than 95%. Indirect enzyme-linked immune sorbent assay (ELISA) and sandwich ELISA analyses show that purified BsDb can bind specifically to VEGF165 and PD-1 simultaneously with affinities of 124.78 nM and 25.07 nM, respectively. Additionally, the BsDb not only effectively inhibits VEGF165-stimulated proliferation, migration, and tube formation in primary human umbilical vein endothelial cells (HUVECs), but also significantly improves proliferation and INF-γ production of activated T cells by blocking PD-1/PD-L1 co-stimulation. Furthermore, the BsDb displays potent antitumor activity in mice bearing HT29 xenograft tumors by inhibiting tumor angiogenesis and activating immune responses in the tumor microenvironment. Based on these results, we have prepared a potential bispecific antibody drug that can co-target both VEGF165 and PD-1 for the first time. This work provides a stable foundation for the development of new strategies by the combination of an angiogenesis inhibition and immune checkpoint blockade for cancer therapy.
Subject(s)
Antibodies, Bispecific/immunology , Antineoplastic Agents, Immunological/immunology , Neoplasms/drug therapy , Programmed Cell Death 1 Receptor/immunology , Single-Chain Antibodies/immunology , Vascular Endothelial Growth Factor A/immunology , Angiogenesis Inhibitors/genetics , Angiogenesis Inhibitors/immunology , Angiogenesis Inhibitors/pharmacology , Angiogenesis Inhibitors/therapeutic use , Animals , Antibodies, Bispecific/genetics , Antibodies, Bispecific/pharmacology , Antibodies, Bispecific/therapeutic use , Antineoplastic Agents, Immunological/metabolism , Antineoplastic Agents, Immunological/pharmacology , Antineoplastic Agents, Immunological/therapeutic use , Cell Line, Tumor , Cell Movement/drug effects , Cloning, Molecular , Female , Gene Expression , Genetic Vectors/genetics , Human Umbilical Vein Endothelial Cells , Humans , Mice, Inbred BALB C , Mice, Nude , Neoplasms/immunology , Pichia/genetics , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Recombinant Proteins/pharmacology , Recombinant Proteins/therapeutic use , Single-Chain Antibodies/genetics , Single-Chain Antibodies/pharmacology , Single-Chain Antibodies/therapeutic useABSTRACT
Lung cancer has several genetic associations identified within the major histocompatibility complex (MHC); although the basis for these associations remains elusive. Here, we analyze MHC genetic variation among 26,044 lung cancer patients and 20,836 controls densely genotyped across the MHC, using the Illumina Illumina OncoArray or Illumina 660W SNP microarray. We impute sequence variation in classical HLA genes, fine-map MHC associations for lung cancer risk with major histologies and compare results between ethnicities. Independent and novel associations within HLA genes are identified in Europeans including amino acids in the HLA-B*0801 peptide binding groove and an independent HLA-DQB1*06 loci group. In Asians, associations are driven by two independent HLA allele sets that both increase risk in HLA-DQB1*0401 and HLA-DRB1*0701; the latter better represented by the amino acid Ala-104. These results implicate several HLA-tumor peptide interactions as the major MHC factor modulating lung cancer susceptibility.
Subject(s)
Chromosome Mapping , Genetic Predisposition to Disease/genetics , Lung Neoplasms/genetics , Major Histocompatibility Complex/genetics , Asian People/genetics , Female , Gene Frequency , Genetic Predisposition to Disease/ethnology , Genotype , HLA Antigens/genetics , Humans , Lung Neoplasms/ethnology , Male , Middle Aged , Peptides/genetics , Polymorphism, Single Nucleotide , White People/geneticsABSTRACT
Genome-wide association studies (GWAS) identified the chromosome 15q25.1 locus as a leading susceptibility region for lung cancer. However, the pathogenic pathways, through which susceptibility SNPs within chromosome 15q25.1 affects lung cancer risk, have not been explored. We analyzed three cohorts with GWAS data consisting 42,901 individuals and lung expression quantitative trait loci (eQTL) data on 409 individuals to identify and validate the underlying pathways and to investigate the combined effect of genes from the identified susceptibility pathways. The KEGG neuroactive ligand receptor interaction pathway, two Reactome pathways, and 22 Gene Ontology terms were identified and replicated to be significantly associated with lung cancer risk, with P values less than 0.05 and FDR less than 0.1. Functional annotation of eQTL analysis results showed that the neuroactive ligand receptor interaction pathway and gated channel activity were involved in lung cancer risk. These pathways provide important insights for the etiology of lung cancer.
Subject(s)
Chromosomes, Human, Pair 15/genetics , Genetic Predisposition to Disease , Lung Neoplasms/genetics , Adolescent , Adult , Aged , Child , Child, Preschool , Cohort Studies , Female , Gene Ontology , Gene Regulatory Networks , Humans , Infant , Infant, Newborn , Male , Middle Aged , Polymorphism, Single Nucleotide , Quantitative Trait Loci/genetics , Reproducibility of Results , Risk Factors , Smoking/adverse effects , Young AdultABSTRACT
Although several lung cancer susceptibility loci have been identified, much of the heritability for lung cancer remains unexplained. Here 14,803 cases and 12,262 controls of European descent were genotyped on the OncoArray and combined with existing data for an aggregated genome-wide association study (GWAS) analysis of lung cancer in 29,266 cases and 56,450 controls. We identified 18 susceptibility loci achieving genome-wide significance, including 10 new loci. The new loci highlight the striking heterogeneity in genetic susceptibility across the histological subtypes of lung cancer, with four loci associated with lung cancer overall and six loci associated with lung adenocarcinoma. Gene expression quantitative trait locus (eQTL) analysis in 1,425 normal lung tissue samples highlights RNASET2, SECISBP2L and NRG1 as candidate genes. Other loci include genes such as a cholinergic nicotinic receptor, CHRNA2, and the telomere-related genes OFBC1 and RTEL1. Further exploration of the target genes will continue to provide new insights into the etiology of lung cancer.
Subject(s)
Genome-Wide Association Study , Lung Neoplasms/genetics , Adenocarcinoma/genetics , Adenocarcinoma of Lung , Adult , Aged , Chromosome Mapping , Family Health , Female , Genetic Predisposition to Disease , Genotype , Humans , Lung Neoplasms/epidemiology , Lung Neoplasms/ethnology , Male , Middle Aged , Polymorphism, Single Nucleotide , Quantitative Trait Loci , Smoking/epidemiology , Telomere Homeostasis/genetics , White People/geneticsABSTRACT
Metastatic disease is the major cause of death from cancer, and immunotherapy and chemotherapy have had limited success in reversing its progression. Researchers have suggested that inflammatory factors in the tumor environment can promote cancer invasion and metastasis, stimulating cancer progression. Thus, novel strategies that target cytokines and modulate the tumor microenvironment may emerge as important approaches for treating metastatic breast cancer. Specific neutralization of pathogenic TNF signaling using a TNFα antibody has gained increasing attention. Considering this, a selective human TNFα neutralized antibody was generated based on nanobody technology. A TNFα-specific nanobody was produced in Pichia pastoris with a molecular mass of 15 kDa and affinity constant of 2.05 nM. In the proliferation experiment, the TNFα nanobody could inhibit the proliferation of the breast cancer cell line MCF-7 induced by hTNFα in a dose-dependent manner. In the microinvasion model, the TNFα nanobody could inhibit the migration of the breast cancer cell lines MCF-7, MDA-MB-231 and the invasiveness of MDA-MB-231 induced by hTNFα in a dose-dependent manner. Drug administration of the combination of paclitaxel with the TNFα nanobody in vivo significantly enhanced the efficacy against 4T-1 breast tumor proliferation and lung metastasis; meanwhile, E-cadherin tumor epithelial marker expression was upregulated, supporting the anti-tumor therapeutic relevance of paclitaxel and the TNFα nanobody on EMT. This study highlights the importance of neutralizing low TNFα levels in the tumor microenvironment to sensitize the chemotherapeutic response, which has attractive potential for clinical applications.
Subject(s)
Antibodies, Neutralizing/pharmacology , Antineoplastic Agents, Phytogenic/pharmacology , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Breast Neoplasms/drug therapy , Cell Movement/drug effects , Lung Neoplasms/prevention & control , Paclitaxel/pharmacology , Single-Domain Antibodies/pharmacology , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Animals , Antibodies, Neutralizing/administration & dosage , Antibody Affinity , Breast Neoplasms/immunology , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Cadherins/metabolism , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Epithelial-Mesenchymal Transition/drug effects , Female , Humans , Lung Neoplasms/immunology , Lung Neoplasms/metabolism , Lung Neoplasms/secondary , MCF-7 Cells , Male , Mice, Inbred BALB C , Rats, Sprague-Dawley , Signal Transduction/drug effects , Single-Domain Antibodies/administration & dosage , Tumor Microenvironment , Tumor Necrosis Factor-alpha/immunology , Tumor Necrosis Factor-alpha/metabolismABSTRACT
OBJECTIVE: To identify risk alleles relevant to the causal and biologic mechanisms of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). METHODS: A genome-wide association study and subsequent replication study were conducted in a total cohort of 1,986 cases of AAV (patients with granulomatosis with polyangiitis [Wegener's] [GPA] or microscopic polyangiitis [MPA]) and 4,723 healthy controls. Meta-analysis of these data sets and functional annotation of identified risk loci were performed, and candidate disease variants with unknown functional effects were investigated for their impact on gene expression and/or protein function. RESULTS: Among the genome-wide significant associations identified, the largest effect on risk of AAV came from the single-nucleotide polymorphism variants rs141530233 and rs1042169 at the HLA-DPB1 locus (odds ratio [OR] 2.99 and OR 2.82, respectively) which, together with a third variant, rs386699872, constitute a triallelic risk haplotype associated with reduced expression of the HLA-DPB1 gene and HLA-DP protein in B cells and monocytes and with increased frequency of complementary proteinase 3 (PR3)-reactive T cells relative to that in carriers of the protective haplotype. Significant associations were also observed at the SERPINA1 and PTPN22 loci, the peak signals arising from functionally relevant missense variants, and at PRTN3, in which the top-scoring variant correlated with increased PRTN3 expression in neutrophils. Effects of individual loci on AAV risk differed between patients with GPA and those with MPA or between patients with PR3-ANCAs and those with myeloperoxidase-ANCAs, but the collective population attributable fraction for these variants was substantive, at 77%. CONCLUSION: This study reveals the association of susceptibility to GPA and MPA with functional gene variants that explain much of the genetic etiology of AAV, could influence and possibly be predictors of the clinical presentation, and appear to alter immune cell proteins and responses likely to be key factors in the pathogenesis of AAV.
Subject(s)
Granulomatosis with Polyangiitis/genetics , HLA-DP beta-Chains/genetics , Microscopic Polyangiitis/genetics , Myeloblastin/genetics , Protein Tyrosine Phosphatase, Non-Receptor Type 22/genetics , T-Lymphocytes/metabolism , alpha 1-Antitrypsin/genetics , Adult , Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis/genetics , Autoantigens/immunology , B-Lymphocytes/metabolism , Case-Control Studies , Female , Gene Expression , Genetic Predisposition to Disease , Genome-Wide Association Study , HLA-DP Antigens/metabolism , HLA-DP beta-Chains/metabolism , Haplotypes , Humans , Male , Middle Aged , Monocytes/metabolism , Myeloblastin/immunology , Neutrophils/metabolism , Odds Ratio , Peroxidase/immunology , Polymorphism, Single Nucleotide , T-Lymphocytes/immunologyABSTRACT
The role of haplotypes and the interaction of haplotypes and smoking in lung cancer risk have not been well characterized. We analyzed data from an Italian population-based, case-control study with 1815 lung cancer patients and 1959 healthy controls in discovery, and performed a validation using a case-control study with 2983 lung cancer patients and 3553 healthy controls of European ancestry for replication. Sliding window haplotype analysis within chromosome 15, evaluating 4722250 haplotypes and pair-wise haplotype analysis identified that CHRNA5 rs588765-rs16969968 was the most significant haplotype associated with lung cancer risk (omnibus P = 8.35×10(-15) in discovery and 7.26×10(-14) in replication), and improved the prediction of case status over that provided by the individual SNPs rs16969968 or rs588765 (likelihood ratio test P = 0.006 for rs16969968 and 3.83×10(-14) for rs588765 in discovery, 0.009 for rs16969968 and 4.62×10(-13) for rs588765 in replication, compared with rs588765-rs16969968). Compared with the wild-type homozygous diplotype, CA/CA homozygote exhibited an approximately 2-fold increase risk for lung cancer (OR = 2.12; 95% CI 1.46-3.07 in discovery, and OR = 2.01; 95% CI 1.51-2.67 in replication). Even among never-smokers, CA/CA homozygote showed an increased risk of lung cancer with borderline significance in discovery (adjusted OR = 1.75, 95% CI 0.96-3.19) and statistical significance in replication (adjusted OR = 2.10, 95% CI 1.12-3.96), compared with combined genotypes (CG/CG + CG/TG). Accordingly, rs588765-rs16969968 may be a genetic marker to lung cancer risk, even among never-smokers.
Subject(s)
Adenocarcinoma/genetics , Carcinoma, Squamous Cell/genetics , Chromosomes, Human, Pair 15/genetics , Lung Neoplasms/genetics , Aged , Case-Control Studies , Female , Genetic Association Studies , Genetic Loci , Genetic Predisposition to Disease , Haplotypes , Humans , Male , Middle Aged , Polymorphism, Single Nucleotide , Smoking/adverse effectsABSTRACT
Lung adenocarcinoma is caused by the combination of genetic and environmental effects, and smoking plays an important role in the disease development. Exploring the gene expression profile and identifying genes that are shared or vary between smokers and nonsmokers with lung adenocarcinoma will provide insights into the etiology of this complex cancer. We obtained RNA-seq data from paired normal and tumor tissues from 34 nonsmoking and 34 smoking patients with lung adenocarcinoma (GEO: GSE40419). R Bioconductor, edgeR, was adopted to conduct differential gene expression analysis between paired normal and tumor tissues. A generalized linear model was applied to identify genes that were differentially expressed in nonsmoker and smoker patients as well as genes that varied between these two groups. We identified 2273 genes that showed differential expression with FDR < 0.05 and |logFC| >1 in nonsmoker tumor versus normal tissues; 3030 genes in the smoking group; and 1967 genes were common to both groups. Sixty-eight and 70% of the identified genes were downregulated in nonsmoking and smoking groups, respectively. The 20 genes such as SPP1, SPINK1, and FAM83A with largest fold changes in smokers also showed similar large and highly significant fold changes in nonsmokers and vice versa, showing commonalities in expression changes for adenocarcinomas in both smokers and nonsmokers for these genes. We also identified 175 genes that were significantly differently expressed between tumor samples from nonsmoker and smoker patients. Gene expression profile varied substantially between smoker and nonsmoker patients with lung adenocarcinoma. Smoking patients overall showed far more complicated disease mechanism and have more dysregulation in their gene expression profiles. Our study reveals pathogenetic differences in smoking and nonsmoking patients with lung adenocarcinoma from transcriptome analysis. We provided a list of candidate genes for further study for disease detection and treatment in both smoking and nonsmoking patients with lung adenocarcinoma.
Subject(s)
Adenocarcinoma/genetics , Gene Expression Regulation, Neoplastic , Lung Neoplasms/genetics , Neoplasm Proteins/biosynthesis , Smoking/genetics , Adenocarcinoma/pathology , Adenocarcinoma of Lung , High-Throughput Nucleotide Sequencing , Humans , Lung Neoplasms/pathology , Neoplasm Proteins/genetics , RNA/genetics , Smoking/pathology , Transcriptome/geneticsABSTRACT
BACKGROUND: The 15q25.1 lung cancer susceptibility locus, containing CHRNA5, could modify lung cancer susceptibility and multiple smoking related phenotypes. However, no studies have investigated the association between CHRNA5 rs3841324, which has been proven to have the highest association with CHRNA5 mRNA expression, and the risk of other smoking-associated cancers, except lung cancer. In the current study we examined the association between rs3841324 and susceptibility to smoking-associated nasopharyngeal carcinoma (NPC). METHODS: In this case-control study we genotyped the CHRNA5 rs3841324 polymorphism with 400 NPC cases and 491 healthy controls who were Han Chinese and frequency-matched by age (±5 years), gender, and alcohol consumption. Univariate and multivariate logistic regression analyses were used to calculate the odds ratio (OR) and 95% confidence intervals (95% CI). RESULTS: We found that individuals with CHRNA5 rs3841324 combined variant genotypes (ins/del+del/del) had a >1.5-fold elevated risk for NPC than those with the ins/ins genotype (adjusted ORâ=â1.52; 95% CI, 1.16-2.00), especially among ever smokers (adjusted ORâ=â2.07; 95% CI, 1.23-3.48). The combined variant genotypes acted jointly with cigarette smoking to contribute to a 4.35-fold increased NPC risk (adjusted ORâ=â4.35; 95% CI, 2.57-7.38). There was a dose-response relationship between deletion alleles and NPC susceptibility (trend test, Pâ=â0.011). CONCLUSIONS: Our results suggest that genetic variants on the 15q25.1 lung cancer susceptibility locus may influence susceptibility to NPC, particularly for smoking-associated NPC. Such work may be helpful to facilitate an understanding of the etiology of smoking-associated cancers and improve prevention efforts.