RIPK3 expression as a potential predictive and prognostic marker in metastatic colon cancer.

BACKGROUND: Colorectal cancer is one of the primary causes of cancer-related deaths and 5-fluorouracil (5-FU) therapy remains the cornerstone of treatment in these patients. Resistance to 5-FU represents a major obstacle; therefore, finding new predictive and prognostic markers is crucial for improvement of patient outcomes. Recently a new type of programmed cell death was discovered-necroptosis, which depends on receptor interacting protein 3 (RIPK3). Preclinical data showed that necroptotic cell death is an important effector mechanism of 5-FU-mediated anticancer activity. PURPOSE: To investigate the predictive and prognostic performance of RIPK3 expression in primary tumors. METHODS: Colon cancer patients (n=74) with metastatic stage were included in this retrospective study and all were treated with first-line 5-FU based chemotherapy. Immunohistochemical staining was performed. RESULTS: The progression free survival for the low expression group of RIPK3 was 5.6 months (95% CI, 4.4-6.8) vs 8.4 months (95% CI, 6.4-10.3) of the group with high expression (p=0.02). Moreover, patients with high expression of RIPK3 were associated with lower risk of disease progression HR 0.61 (95% CI, 0.38-0.97; p=0.044). Patients with high expression levels of RIPK3 also had significantly longer mean overall survival (OS) of 29.3 months (95% CI, 20.8-37.8) as compared with those with low expression: 18.5 months (95% CI, 15.06-21.9) (p= 0.036). In addition, univariate analysis showed that high level of RIPK3 expression was associated with a longer OS HR 0.59 (95% CI, 0.35-0.98; p=0.044). CONCLUSIONS: This study suggests that expression of RIPK3 in primary tumors of metastatic colon cancer patients should be further investigated for its potential as a promising predictive and prognostic marker.

Dual inhibition of Met kinase and angiogenesis to overcome HGF-induced EGFR-TKI resistance in EGFR mutant lung cancer.

Acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) is a serious problem in the management of EGFR mutant lung cancer. We recently reported that hepatocyte growth factor (HGF) induces resistance to EGFR-TKIs by activating the Met/PI3K pathway. HGF is also known to induce angiogenesis in cooperation with vascular endothelial growth factor (VEGF), which is an important therapeutic target in lung cancer. Therefore, we hypothesized that dual inhibition of HGF and VEGF may be therapeutically useful for controlling HGF-induced EGFR-TKI-resistant lung cancer. We found that a dual Met/VEGF receptor 2 kinase inhibitor, E7050, circumvented HGF-induced EGFR-TKI resistance in EGFR mutant lung cancer cell lines by inhibiting the Met/Gab1/PI3K/Akt pathway in vitro. HGF stimulated VEGF production by activation of the Met/Gab1 signaling pathway in EGFR mutant lung cancer cell lines, and E7050 showed an inhibitory effect. In a xenograft model, tumors produced by HGF-transfected Ma-1 (Ma-1/HGF) cells were more angiogenic than vector control tumors and showed resistance to gefitinib. E7050 alone inhibited angiogenesis and retarded growth of Ma-1/HGF tumors. E7050 combined with gefitinib induced marked regression of tumor growth. Moreover, dual inhibition of HGF and VEGF by neutralizing antibodies combined with gefitinib also markedly regressed tumor growth. These results indicate the therapeutic rationale of dual targeting of HGF-Met and VEGF-VEGF receptor 2 for overcoming HGF-induced EGFR-TKI resistance in EGFR mutant lung cancer.

Neutrophil to lymphocyte ratio as a potential predictive marker for treatment with pembrolizumab as a second line treatment in patients with non-small cell lung cancer.

The aim of this multicentric retrospective study is to evaluate the predictive and prognostic performance of neutrophil to lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR) and their dynamics in patients with non-small cell lung cancer (NSCLC) treated with pembrolizumab as a second line. Patients with metastatic NSCLC (n = 119), whose tumors expressed programmed death-ligand 1 (PD-L1) ≥ 1%, were retrospectively analyzed between Apr 2017 and Apr 2019. All patients received platinum-containing chemotherapy as a first line treatment. Pre-treatment NLR was calculated by dividing the number of neutrophils by the number of lymphocytes in peripheral blood before the first pembrolizumab infusion. Progression free survival (PFS) and overall survival (OS) was compared by Kaplan-Meier method and Cox Proportional Hazard model. Patients with NLR > 5 before immunotherapy showed significantly shorter mean PFS of 6.86 months (95% CI: 5.81-7.90) as compared to those with NLR ≤ 5 of 18.82 months (95% CI: 15.87-21.78) (long rank test p < 0.001). Furthermore in the multivariate analysis, only NLR > 5 was an independent predictive factor for shorter PFS (HR: 4.47, 95% CI: 2.20-9.07, p < 0.001). In multivariate analysis, presence of bone metastases (HR: 2.08, 95% CI: 1.10-4.94, p = 0.030), NLR > 5 before chemotherapy (HR: 8.09, 95% CI: 2.35-27.81, p = 0.001) and high PLR before chemotherapy (HR: 2.81, 95% CI: 1.13-6.97, p = 0.025) were found to be independent negative prognostic factors for poor OS. Our data suggests that NLR ≤ 5 is a potential predictive marker, which may identify patients appropriate for immunotherapy as a second line treatment.

Quantitative Analysis of Tumor-associated Tissue Eosinophilia in Recurring Bladder Cancer.

The relatively high incidence of recurrence of bladder cancer is a serious problem in clinical practice. At present, there are no objective microscopic criteria for evaluation of the tendency for local relapse. Besides the phenotypic properties of the tumor parenchymal cells, possible signs in regard to recurrence could also be derived from the peculiarities of the tumor stroma. The stromal reaction, manifested by inflammatory infiltration in the tumor is considered to influence the biological behavior of tumors. Also, a relationship has been reported between the number of eosinophils and the survival of patients. The aim of the present study is to analyze tumor-associated tissue eosinophilia (TATE) and to compare TATE density in the initial foci of age and gender-matched 156 cases of recurrent and non-recurrent bladder cancers; the tumors that have relapsed within six months after removal and contained statistically significant greater numbers of eosinophils in primary cancer sites. These results suggest that TATE may be one of the probable prognostic signs for local relapse of urothelial cancer.

Correlation Between Ki-67 Index, World Health Organization Grade and Patient Survival in Glial Tumors With Astrocytic Differentiation.

Background Glioblastoma multiforme (GBM) is a class IV astrocytic tumor, the most malignant of the four groups of World Health Organization (WHO) tumors with astrocytic differentiation. Aim The aim of this study was to estab-lish whether a correlation exists between the Ki-67 index of tumors with astrocytic differentiation, WHO grade, and patient survival. Materials and methods A retrospective non-clinical approach to patient selection was chosen for the aim of the study. A total of 47 patients diagnosed and treated for CNS tumors with astrocytic differentiation in the St. Marina University Hospital, Varna, Bulgaria, from September 2012 to July 2016 were retrospectively included into the study cohort. The cases were tested for their immunohistochemistry (IHC) reaction with Ki-67 after their original Hematoxylin and Eosin and IHC slides were reviewed by a single author and blind coded. The Ki-67 positivity index of the nuclei was estimated after digitalization of the slides and calculated by the ImmunoRatio automated count-ing tool. The individual Ki-67 index and patient survival of each case were statistically compared. Results The histopathological groups, after the blind Ki-67 index automated calculation was carried out, revealed no WHO grade I, two WHO grade II samples, four WHO grade III samples and 41 WHO grade IV cases, and these were included in the analysis. The two samples of WHO grade II astrocytic tumors had a mean Ki-67 index of 25%; however, they comprised tumors with an individual index of 43% and 7%, both individual values with a highly unlikely index for this group. The four samples of WHO grade III had a mean Ki-67 index of 4%, standard deviation ±2.16 (p>0.05), with the lowest index being 1% and the highest one being 6%. Both WHO grade II and III did not include enough samples to allow for a proper statistical analysis of patient survival. The 41 GBM cases had a mean Ki-67 index of 17.34%, standard deviation ±10.79 (p>0.05). Statistical analysis of the Ki-67 index divid-ed dichotomously into two groups and patient survival revealed that cases with a high Ki-67 index had no significant difference in survival when compared to those with low expression. Conclusions Based on the reported results, the mean Ki-67 percentage of positive nuclei in GBM tumor sam-ples cannot be used to estimate the survival of patients. However, Ki-67 remains a valuable IHC pathological tool.

Role of the pretreatment 18F-fluorodeoxyglucose positron emission tomography maximal standardized uptake value in predicting outcomes of colon liver metastases and that value's association with Beclin-1 expression.

The current study sought to evaluate the predictive and prognostic performance of the maximum standardized uptake value (SUVmax) prior to treatment in 43 patients with colon cancer and unresectable liver metastases. Patients with colon cancer who underwent 18F-FDG-PET/computed tomography (CT) scans for staging before the start of first-line 5-fluorouracil-based chemotherapy were retrospectively analyzed. Expression of Beclin-1 in cancer cells was evaluated in primary tumors using immunohistochemical staining. The pretreatment SUVmax for liver metastases was not able to predict progression-free survival but was significantly associated with poorer overall survival, with a hazard ratio of 2.05 (95 % CI, 1.016-4.155). Moreover, a negative correlation was noted between SUVmax and expression of a marker of autophagy - Beclin-1 (rho = -0.42, p = 0.006). This suggests that the pretreatment SUVmax in 18F-FDG PET/CT is a useful tool to help predict survival outcome in patients with colon cancer and unresectable liver metastases and may significantly distinguish between patients with low and high levels of Beclin-1 expression (AUC = 0.809, 95% CI: 0.670-0.948, p = 0.001).

Serum levels of RIPK3 and troponin I as potential biomarkers for predicting impaired left ventricular function in patients with myocardial infarction with ST segment elevation and normal troponin I levels prior percutaneous coronary intervention.

The current study examined the serum levels of receptor-interacting protein kinase 3 (RIPK3) in 51 patients with New York Heart Association (NYHA) class III-IV heart failure, 53 patients with myocardial infarction with ST elevation (STEMI), and 19 healthy subjects serving as a control group. An enzyme-linked immunoadsorbent assay (ELISA) was used to measure the levels of RIPK3 expression in serum. The area under the receiver operating characteristic curve (AUC) was then used to evaluate the predictive performance of RIPK3 and troponin I in patients with STEMI. In patients with normal levels of troponin I prior to percutaneous coronary intervention (PCI), serum levels of RIPK3 and troponin I after PCI were sufficient to differentiate patients with a preserved left ventricular ejection fraction (LVEF) from those with impaired left ventricular function after PCI (AUC = 0.780 (95% CI: 0.565-0.995, p = 0.043) with a sensitivity of 76.9% and a specificity of 71.4% vs. AUC = 0.735 (95% CI: 0.530-0.941, p = 0.038) with a sensitivity of 88.2% and a specificity of 63.6% at the optimal cutoff values, respectively). Moreover, elevated levels of troponin I after PCI were associated with an increased risk of an LVEF < 50% prior to discharge (odds ratio, 1.014; 95 % CI, 1.001 to 1.027; p = 0.03), while elevated levels of RIPK3 were not associated with such a risk. The current findings suggest that in patients with normal levels of troponin I prior to PCI, serum levels of RIPK3 and troponin I can serve as a potential marker to identify patients with a decreased LVEF, thus possibly allowing an early shift to more intensive therapy.

Serum expression levels of miR-17, miR-21, and miR-92 as potential biomarkers for recurrence after adjuvant chemotherapy in colon cancer patients.

The present study examined whether miR-17, miR-21, miR-29a, and miR-92 that are dysregulated in colon cancer (CC) can serve as potential predictive markers for relapse of disease after radical surgery and adjuvant chemotherapy. Real-time reverse transcription quantitative polymerase chain reaction was used to measure the expression levels of the miRNAs in serum samples from 37 patients with CC and 7 healthy individuals, tested as a control group. The area under the receiver operating characteristic curve (AUC) was then used to evaluate the predictive performance of the four miRNAs alone or in combination and compare it with carcinoembryonic antigen. The expression of miR-17, miR-21 and miR-92 were significantly higher in serum of patients with disease relapse. The AUCs for miR-17, miR-21, miR-92 for Nx patients were 0.844, 0.948, and 0.935, respectively (p < 0.05). Combining the four miRNAs for stage III patients increased the diagnostic performance, yielding an AUC of 0.881, with a sensitivity of 83.3% and a specificity of 85.7% (p < 0.05). Our study suggests that the expression levels of serum miR-21, miR-17, and miR-92 in patients with CC who underwent radical surgery and adjuvant chemotherapy may have diagnostic value for differentiating between recurred and non-recurred patients.

Transient PI3K inhibition induces apoptosis and overcomes HGF-mediated resistance to EGFR-TKIs in EGFR mutant lung cancer.

PURPOSE: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI), such as gefitinib and erlotinib, show favorable response to EGFR mutant lung cancer. However, the responders acquire resistance almost without exception. We recently reported that hepatocyte growth factor (HGF) induces EGFR-TKI resistance by activating MET that restores downstream mitogen activated protein kinase (MAPK)/extracellular signal regulated kinase (ERK)1/2 and phosphoinositide 3-kinase (PI3K)/Akt signaling. The purpose of this study was to determine whether inhibition of PI3K, a downstream molecule of both EGFR and MET, could overcome HGF-mediated EGFR-TKI resistance in EGFR mutant lung cancer cells PC-9 and HCC827. EXPERIMENTAL DESIGN: We explored therapeutic effect of a class I PI3K inhibitor PI-103 on HGF-induced EGFR-TKI resistance in vitro and in vivo. RESULTS: Unlike gefitinib or erlotinib, continuous exposure with PI-103 inhibited proliferation of PC-9 and HCC827 cells, even in the presence of HGF. On the other hand, in gefitinib-resistant xenograft model by using PC-9 cells mixed with HGF high producing fibroblasts, PI-103 monotherapy did not inhibit tumor growth. However, PI-103 combined with gefitinib successfully regressed gefitinib-resistant tumor. In vitro experiments by considering short half-life of PI-103 reveal that transient exposure of PI-103 combined with gefitinib caused sustained inhibition of Akt phosphorylation, but not ERK1/2 phosphorylation, resulting in induction of tumor cell apoptosis even in the presence of HGF. CONCLUSIONS: These results indicate that transient blockade of PI3K/Akt pathway by PI-103 and gefitinib could overcome HGF-mediated resistance to EGFR-TKIs by inducing apoptosis in EGFR mutant lung cancer.