Assessment of mirvetuximab soravtansine immunogenicity in patients with folate receptor alpha-positive ovarian cancer.

Aim: The aim of this research was to evaluate the immunogenicity of mirvetuximab soravtansine (MIRV), an antibody-drug conjugate in patients with folate receptor alpha-positive ovarian cancer across four clinical studies.Materials & methods: An assay was developed and validated for the detection of antidrug antibodies (ADAs) against MIRV. A cell-based method was also developed and validated for the detection of neutralizing anti-MIRV antibodies (NAbs). Both ADAs and NAbs were assessed across four clinical studies in 734 patients.Results: Across studies, MIRV demonstrated low immunogenicity with 7.8% of patients with treatment-emergent ADAs, 7.2% with treatment-unaffected ADAs, and 0.5% with treatment-enhanced ADAs. MIRV trough concentrations were comparable in ADA-negative and ADA-positive individuals. Limited data suggest that MIRV ADAs may be associated with decreased efficacy. Due to the very limited number of NAb-positive individuals, no conclusions could be drawn on the effect of NAb on efficacy.Conclusion: Both the validation tests and the data from the MIRV clinical studies demonstrated that these assays were suitable and reliable for the detection of MIRV ADAs and NAbs. These validated assays will continue to be used to monitor MIRV immunogenicity in future clinical trials.

Mirvetuximab soravtansine (MIRV) is a new drug of cancer treatment that targets a protein called folate receptor alpha, which is often found in certain ovarian cancers that do not respond to platinum-based therapies. To check if patients' immune systems react to this drug, we developed tests to look for antibodies that may form against MIRV. These antibodies can affect how well the drug works.The study looked at 734 patients in four different clinical trials. It found that MIRV triggered an immune response in only a small number of patients ­ 7.8% developed new antibodies after starting treatment. However, most of these antibodies did not seem to impact the effectiveness of the drug, and patients with or without antibodies had similar levels of the drug in their bodies. Some data hinted that having antibodies might slightly reduce how well the drug works, but there were not enough patients who developed these antibodies to be certain. Another test was done to check if certain antibodies blocked MIRV from working, but very few patients (31 out of 734) had these, so no conclusions could be made.Overall, the tests were shown to work well and will continue to be used in future studies to monitor how patients' immune systems respond to MIRV.

Exposure-response relationships of mirvetuximab soravtansine in patients with folate receptor-α-positive ovarian cancer: Justification of therapeutic dose regimen.

AIMS: This study aimed to investigate exposure-response (ER) relationships in efficacy and safety for mirvetuximab soravtansine (MIRV) which is a first-in-class antibody-drug conjugate approved for the treatment of folate receptor-α-positive platinum-resistant ovarian cancer. METHODS: MIRV was characterized in 4 clinical studies. Exposure metrics for MIRV, its payload and a metabolite were derived from a population pharmacokinetic model. Efficacy was analysed in MIRV-treated patients (n = 215) in a recent confirmatory, randomized, chemotherapy-controlled MIRASOL trial and safety was evaluated in patients pooled across all 4 clinical studies (n = 757). RESULTS: In the MIRASOL trial (NCT04209855), MIRV demonstrated significant benefit over chemotherapy in progression-free survival (PFS), objective response rate (ORR) and overall survival (OS). The most common adverse events (AEs) included ocular disorders, peripheral neuropathy and pneumonitis. For PFS, ORR and OS, the trough concentration of MIRV was the predictor consistently found in ER models for efficacy. In contrast, for ocular AEs (as well as the time to onset of ocular AEs) and peripheral neuropathy, the area under the concentration-time curve (AUC) of MIRV was identified as the exposure metric in ER models for safety. No exposure parameters were found to correlate with pneumonitis. Covariates in all models did not show clinically meaningful impact on efficacy or safety. Logistic regression models for ORR and ocular AEs based on AUC of MIRV were used to justify the clinical dose regimen approved for MIRV. CONCLUSION: The trough concentration of MIRV correlated with efficacy whereas the AUC of MIRV was associated with major AEs. The ER relationships supported the selected therapeutic dose regimen.

Population pharmacokinetics of mirvetuximab soravtansine in patients with folate receptor-α positive ovarian cancer: The antibody-drug conjugate, payload and metabolite.

AIMS: Mirvetuximab soravtansine is a first-in-class antibody-drug conjugate recently approved for the treatment of folate receptor-α positive ovarian cancer. The aim of this study was to develop a population pharmacokinetic model to describe the concentration-time profiles of mirvetuximab soravtansine, the payload (DM4) and a metabolite (S-methyl-DM4). METHODS: Mirvetuximab soravtansine was administered intravenously from 0.15 to 7 mg/kg to 543 patients with predominantly platinum-resistant ovarian cancer in 3 clinical studies, and the plasma drug concentrations were analysed using a nonlinear mixed-effects modelling approach. Stepwise covariate modelling was performed to identify covariates. RESULTS: We developed a semi-mechanistic population pharmacokinetic model that included linear and nonlinear routes for the elimination of mirvetuximab soravtansine and a target compartment for the formation and disposition of the payload and metabolite in tumour cells. The clearance and volume of the central compartment were 0.0153 L/h and 2.63 L for mirvetuximab soravtansine, 8.83 L/h and 3.67 L for DM4, and 2.04 L/h and 6.3 L for S-methyl-DM4, respectively. Body weight, serum albumin and age were identified as statistically significant covariates. Exposures in patients with renal or hepatic impairment and who used concomitant cytochrome P450 (CYP) 3A4 inhibitors were estimated. CONCLUSION: There is no need for dose adjustment due to covariate effects for mirvetuximab soravtansine administered at the recommended dose of 6 mg/kg based on adjusted ideal body weight. Dose adjustment is not required for patients with mild or moderate renal impairment, mild hepatic impairment, or when concomitant weak and moderate CYP3A4 inhibitors are used.

Chemical complementarity between immune receptor CDR3s and candidate cancer antigens correlating with reduced survival: evidence for outcome mitigation with corticosteroid treatments.

The evaluation of physicochemical characteristics of extensive adaptive immune receptor (IR) recombination sequence collections has led to the discovery of many correlations of those sequences and a variety of diseases, including cancer. In the cancer setting, these evaluations have recently focused on the adaptive IR, complementarity determining region-3 (CDR3) amino acid (AA) sequences, which play a major role in antigen binding. For example, the chemical complementarities of the tumor resident, CDR3 AA sequences and the BRAFV600E mutant, common in melanoma, have proved informative with regard to outcomes. Many of these evaluations led to the conclusion that a high affinity match, efficiently, algorithmically designated as a high chemical complementarity score (CS) for the patient specific, IR CDR3 AA sequences and the cancer antigens, correlated with improved survival outcomes. In this report, the complementarity scoring algorithms were used to investigate the opposite phenomenon, high complementarity chemistry between CRD3 AAs and cancer antigens that correlated with a worse survival, an approach that revealed potential risk stratification biomarkers for lung adenocarcinoma, lung squamous carcinoma, and likely other cancer types. Most importantly, analyses suggested that high IR CDR3 AA-candidate antigen CS, low overall survival results for low grade glioma were mitigated by neoadjuvant corticosteroid treatments. Overall, the analyses of this report, coupled with earlier work establishing the CS approach for identifying likely good outcomes, have the potential to distinguish patients who will benefit from (i) immune activating or (ii) immune augmenting or (iii) even immunosuppressive treatment strategies.Communicated by Ramaswamy H. Sarma.

Clinical and molecular implications of RGS2 promoter genetic variation in severe asthma.

BACKGROUND: Regulator of G protein signaling (RGS) 2 terminates bronchoconstrictive Gαq signaling; murine RGS2 knockout demonstrate airway hyperresponsiveness. While RGS2 promoter variants rs2746071 and rs2746072 associate with a clinical mild asthma phenotype, their impact on human airway smooth muscle (HASM) contractility and asthma severity outcomes is unknown. OBJECTIVE: We sought to determine whether reductions in RGS2 expression seen with these 2 RGS2 promoter variants augment HASM contractility and associate with an asthma severity phenotype. METHODS: We transfected HASM with a range of RGS2-specific small interfering RNA (siRNA) concentrations and determined RGS2 protein expression by Western blot analysis and intracellular calcium flux induced by histamine (a Gαq-coupled H1 receptor bronchoconstrictive agonist). We conducted regression-based genotype association analyses of RGS2 variants from 611 patients from the National Heart, Lung, and Blood Institute Severe Asthma Research Program 3. RESULTS: RGS2-specific siRNA caused dose-dependent increases in histamine-stimulated bronchoconstrictive intracellular calcium signaling (2-way ANOVA, P < .0001) with a concomitant decrease in RGS2 protein expression. RGS2-specific siRNA did not affect Gαq-independent ionomycin-induced intracellular calcium signaling (P = .42). The minor allele frequency of rs2746071 and rs2746072 was 0.46 and 0.28 among African American/non-Hispanic Black patients and was 0.28 and 0.27 among non-Hispanic White patients, among whom these single nucleotide polymorphisms were in stronger linkage disequilibrium (r2 = 0.97). Among non-Hispanic White patients, risk allele homozygotes for rs2746072 and rs2746071 each had nearly 2-fold greater asthma exacerbation rates relative to alternative genotypes with wild-type alleles (Padditive = 2.86 × 10-5/Precessive = 5.22 × 10-6 and Padditive = 3.46 × 10-6/Precessive = 6.74 × 10-7, respectively) at baseline, which was confirmed by prospective longitudinal exacerbation data. CONCLUSION: RGS2 promoter variation associates with a molecular and clinical phenotype characterized by enhanced bronchoconstrictive stimulation in vitro and higher asthma exacerbations rates in non-Hispanic White patients.

TH17 cells and corticosteroid insensitivity in severe asthma.

Asthma is classically described as having either a type 2 (T2) eosinophilic phenotype or a non-T2 neutrophilic phenotype. T2 asthma usually responds to classical bronchodilation therapy and corticosteroid treatment. Non-T2 neutrophilic asthma is often more severe. Patients with non-T2 asthma or late-onset T2 asthma show poor response to the currently available anti-inflammatory therapies. These therapeutic failures result in increased morbidity and cost associated with asthma and pose a major health care problem. Recent evidence suggests that some non-T2 asthma is associated with elevated TH17 cell immune responses. TH17 cells producing Il-17A and IL-17F are involved in the neutrophilic inflammation and airway remodeling processes in severe asthma and have been suggested to contribute to the development of subsets of corticosteroid-insensitive asthma. This review explores the pathologic role of TH17 cells in corticosteroid insensitivity of severe asthma and potential targets to treat this endotype of asthma.

PEG-GO@XN nanocomposite suppresses breast cancer metastasis via inhibition of mitochondrial oxidative phosphorylation and blockade of epithelial-to-mesenchymal transition.

Metastatic breast cancer is a significant contributor to mortality among women, but its complex regulation represents a barrier to precision targeting. In the present study, a graphene-based nanocomposite which probes and selectively inhibits cancer cell motility is described. By controllable coupling of prenylated chalcone xanthohumol, an efficient inhibitor of mitochondrial electron transport chain complex I, with PEGylated graphene oxide nanosheet, a PEG-GO@XN nanocomposite with good stability and biocompatibility is synthesized. PEG-GO@XN is capable of inhibiting mitochondrial oxidative phosphorylation selectively in MDA-MB-231 and MDA-MB-436 metastatic breast cancer cells. PEG-GO@XN reduces the production of ATP, impairs the formation of F-actin cytoskeleton in the lamellipodia, and blocks the migration and invasion of breast cancer cells in vitro, without interfering the proliferation and metabolism of non-cancerous cells. More importantly, PEG-GO@XN suppresses the metastasis of MDA-MB-231 cells to lung in nude mice. PEG-GO@XN abolishes the TGF-ß1-induced down-regulation of E-cadherin and up-regulation of N-cadherin, vimentin, Snail and Twist, thus causes the maintenance of "epithelial-like" rather than the "mesenchymal-like" features, and decreases the motility potential of breast cancer cells. Taken together, this research unveils the enormous potential of PEG-GO@XN to suppress metastatic breast cancer by selective targeting oxidative phosphorylation and epithelial-mesenchymal transition of cancer cells and thereby providing insights on metastatic cancer treatment.

Anti-EGFR Single-Chain Fv Antibody Fragment Displayed on the Surface of Ferritin H-Chain Protein Nanoparticle for Asthma Therapy.

Epidermal growth factor receptor (EGFR)-dependent signaling contributes to the pathophysiology of asthma. However, these findings have not been translated into a clinical application. We recently generated ferritin H-chain protein (FTH1)-based nanoparticles with an anti-EGFR single-chain Fv (anti-EGFR scFv) on the surface of FTH1, namely, anti-EGFR scFv-FTH1/FTH1 nanoparticles. In the present study, we found that these nanoparticles could specifically bind to EGFR-expressing cells, leading to downregulation of EGFR and mucin 5AC (MUC5AC) protein expression and growth suppression of House Dust Mite (HDM)-stimulated human bronchial epithelial 16HBE and lipopolysaccharides (LPS)-activated murine macrophage-like RAW264.7 cells. In vivo, intraperitoneal administration of anti-EGFR scFv-FTH1/FTH1 nanoparticles, but not FTH1 nanoparticles, alleviated the major pathological symptoms including airway hyperresponsiveness, airway inflammation, goblet cell hyperplasia, mucus hyperproduction, and increased release of Th2 cytokines in an allergen ovalbumin (OVA)-induced asthma mouse model. Importantly, during the dosing period these nanoparticles were safe for both heathy and asthmatic mice, and more effective in controlling airway inflammation than cetuximab, an EGFR monoclonal antibody. Altogether, our studies provide insights into the control of airway inflammation for treatment of asthma by targeting EGFR. The similar strategy can be used to fabricate scFv-based recombinant protein nanoparticles for other clinical applications.

Airway relaxation mechanisms and structural basis of osthole for improving lung function in asthma.

Overuse of ß2-adrenoceptor agonist bronchodilators evokes receptor desensitization, decreased efficacy, and an increased risk of death in asthma patients. Bronchodilators that do not target ß2-adrenoceptors represent a critical unmet need for asthma management. Here, we characterize the utility of osthole, a coumarin derived from a traditional Chinese medicine, in preclinical models of asthma. In mouse precision-cut lung slices, osthole relaxed preconstricted airways, irrespective of ß2-adrenoceptor desensitization. Osthole administered in murine asthma models attenuated airway hyperresponsiveness, a hallmark of asthma. Osthole inhibited phosphodiesterase 4D (PDE4D) activity to amplify autocrine prostaglandin E2 signaling in airway smooth muscle cells that eventually triggered cAMP/PKA-dependent relaxation of airways. The crystal structure of the PDE4D complexed with osthole revealed that osthole bound to the catalytic site to prevent cAMP binding and hydrolysis. Together, our studies elucidate a specific molecular target and mechanism by which osthole induces airway relaxation. Identification of osthole binding sites on PDE4D will guide further development of bronchodilators that are not subject to tachyphylaxis and would thus avoid ß2-adrenoceptor agonist resistance.

KAP1-associated transcriptional inhibitory complex regulates C2C12 myoblasts differentiation and mitochondrial biogenesis via miR-133a repression.

The differentiation of myoblasts plays a key role in the growth of biological individuals and the reconstruction of muscle tissue. Several microRNAs are significantly upregulated during the differentiation of myoblasts and their target genes have been explored. However, the molecular mechanisms underlying the transcriptional regulation of microRNAs remain elusive. In the present study, we found that the expression of miR-133a is increased during the differentiation of C2C12 myoblasts. miR-133a mimic is sufficient to induce the biogenesis of mitochondria and differentiation of C2C12 myoblasts whereas miR-133a inhibitor abolishes cell differentiation. Using CRISPR affinity purification in situ of regulatory elements (CAPTURE) technique, we further dissected the regulatory mechanisms of miR-133a expression and found that KAP1-associated transcription complex accounts for the suppression of miR-133a in C2C12 myoblasts. Knockdown of KAP1 increased the expression of miR-133a, which contributed to the biogenesis of mitochondria and differentiation of C2C12 myoblasts. To our knowledge, this is the first study using the CAPTURE technology to identify the regulatory factors of miR-133a during cell differentiation, which may provide new ideas for understanding the precision regulatory machinery of microRNAs during different biological processes.

TGF-ß1-induced miR-424 promotes pulmonary myofibroblast differentiation by targeting Slit2 protein expression.

Idiopathic pulmonary fibrosis (IPF) is a devastating interstitial lung disease with irreversible loss of lung tissue and function. Myofibroblasts in the lung are key cellular mediators of IPF progression. Transforming growth factor (TGF)-ß1, a major profibrogenic cytokine, induces pulmonary myofibroblast differentiation, and emerging evidence has established the importance of microRNAs (miRs) in the development of IPF. The objective of this study was to define the pro-fibrotic roles and mechanisms of miRs in TGF-ß1-induced pulmonary myofibroblast differentiation. Using RNA sequencing, we identified miR-424 as an important TGF-ß1-induced miR in human lung fibroblasts (HLFs). Quantitative RT-PCR confirmed that miR-424 expression was increased by 2.6-fold in HLFs in response to TGF-ß1 and was 1.7-fold higher in human fibrotic lung tissues as compared to non-fibrotic lung tissues. TGF-ß1-induced upregulation of miR-424 was blocked by the Smad3 inhibitor SIS3, suggesting the involvement of this canonical TGF-ß1 signaling pathway. Transfection of a miR-424 hairpin inhibitor into HLFs reduced TGF-ß1-induced expression of classic myofibroblast differentiation markers including ɑ-smooth muscle actin (ɑ-SMA) and connective tissue growth factor (CTGF), whereas a miR-424 mimic significantly enhanced TGF-ß1-induced myofibroblast differentiation. In addition, TGF-ß1 induced Smad3 phosphorylation in HLFs, and this response was reduced by the miR-424 inhibitor. In silico analysis identified Slit2, a protein that inhibits TGF-ß1 profibrogenic signaling, as a putative target of regulation by miR-424. Slit2 is less highly expressed in human fibrotic lung tissues than in non-fibrotic lung tissues, and knockdown of Slit2 by its siRNA enhanced TGF-ß1-induced HLF differentiation. Overexpression of a miR-424 mimic down-regulated expression of Slit2 but not the Slit2 major receptor ROBO1 in HLFs. Luciferase reporter assays showed that the miR-424 mimic represses Slit2 3' untranslated region (3'-UTR) reporter activity, and mutations at the seeding regions in the 3'-UTR of Slit2 abolish this inhibition. Together, these data demonstrate a pro-fibrotic role of miR-424 in TGF-ß1-induced HLF differentiation. It functions as a positive feed-back regulator of the TGF-ß1 signaling pathway by reducing expression of the negative regulator Slit2. Thus, targeting miR-424 may provide a new therapeutic strategy to prevent myofibroblast differentiation and IPF progression.

Antiviral T Cell Receptor Complementarity Determining Region-3 Sequences Are Associated with a Worse Cancer Outcome: A Pancancer Analysis.

Human papilloma virus has a clearly demonstrated role in cervical and head and neck cancers, but viral etiology for other solid tumors is less well understood. To expand this area of research, we obtained and analyzed the immune receptor recombinations available from both blood and tumor samples, through mining of exome files produced from those sources, for 32 cancer types represented by the cancer genome atlas (TCGA). Among TCGA data sets, the recovery frequency for antiviral complementarity determining region-3 sequences (CDR3s), for T cell receptor-alpha and T cell receptor-beta, ranged from 0% to 21% of the patients, for the different cancer types, with breast, lung, pancreatic, and thymus cancers representing the highest of that range, particularly for tumor tissue resident T cells. In several cases, recovery of the antiviral CDR3s associated with distinct survival rates, and in all of these cases, the recovery of an antiviral CDR3 associated with a worse survival rate.

B-cell Receptor Recombinations in Lung Adenocarcinoma Exome Files Correlate With a Higher Overall Survival Rate.

BACKGROUND/AIM: While there has been a rapid development in genomic data mining approaches for T-cell receptor recombinations (TcR), less emphasis has been placed on B-cell receptor (BcR) recombinations. MATERIALS AND METHODS: We obtained lung cancer exome files from the cancer genome atlas (TCGA) and mined the files for TcR and BcR recombination reads. RESULTS: There was a robust detection of BcR light chain recombination reads in lung adenocarcinoma (TCGA-LUAD) samples, and there was a correlation between the detection of light chain recombination reads and a more favorable outcome. This result was supported by analyses of the expression of B-cell markers as indicated by LUAD RNASeq files. CONCLUSION: BcR and TcR recombination reads recovered from LUAD WXS files, either alone or in combination with the human leukocyte antigen (HLA) type, are likely to have prognostic value.

Transforming growth factor (TGF)-ß1-induced miR-133a inhibits myofibroblast differentiation and pulmonary fibrosis.

Transforming growth factor (TGF)-ß1, a main profibrogenic cytokine in the progression of idiopathic pulmonary fibrosis (IPF), induces differentiation of pulmonary fibroblasts to myofibroblasts that produce high levels of collagen, leading to concomitantly loss of lung elasticity and function. Recent studies implicate the importance of microRNAs (miRNAs) in IPF but their regulation and individual pathological roles remain largely unknown. We used both RNA sequencing and quantitative RT-PCR strategies to systematically study TGF-ß1-induced alternations of miRNAs in human lung fibroblasts (HFL). Our data show that miR-133a was significantly upregulated by TGF-ß1 in a time- and concentration-dependent manner. Surprisingly, miR-133a inhibits TGF-ß1-induced myofibroblast differentiation whereas miR-133a inhibitor enhances TGF-ß1-induced myofibroblast differentiation. Interestingly, quantitative proteomics analysis indicates that miR-133a attenuates myofibroblast differentiation via targeting multiple components of TGF-ß1 profibrogenic pathways. Western blot analysis confirmed that miR-133a down-regulates TGF-ß1-induced expression of classic myofibroblast differentiation markers such as ɑ-smooth muscle actin (ɑ-SMA), connective tissue growth factor (CTGF) and collagens. miRNA Target Searcher analysis and luciferase reporter assays indicate that TGF-ß receptor 1, CTGF and collagen type 1-alpha1 (Col1a1) are direct targets of miR-133a. More importantly, miR-133a gene transferred into lung tissues ameliorated bleomycin-induced pulmonary fibrosis in mice. Together, our study identified TGF-ß1-induced miR-133a as an anti-fibrotic factor. It functions as a feed-back negative regulator of TGF-ß1 profibrogenic pathways. Thus, manipulations of miR-133a expression may provide a new therapeutic strategy to halt and perhaps even partially reverse the progression of IPF.

Immune receptor recombinations from breast cancer exome files, independently and in combination with specific HLA alleles, correlate with better survival rates.

PURPOSE: Immune characterizations of cancers, including breast cancer, have led to information useful for prognoses and are considered to be important in the future of refining the use of immunotherapies, including immune checkpoint inhibitor therapies. In this study, we sought to extend these characterizations with genomics approaches, particularly with cost-effective employment of exome files. METHODS: By recovery of immune receptor recombination reads from the cancer genome atlas (TCGA) breast cancer dataset, we observed associations of these recombinations with T-cell and B-cell biomarkers and with distinct survival rates. RESULTS: Recovery of TRD or IGH recombination reads was associated with an improved disease-free survival (p = 0.047 and 0.045, respectively). Determination of the HLA types using the exome files allowed matching of T-cell receptor V- and J-gene segment usage with specific HLA alleles, in turn allowing a refinement of the association of immune receptor recombination read recoveries with survival. For example, the TRBV7, HLA-C*07:01 combination represented a significantly worse, disease-free outcome (p = 0.014) compared to all other breast cancer samples. By direct comparisons of distinct TRB gene segment usage, HLA allele combinations revealed breast cancer subgroups, within the entire TCGA breast cancer dataset with even more dramatic survival distinctions. CONCLUSIONS: In sum, the use of exome files for recovery of adaptive immune receptor recombination reads, and the simultaneous determination of HLA types, has the potential of advancing the use of immunogenomics for immune characterization of breast tumor samples.

TRB-J1 usage, in combination with the HLA-A*01:01 allele, represents an apparent survival advantage for uterine corpus endometrial carcinoma: Comparisons with microscopic assessments of lymphocyte infiltrates.

The opportunity for the highly efficient recovery of immune receptor recombination data from cancer specimens, including the ready assessment of immune receptor V and J usage, raises the issue of establishing precise values of assessing the immune receptor status as opposed to obtaining basic information regarding lymphocyte infiltration, in the cancer setting. In this report, we obtained the lymphocyte infiltration percentages from the cancer digital slide archive representing uterine corpus endometrial carcinoma (UCEC) and correlated these data with recovery of the immune receptor recombination reads from corresponding UCEC exome files. Results indicated a basic correlation of the recovery of productive T-cell receptor beta (TRB) recombination reads with lymphocyte infiltration percentages. However, the recovery of specific immune receptor recombination reads did not indicate the same survival outcomes as microscope detection of lymphocyte infiltrate percentages. To further exploit the value of recovery of the TRB recombination reads from the UCEC exome files, we determined the survival outcomes for combinations of TRB gene segment usage and HLA class I alleles, with the most important result being that the combination of HLA-A*01:01 and TRB-J1 segment usage reflected a strikingly high survival rate. Overall, this report emphasized the increased value of the knowledge of the immune receptor recombinations, in comparison with basic lymphocyte infiltration percentages, in assessing cancer survival rates.

Up-regulated miR-133a orchestrates epithelial-mesenchymal transition of airway epithelial cells.

Dysregulation of microRNAs (miRNAs) contributes to epithelial-mesenchymal transition (EMT) of cancer, but the pathological roles of miRNAs in airway EMT of lung diseases remains largely unknown. We performed sequencing and real-time PCR analysis of the miRNA expression profile of human airway epithelial cells undergoing EMT, and revealed miR-133a to be one of the most common up-regulated miRNAs. MiR-133a was previously reported to be persistently up-regulated in airway epithelial cells of smokers. We found that mice exposed to cigarette smoke (CS) showed airway hyper-responsiveness, a typical symptom occurring in CS-related lung diseases, up-regulation of miR-133a and EMT marker protein N-cadherin in airway epithelium. Importantly, miR-133a overexpression induces airway epithelial cells to undergo spontaneous EMT via down-regulation of grainyhead-like 2 (GRHL2), an epithelial specific transcriptional factor. Loss of GRHL2 causes down-regulation of epithelial splicing regulatory protein 1 (ESRP1), a central coordinator of alternative splicing processes that are critical in the regulation of EMT. Down-regulation of ESRP1 induces isoform switching of adherens junction-associated protein p120-catenin, and leads to the loss of E-cadherin. Our study is the first to demonstrate that up-regulated miR-133a orchestrates airway EMT via alternative splicing processes, which points to novel therapeutic possibilities for the treatment of CS-related lung disease.

Novel role of Giα2 in cell migration: Downstream of PI3-kinase-AKT and Rac1 in prostate cancer cells.

Tumor cell motility is the essential step in cancer metastasis. Previously, we showed that oxytocin and epidermal growth factor (EGF) effects on cell migration in prostate cancer cells require Giα2 protein. In the current study, we investigated the interactions among G-protein coupled receptor (GPCR), Giα2, PI3-kinase, and Rac1 activation in the induction of migratory and invasive behavior by diverse stimuli. Knockdown and knockout of endogenous Giα2 in PC3 cells resulted in attenuation of transforming growth factor ß1 (TGFß1), oxytocin, SDF-1α, and EGF effects on cell migration and invasion. In addition, knockdown of Giα2 in E006AA cells attenuated cell migration and overexpression of Giα2 in LNCaP cells caused significant increase in basal and EGF-stimulated cell migration. Pretreatment of PC3 cells with Pertussis toxin resulted in attenuation of TGFß1- and oxytocin-induced migratory behavior and PI3-kinase activation without affecting EGF-induced PI3-kinase activation and cell migration. Basal- and EGF-induced activation of Rac1 in PC3 and DU145 cells were not affected in cells after Giα2 knockdown. On the other hand, Giα2 knockdown abolished the migratory capability of PC3 cells overexpressing constitutively active Rac1. The knockdown or knockout of Giα2 resulted in impaired formation of lamellipodia at the leading edge of the migrating cells. We conclude that Giα2 protein acts at two different levels which are both dependent and independent of GPCR signaling to induce cell migration and invasion in prostate cancer cells and its action is downstream of PI3-kinase-AKT-Rac1 axis.

Elucidating feed-forward apoptosis signatures in breast cancer datasets: Higher FOS expression associated with a better outcome.

Overstimulation of pro-proliferative pathways and high level expression of pro-proliferative transcription factors (TFs) can lead to apoptosis. This is likely due to TF binding sites for pro-proliferative TFs common to pro-proliferative and pro-apoptosis-effector genes. Certain clinical datasets have indicated that molecular markers associated with higher proliferation rates lead to improved outcomes for patients with cancer. These observations have been extensively assessed on a general basis, however there has been little work dissecting feed-forward apoptosis signaling pathways that may represent specific distinctions between a pro-proliferative mechanism and a pro-apoptotic mechanism in samples from patients with cancer. Using The Cancer Genome Atlas datasets and bioinformatic approaches, the present study reports that higher FOS expression levels, along with higher FOS target apoptosis-effector gene expression, is associated with an increased survival, while higher POU2F1 expression is associated with a reduced survival (average difference of 25.9 months survival). In summary, in the datasets examined FOS represents an apoptosis-driver and high POU2F1 represents a driver mechanism for cancer development.