Integrated investigation of the prognostic role of HLA LOH in advanced lung cancer patients with immunotherapy.

Although multiple studies have shown that loss of heterozygosity (LOH) at the human leukocyte antigen (HLA) locus is one of the mechanisms of immune escape, the effect of HLA LOH on the immunotherapy response of patients is still unclear. Based on the data of 425 Chinese lung cancer patients, the genomic characteristics with different HLA LOH statuses were analyzed. The driver genes mutation frequency, oncogenic signaling pathways mutation frequency, tumor mutational burden (TMB) and chromosomal instability (CIN) score in the HLA LOH high group was significantly higher than in the HLA LOH negative group. Transcriptome analyses revealed that pre-existing immunologically active tumor microenvironment (TME) was associated with HLA LOH negative patients. Non-small cell lung cancer (NSCLC) patients, especially for lung squamous cell carcinomas (LUSC), with HLA LOH negative have a longer survival period than those with HLA LOH. In addition, the combination of HLA LOH with TMB or programmed cell death-Ligand 1 (PD-L1) expression can further distinguish responders from non-responders. Furthermore, a comprehensive predictive model including HLA LOH status, TMB, PD-L1 expression and CD8+ T cells was constructed and exhibited a higher predictive value, which may improve clinical decision-making.

Identification of a prognostic immune-related signature for small cell lung cancer.

PURPOSE: As a subgroup of lung cancer, small cell lung cancer (SCLC) is characterized by a short tumor doubling time, high rates of early occurred distant cancer spread, and poor outcomes. Despite its exquisite sensitivity to chemotherapy and radiotherapy, acquired drug resistance and tumor progression are typical. This study aimed to develop a robust signature based on immune-related genes to predict the outcome of patients with SCLC. METHODS: The expression data of 77 SCLC patients from George's cohort were divided into training set and testing set, and 1534 immune-related genes from ImmPort database were used to generate and validate the signature. Cox proportional hazards and the Kaplan-Meier analysis were used for developing and testing the prognostic signature. Single-sample gene set enrichment analysis was used to determine immune cell infiltration phenotypes. RESULTS: A 10-gene model comprising NR3C1, NR1D2, TANK, ARAF, HDGF, INHBE, LRSAM1, PLXNA1, PML, and SP1 with the highest frequency after 1000 interactions, was chosen to construct immune-related signature. This signature showed robust predictive value for SCLC patients' survival in both training and testing sets. This signature was weakly associated with the clinic pathological values like TNM stage. Furthermore, patients with low risk presented with activation of immune signal pathways, and specific immune cell infiltration with high levels of CD56bright NK cells but low levels of CD8+ T cells, mast cells, and helper T cells. CONCLUSION: The present study developed immune-related signature that may help predict the prognosis of SCLC patients, which reflects an unappreciated level of heterogeneity of immunophenotype associated with diverse prognosis for specific subsets in this highly lethal cancer type.

Decreased human antigen R expression confers resistance to tyrosine kinase inhibitors in epidermal growth factor receptor-mutant lung cancer by inhibiting Bim expression.

Primary resistance to epidermal growth factor receptor­tyrosine kinase inhibitors (EGFR­TKIs) is an obstacle for the treatment of non­small cell lung cancer (NSCLC); however, the associated mechanisms are not well understood. Studies have reported that Bim expression levels may be associated with the efficacy of EGFR­TKI treatment in NSCLC patients harboring EGFR mutations. Human antigen R (HuR) regulates the mRNA and protein expression of target genes, including certain B­cell lymphoma 2 family members. The present study investigated whether HuR mediates resistance to EGFR­TKIs via the regulation of Bim. The results demonstrated that decreased levels of HuR and Bim protein expression are associated with primary resistance to EGFR­TKIs and reduced median progression­free survival in NSCLC patients. In vitro assays also revealed that knockdown of HuR resulted in primary EGFR­TKI resistance and reduced gefitinib­induced apoptosis in HCC827 cells by decreasing Bim expression. Furthermore, elevated HuR expression restored gefitinib sensitivity and enhanced gefitinib­induced apoptosis in H1650 cells by increasing Bim expression. In vivo, it was further demonstrated that overexpression of HuR was able to restore the gefitinib sensitivity of H1650 cells. Therefore, altered HuR/Bim expression is proposed to be a novel mechanism of EGFR­TKI resistance in NSCLC.

PIK3CA mutations confer resistance to first-line chemotherapy in colorectal cancer.

Chemotherapy represents an important treatment option for colorectal cancer (CRC), but only half of the patients benefit from these regimens. We explored the potential predicting value and mechanism of PIK3CA mutation in CRC chemotherapy. CRC specimens from 440 patients were retrospectively collected and examined with a fluorescence PCR-based method. The correlation of first-line chemotherapy response and PIK3CA mutation was evaluated according to follow-up and medical records. The underlying mechanism of PIK3CA mutation in chemotherapy resistance was assessed with CRC tumors and primary cells. The mutation frequency of the PIK3CA gene in CRC patients was 9.55%, which was correlated with late TNM staging and lower histological grade. The CRC patients with PIK3A mutation showed worse response to first-line chemotherapy than those without PIK3CA mutation. PIK3A mutation tumor cells showed poor sensitivity to first-line chemotherapy in vitro and in vivo. PIK3CA mutation induced PI3K/Akt signaling activation to increase LGR5+ CRC stem cells survival and proliferation, from which lead to chemotherapy resistance. Furthermore, PIK3CA mutation/LGR5+ expression was an independent detrimental factor for CRC patients. Our findings indicated that PIK3CA mutation induced PI3K/Akt activation contributed to CRC stem cells survival and proliferation, from which cells further resistance to chemotherapy. PIK3CA mutation/LGR5+ expression was a potential biomarker for monitoring chemotherapy resistance in CRC.

Strategies targeting angiogenesis in advanced non-small cell lung cancer.

Tumor angiogenesis is a frequent event in the development and progression of non-small cell lung cancer (NSCLC) and has been identified as a promising therapeutic target. The vascular endothelial growth factor (VEGF) family and other angiogenic factors, including fibroblast growth factor and platelet-derived growth factor, promote the growth of newly formed vessels from preexisting vessels and change the tumor microenvironment. To date, two antiangiogenic monoclonal antibodies, bevacizumab and ramucirumab, which target VEGF-A and its receptor VEGF receptor-2, respectively, have been approved for the treatment of locally advanced or metastatic NSCLC when added to first-line standard chemotherapy. Numerous oral multitargeting angiogenic small molecule tyrosine kinase inhibitors (TKIs) have been widely evaluated in advanced NSCLC, but only nintedanib in combination with platinum-based doublet chemotherapy has demonstrated a survival benefit in the second-line setting. Additionally, small-molecule TKIs remain the standard of care for patients with mutated EGFR, ALK or ROS1. Moreover, immune checkpoint inhibitors that target the programmed cell death protein 1 (PD-1) and programmed cell death protein ligand 1 (PD-L1) are changing the current strategy in the treatment of advanced NSCLC without driver gene mutations. The potential synergistic activity of antiangiogenic agents and TKIs or immunotherapy is an interesting topic of research. This review will summarize the novel antiangiogenic agents, antiangiogenic monotherapy, as well as potential combination therapeutic strategies for the clinical management of advanced NSCLC.

Pretreatment Neutrophil to Lymphocyte Ratio Is Associated with Poor Survival in Patients with Stage I-III Non-Small Cell Lung Cancer.

BACKGROUND: Neutrophil-to-lymphocyte ratio (NLR) has been shown to be a prognostic indicator in several types of cancer. We aimed to investigate the association between NLR and survival in surgery-treated non-small cell lung cancer (NSCLC) patients. STUDY DESIGN: This large retrospective study included 1,245 patients who underwent initial surgery for stage I-III NSCLC at The University of Texas MD Anderson Cancer Center between December 2002 and November 2010. We analyzed the relationship of NLR with clinicopathological variables, local recurrence-free survival (LRFS), distant recurrence-free survival (DRFS), recurrence-free survival (RFS), overall survival (OS), and disease-specific survival (DSS) in patients with high or low NLR using Kaplan-Meier method. Hazard ratios (HRs) with 95% confidence intervals (CIs) were used to assess the prognostic strength of NLR. RESULTS: There was a statistically significant association between the pretreatment NLR and histology type (P = 0.003) and tumor grade (P = 0.028). At a median follow-up time of 50.6 months, high NLR was associated with reduced DRFS (P = 0.011), OS (P < 0.0001) and DSS (P = 0.004); it was not associated with LRFS and RFS. Multivariable Cox analysis further revealed that NLR (P = 0.027), pathologic stage (P < 0.0001) and lymphovascular invasion (P < 0.0001) were strong independent predictors for DRFS. NLR was also an independent marker predicting poor OS (P = 0.002) and DSS (P = 0.017). CONCLUSION: The pretreatment NLR can serve as a biomarker to predict distant recurrence and death in stage I-III NSCLC patients. Combination of NLR and pathologic stage can better predict the OS and DSS in stage I-II NSCLC patients.

Intrinsic resistance to EGFR tyrosine kinase inhibitors in advanced non-small-cell lung cancer with activating EGFR mutations.

Identifying activating EGFR mutations is a useful predictive strategy that helps select a population of advanced non-small-cell lung cancer (NSCLC) patients for treatment with EGFR tyrosine kinase inhibitors (TKIs). Patients with sensitizing EGFR mutations (predominantly an in-frame deletion in exon 19 and an L858R substitution) are highly responsive to first-generation EGFR TKIs, such as gefitinib and erlotinib, and show improved progression-free survival without serious side effects. However, all patients with activating EGFR mutations who are initially responsive to EGFR TKIs eventually develop acquired resistance after a median progression-free survival of 10-16 months, followed by disease progression. Moreover, ~20%-30% of NSCLC patients have no objective tumor regression on initial EGFR TKI treatment, although they harbor an activating EGFR mutation. These patients represent an NSCLC subgroup that is defined as having intrinsic or primary resistance to EGFR TKIs. Different mechanisms of acquired EGFR TKI resistance have been identified, and several novel compounds have been developed to reverse acquired resistance, but little is known about EGFR TKI intrinsic resistance. In this review, we summarize the latest findings involving mechanisms of intrinsic resistance to EGFR TKIs in advanced NSCLC with activating EGFR mutations and present possible therapeutic strategies to overcome this resistance.

Cytoplasmic expression of the ELAV-like protein HuR as a potential prognostic marker in esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma (ESCC) is one of the most frequent cancers and a leading cause of death from cancer in China. The human ELAV-like protein HuR has been found to contribute to cancer development and progression through stabilizing a group of cellular mRNAs of cancer-related genes. In this study, we investigated the expression of HuR in a cohort of ESCC patients using immunohistochemical staining. HuR detected in the cytoplasm of cancer cells was positive in 46.6% of 58 ESCC specimens; 75.9% of these specimens had nuclear immunoreactivity for HuR. Cytoplasmic HuR expression was higher in cancer tissues compared to 20 matched adjacent noncancerous tissues. A clinicopathological study showed that cytoplasmic HuR expression was positively associated with lymph node metastasis, depth of tumor invasion, and advanced stage, whereas nuclear HuR expression was not correlated with any clinicopathological factors. Patients positive for cytoplasmic HuR expression had a cumulative 5-year survival rate of 25.3%, whereas it was 43.8% for patients negative for cytoplasmic HuR expression. In a multivariate analysis, cytoplasmic HuR expression was an independent prognostic factor, whereas nuclear positivity for HuR was not. Our results indicate that high cytoplasmic HuR expression is associated with positive lymph node metastasis, deep tumor invasion, high stage, and poor survival in ESCC. Thus, HuR is the first mRNA stability protein whose expression is associated with poor survival in esophageal cancer.

Direct sequencing and amplification refractory mutation system for epidermal growth factor receptor mutations in patients with non-small cell lung cancer.

Treatment with epidermal growth factor receptor (EGFR) tyrosine inhibitors (EGFR-TKIs) provides encouraging outcomes for advanced non-small cell lung cancer (NSCLC) patients with EGFR mutations. Pleural effusion is a common complication of NSCLC. We compared direct DNA sequencing and ADx Amplification Refractory Mutation System (ADx-ARMS) to detect EGFR mutations in malignant pleural effusion samples. We obtained 24 samples from pleural effusion fluid of NSCLC patients. Three common types of EGFR mutations were examined by direct sequencing and ADx-ARMS analysis. The sensitivity of the methods was compared and the relationship between EGFR mutations and response rates of the patients determined. In 14/24 patients, we detected EGFR mutations (58.3%) by ADx-ARMS, and in 10 samples (41.7%) by direct sequencing. In 6 samples, EGFR mutations were on exon 19, and in 8 samples, mutations were on exon 21 by ADx-ARMS. By contrast, we found EGFR mutations in 4 samples on exon 19, and in 6 samples on exon 21 by direct sequencing. Neither method showed mutations on exon 20. Among the 24 patients, there was 83.3% concordance for the methods. In 18/24 patients, gefitinib treatment was administered, including 10 patients with mutations who showed improved response compared to 8 of the wild-type patients (P<0.05). In conclusion, EGFR mutation analysis by ADx-ARMS was the most sensitive compared to direct sequencing, and provided more reliable EGFR mutation assessments. ADx-ARMS could be introduced into the clinical practice to identify NSCLC patients likely to benefit from TKI treatment, especially those with malignant pleural effusion.

Association study of single nucleotide polymorphisms in XRCC1 gene with risk of hepatocellular carcinoma in Chinese Han population.

Hepatocellular carcinoma (HCC) is one of the most frequently causing cancer-related deaths worldwide. Previous evidence suggests that the X-ray repair cross-complementing group 1 gene (XRCC1) is an important candidate gene for influencing the risk of HCC. The aim of this study was to assess the association of XRCC1 genetic polymorphisms with the risk of HCC in Chinese Han population. A total of 1314 subjects, including 651 HCC patients and 663 healthy controls, were enrolled in this case-control study. Two genetic variants (c.1254C>T and c.1517G>C) in XRCC1 gene were genotyped by created restriction site-polymerase chain reaction (CRS-PCR) and PCR-restriction fragment length polymorphism (PCR-RFLP) methods. Our data indicated that the allele and genotype frequencies of these two genetic variants were statistical difference in HCC cases and healthy controls. Association analyses suggested that these two genetic variants were statistically associated with the increased risk of HCC in all genetic models (for c.1254C>T, TT versus CC: OR = 2.30, 95% CI 1.61-3.28; CT versus CC: OR = 1.32, 95% CI 1.05-1.67; TT/CT versus CC: OR = 1.50, 95% CI 1.20-1.86; TT versus CT/CC: OR = 2.00, 95% CI 1.43-2.80; T versus C: OR = 1.47, 95% CI 1.25-1.73; for c.1517G>C, CC versus GG: OR = 1.90, 95% CI 1.34-2.69; GC versus GG: OR = 1.56, 95% CI 1.24-1.97; CC/GC versus GG: OR = 1.63, 95% CI 1.31-2.03; CC versus GC/GG: OR = 1.52, 95% CI 1.10-2.11; C versus G: OR = 1.45, 95% CI 1.23-1.70). The allele-T of c.1254C>T and allele-C of c.1517G>C genetic variants may contribute to HCC susceptibility in Chinese Han population.

Multiple functions of the RNA-binding protein HuR in cancer progression, treatment responses and prognosis.

The human embryonic lethal abnormal vision-like protein, HuR, is a member of the Hu family of RNA-binding proteins. Over the past decade, this ubiquitously expressed protein has been extensively investigated in cancer research because it is involved in the regulation of mRNA stability and translation in many cell types. HuR activity and function is associated with its subcellular distribution, transcriptional regulation, translational and post-translational modifications. HuR regulation of target mRNAs is based on the interaction between the three specific domains of HuR protein and one or several U- or AU-rich elements (AREs) in the untranslated region of target mRNAs. A number of cancer-related transcripts containing AREs, including mRNAs for proto-oncogenes, cytokines, growth factors, and invasion factors, have been characterized as HuR targets. It has been proposed that HuR has a central tumorigenic activity by enabling multiple cancer phenotypes. In this review, we comprehensively survey the existing evidence with regard to the diverse functions of HuR in caner development and progression. The current data also suggest that HuR might be a novel and promising therapeutic target and a marker for treatment response and prognostic evaluation.

Cytoplasmic HuR expression correlates with P-gp, HER-2 positivity, and poor outcome in breast cancer.

HuR is an ubiquitously expressed RNA-binding protein that stabilizes messenger RNA and regulates translation. This protein has been shown to play an important role in carcinogenesis and cancer progression. P-glycoprotein (P-gp) is the product of the multidrug resistance 1 gene, and the overexpression of P-gp induces multidrug resistance and represents a major obstacle in cancer chemotherapy. The purpose of this study was to determine the expression of HuR and P-gp in human breast cancer tissues and analyze the relationship between HuR or P-gp expression and the clinical-pathological variables and patient outcomes. Immunohistochemistry was used to determine HuR and P-gp expression in 82 human breast cancer tissues and 20 matched adjacent noncancerous tissues. Additionally, 16 benign breast tumor samples were used as controls. The overexpression of cytoplasmic HuR was found in breast cancer but not in the matched adjacent noncancerous tissues or benign breast tumors. The expression levels of cytoplasmic HuR were significantly associated with increased age, high nuclear grade, and the positive expression of the ER, PR, and HER-2/neu. HuR was also associated with the expression of P-gp protein. Furthermore, univariate analysis indicates that patients with high expression levels of cytoplasmic HuR or P-gp had significantly reduced survival compared to patients with low expression levels. A multivariate analysis showed that age at diagnosis, nuclear grade, and cytoplasmic HuR positivity were independent indicators for disease-free survival and overall survival in patients with breast cancer. In conclusion, cytoplasmic HuR expression detected by immunohistochemical staining is a negative prognostic indicator for survival in patients with breast cancer.

Effects of sulfated polysaccharide from Masson pine (Pinus massoniana) pollen on the proliferation and cell cycle of HepG2 cells.

AIM: To explore the inhibitory effect of sulfated polysaccharide from Masson pine (Pinus massoniana) pollen (SPPM60) on G2/M phase of human liver cancer HepG2 cells and its mechanism. METHODS: The proliferation rate of HepG2 cells was evaluated by methyl thiazolyl tetrazolium (MTT) colorimetric assay. The cycles of HepG2 cells were measured by flow cytometry when 200µg/ml concentration of SPPM60 was adopted, the expression of the genes related to cell cycle was detected by real-time PCR. RESULTS: SPPM60 inhibited the proliferation of HepG2 cells and the inhibition rate was elevated with increase of SPPM60 concentration. After treatment with 200µg/ml of SPPM60, the percentage of S phase cells was decreased, but that of G2/M phase was significantly increased (72h vs control: 32.96±0.33% vs 18.59±0.04%, 3.44±0.05% vs 18.30±0.08%, P<0.01). The results of real-time PCR showed that SPPM60 could down-regulate the mRNA levels of CDK1 and CyclinB (P<0.01), and up-regulate the expression of p53 and p21 (P<0.05). CONCLUSION: SPPM60 causes arrest of HepG2 cells at G2/M phase, and the mechanism is related to the down-regulation of CDK1 and CyclinB and up-regulation of p53 and p21 expression.

Lymphatic microvessel density and vascular endothelial growth factor-C and -D as prognostic factors in breast cancer: a systematic review and meta-analysis of the literature.

The use of lymphatic microvessel density (LVD) and pro-lymphangiogenic mediators as prognostic factors for survival in breast cancer remains controversial. We searched the electronic databases PubMed and EMBASE without language restrictions for relevant literature to aggregate the survival results. To be eligible, every study had to include the assessment of the LVD or the expression of vascular endothelial growth factor (VEGF)-C or -D in patients with breast cancer and provide a survival comparison, including disease-free survival (DFS) or overall survival (OS), according to the LVD, VEGF-C or VEGF-D status. Across all studies, 56.64 % of patients were considered to have a VEGF-C-positive tumor, and 65.54 % of patients had VEGF-D-positive tumors. High LVD had an unfavorable impact on DFS, with a pooled hazard ratio (HR) of 2.222 (95 % CI 1.579-3.126) and an OS with a HR of 2.493 (95 % CI 1.183-5.25). According to the different lymphatic makers, the subgroup HR in the D2-40 studies was 2.431 (95 % CI 1.622-3.644) for DFS and 4.085 (95 % CI 1.896-8.799) for OS. VEGF-C overexpression, as assessed by immunochemistry, was a prognostic factor for decreased DFS (HR 2.164; 95 % CI 1.256-3.729) and for decreased OS (HR 2.613; 95 % CI 1.637-4.170). VEGF-D overexpression was a significant although weak prognostic factor for DFS only when assessed by immunochemistry, with a HR of 2.108 (95 % CI 1.014-4.384). Our meta-analysis demonstrated that LVD, VEGF-C and VEGF-D could predict poor prognosis in patients with breast cancer. However, standardization of the assessment of LVD and for the expression of lymphangiogenesis factors is needed.

Clinical significance and role of lymphatic vessel invasion as a major prognostic implication in non-small cell lung cancer: a meta-analysis.

BACKGROUND: Lymphatic vessel invasion (LVI) exerts an important process in the progression and local spread of cancer cells. However, LVI as a prognostic factor for survival in non-small cell lung cancer (NSCLC) remains controversial. METHODOLOGY/PRINCIPAL FINDINGS: A meta-analysis of published studies from PubMed and EMBASE electronic databases was performed to quantity the effects of LVI on both relapse-free survival and overall survival for patients with NSCLC. Hazard ratios (HRs) with 95% confidence intervals (95% CIs) were used to assess the strength of these effects. This meta-analysis included 18,442 NSCLC patients from 53 eligible studies. LVI appeared in 32.1% (median; range, 2.8% to 70.9%) of tumor samples. In all, patients with LVI were 2.48 times more likely to relapse by univariate analysis (95% CI: 1.92-3.22) and 1.73 times by multivariate analysis (95% CI: 1.24-2.41) compared with those without LVI. For the analyses of LVI and overall survival, the pooled HR estimate was 1.97 (95% CI: 1.75-2.21) by univariate analysis and 1.59 (95% CI: 1.41-1.79) by multivariate analysis. Multivariate analysis showed a risk was 91% higher for recurrence (HR =1.91, 95% CI: 1.14-2.91) and 70% higher for mortality (HR=1.70, 95% CI: 1.38-2.10) in LVI-positive I stage patients compared with LVI-negative I stage patients. Subgroup analyses showed similar significant adjusted risks for recurrence and death in adenocarcinomas, and a significant adjusted risk for death in studies that utilized elastic staining with or without immunohistochemistry in defining LVI. CONCLUSIONS/SIGNIFICANCE: The present study indicates that LVI appears to be an independent poor prognosticator in surgically managed NSCLC. NSCLC patients with LVI would require a more aggressive treatment strategy after surgery. However, large, well-designed prospective studies with clinically relevant modeling and standard methodology to assess LVI are required to address some of these important issues.