Signal Transduction In Tumor Cells Research Articles

Research progress on anti-tumor mechanism of TAOK kinases

Thousand and one amino-acid protein kinases(TAOKs), as a key member of the mitogen-activated protein kinase (MAPK) cascade, has recently attracted widespread attention in the field of anti-cancer research. There are three members of this subfamily: TAOK1, TAOK2, and TAOK3. Studies have shown that members of the TAOK family participate in regulating cell proliferation, apoptosis, migration, and invasion through various pathways, thereby playing an important role in tumorigenesis and progression. This review summarizes the functions of TAOK kinases in tumor cell signal transduction, cell cycle regulation, and the tumor microenvironment, with a particular emphasis on its potential as a target for anti-cancer drugs. Future research will further elucidate the specific mechanisms of action of TAOK kinase in different types of tumors and explore its clinical application prospects.

A prognosis marker MUC1 correlates with metabolism and drug resistance in bladder cancer: a bioinformatics research

BackgroundMUC1 is a type I transmembrane protein that plays an important role in tumor cell signal transduction. Although current studies have shown that MUC1 is upregulated in bladder cancer (BC), the specific mechanism is still unclear.MethodsWe performed expression analysis, gene set enrichment analysis, survival analysis, immune infiltration analysis, drug sensitivity analysis, and metabolism-related gene expression analysis on TCGA-BLCA, GES31684 and GSE13507.ResultsThe expression of MUC1 in the tumor and lymphatic metastasis positive samples was significantly increased. Genes related to MUC1 expression were significantly enriched in immune response, ribosomes, exosomes, and energy metabolism. The results of the immune infiltration analysis showed that M1 macrophages in BC with high MUC1 expression were significantly decreased. Expression of MUC1 increases drug resistance in BC patients. In addition, MUC1 increases glycolysis, glucose uptake, and lactate production by inducing metabolic reprogramming.ConclusionMUC1 has a significant effect on the metabolism and immune cell infiltration of BC, which may be the cause of increased drug resistance, and can be used as a molecular target for the diagnosis and treatment of BC.

A natural xanthone suppresses lung cancer growth and metastasis by targeting STAT3 and FAK signaling pathways

BackgroundNonsmall-cell lung cancer (NSCLC) is one of the most common malignant tumors, and the current drugs have not achieved ideal therapeutic effects. The abnormal activation of STAT3 and FAK signal transduction in tumor cells is highly correlated with their proliferation and migration ability. Therefore, bioactive compounds that can inhibit STAT3 and FAK activation have the potential to become agents to treat NSCLC. PurposeThis study aims to discover new antitumor compounds from Garcinia xipshuanbannaensis and investigate the molecular mechanism by which they inhibit lung cancer proliferation and metastasis in vivo and in vitro, all of which may lead to obtainment of a potential antitumor agent. MethodsXipsxanthone H was obtained by various chromatography methods (including silica gel, medium pressure liquid chromatography (MPLC), and preparative high-performance liquid chromatography (HPLC)). 1D and 2D nuclear magnetic resonance (NMR) spectra were used to analyze the structure. Cell viability and wound healing assays were employed to detect changes in the proliferation and migration of cancer cells. Cell cycle and apoptosis were analyzed by flow cytometry. The protein expression of STAT3 and FAK signaling pathways affected by xipsxanthone H was determined by Western blotting. The zebrafish model was used to evaluate the in vivo effects of xipshantone H on tumor proliferation and metastasis. Molecular docking was utilized to explore the interaction between xipsxanthone H and STAT3. Cellular thermal shift assays (CETSAs) were employed to explore the possible target of xipsxanthone H. ResultsThe novel compound xipsxanthone H was purified and characterized from G. xipshuanbannaensis. Xipsxanthone H exhibited strong anti-proliferation activity in a variety of tumor cell lines. In addition to inducing reactive oxygen species (ROS) production and arresting the cell cycle, mechanistic studies demonstrated that xipsxanthone H suppressed STAT3 and FAK phosphorylation and regulated the downstream protein expression of the STAT3 and FAK signaling pathways. The in vivo studies using the zebrafish model revealed that xipsxanthone H inhibited tumor proliferation, metastasis, and angiogenesis. ConclusionsA new xanthone was obtained from G. xipshuanbannaensis, and this compound had the property of inhibiting tumor proliferation and metastasis by targeting STAT3 and FAK signaling pathways in NSCLC. These findings suggested that xipsxanthone H might be a potential candidate agent for NSCLC treatment.

Functional signaling test identifies HER2 negative breast cancer patients who may benefit from c-Met and pan-HER combination therapy

BackgroundResearch is revealing the complex coordination between cell signaling systems as they adapt to genetic and epigenetic changes. Tools to uncover these highly complex functional linkages will play an important role in advancing more efficacious disease treatments. Current tumor cell signal transduction research is identifying coordination between receptor types, receptor families, and transduction pathways to maintain tumor cell viability despite challenging tumor microenvironment conditions.MethodsIn this report, coactivated abnormal levels of signaling activity for c-Met and HER family receptors in live tumor cells were measured by a new clinical test to identify a subpopulation of breast cancer patients that could be responsive to combined targeted therapies. The CELsignia Multi-Pathway Signaling Function (CELsignia) Test uses an impedance biosensor to quantify an individual patient’s ex vivo live tumor cell signaling response in real-time to specific HER family and c-Met co-stimulation and targeted therapies.ResultsThe test identified breast tumors with hyperactive HER1, HER2, HER3/4, and c-Met coordinated signaling that express otherwise normal amounts of these receptors. The supporting data of the pre-clinical verification of this test included analyses of 79 breast cancer patients’ cell response to HER and c-Met agonists. The signaling results were confirmed using clinically approved matching targeted drugs, and combinations of targeted drugs in addition to correlative mouse xenograft tumor response to HER and c-Met targeted therapies.ConclusionsThe results of this study demonstrated the potential benefit of a functional test for identifying a subpopulation of breast cancer patients with coordinated abnormal HER and c-Met signaling for a clinical trial testing combination targeted therapy.4ydEpWcWmteHkpnJtfDHYnVideo

Emerging roles of protein palmitoylation and its modifying enzymes in cancer cell signal transduction and cancer therapy.

Protein palmitoylation is an increasingly investigated form of post-translational lipid modification that affects protein localization, accumulation, secretion and function. Recently, emerging findings have revealed that protein palmitoylation is crucial for many tumor-related signaling pathways, such as EGFR, RAS, PD-1/PD-L1 signaling, affecting the occurrence, progression and therapeutic response of tumors. Protein palmitoylation and its modifying enzymes, including palmitoylases and depalmitoylases, are expected to be new targets for effective tumor treatment. Recognizing the significance of palmitoylation modification on protein stability, localization and downstream signal regulation, this review focuses on the regulatory roles of protein palmitoylation and its modifying enzymes in tumor cell signal transduction, aiming to bring new ideas for effective cancer prevention and treatment.

Abstract PO-089: Morphokinetic single-cell analysis and machine learning as a tool to characterize breast cancer cell motility and response to therapy

Abstract Triple-negative breast cancer (TNBC) is a type of aggressive breast cancer lacking the expression of estrogen receptors (ER), progesterone receptors (PR), and human epidermal growth factor receptor-2 (HER-2). TNBC patients have a high propensity for distant metastasis and limited treatment options. Cancer cell motility and invasion are fundamental steps in metastasis. MET, a tyrosine kinase receptor, and its ligand, Hepatocyte Growth Factor/Scatter Factor (HGF/SF), induce specific signal transduction in tumor cells leading to cell motility and metastasis. MET's induced tumorigenesis and metastatic processes make it an ideal target for anti-cancer therapy. We characterized the motility patterns of m-Cherry labeled TNBC cells (BT-549, MDA-MB-231) and ER-positive cells (MCF7) subjected to time-lapse fluorescence wound healing assay. We also studied the effect of MET inhibition, chemotherapy, and their combined impact on breast cancer motility. Time-lapse images were subjected to commercial packages for segmentation and tracking - allowing us to extract morphokinetic (MK) information at a single-cell resolution. To better understand cell motility patterns, we developed the Tool for Analysis of Single-Cell (TASC) infrastructure, which utilizes unsupervised machine learning methods to demonstrate a high-dimensional feature set at a single-cell resolution. We compared our experimental MK results to simulations of three modified classical physical models: Lévy flight (LF), fractal Brownian motion, and random walk (RW) with or without a back-propagating wave (BPW). TASC analysis demonstrated a fundamental difference between ER-positive and TNBC cells: both cell types contained two subpopulations harboring similar MK characteristics. TNBC cells presented an additional subpopulation characterized by 1) dominantly increasing cumulative kinetics values (mean square displacement - MSD), 2) highest temporal wave values (velocity starting-time), and 3) a decrease in non-cumulative kinetics values. The combined treatment of TNBC cells with MET inhibition and chemotherapy eliminates this high-motility group. Cell motility models and motion simulations were compared using Wasserstein's analysis. Untreated TNBC cells showed high similarities to RW with BPW simulations, while MET-activated TNBC cells transformed into LF without BPW simulations, indicating more aggressive behavior. TASC analysis revealed a previously undefined high motility subpopulation of TNBC that was eradicated by combining MET inhibition and chemotherapy. The primary benefit of TASC is the ability to cluster and characterize similar behaviors across different motility models while still detecting subtle changes within each one. MK analysis combined with TASC can lead to discovering novel targets for precise therapy. Moreover, this infrastructure may determine patient tumor susceptibility to chemotherapy and biological treatments and operate as an analytical tool for personalized, targeted therapy. Citation Format: Ilan Tsarfaty, Or Megides, Ohad Doron, Hagar Alaloof, Ori Moskowitz, Judith Horev. Morphokinetic single-cell analysis and machine learning as a tool to characterize breast cancer cell motility and response to therapy [abstract]. In: Proceedings of the AACR Virtual Special Conference on Artificial Intelligence, Diagnosis, and Imaging; 2021 Jan 13-14. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(5_Suppl):Abstract nr PO-089.

Abstract 535: T-cell recruitment tumor lysis via a novel c-MET/CD3 bispecific antibody

Abstract Anti-receptor tyrosine kinases (RTKs) therapy has been used for the treatment of a number of solid tumors in clinic. C-MET, also known as the hepatocyte growth factor receptor (HGFR), is a representative member of the RTK family. C-MET overexpression is commonly observed in tumor biopsies and associated with poor patient survival. Various kinase inhibitor and monoclonal antibody (e.g. Emibetuzumab and Onartuzumab) have been developed to block the HGF/c-MET interactions, however, those kinase-inhibiting-based therapeutics have shown limited success in clinical trials. There is an urgent need to develop new treatments targeting c-MET to overcome these limitations. Bispecific antibodies that recruit immune cells to the tumor microenvironment have been considered as effective therapeutic candidates. An example of this design strategy is the FDA-approved bispecific antibody blinatumomab. In this study, we have constructed an asymmetric bispecific antibody, which binds both c-MET and CD3, using a novel platform that is monovalent to epitope. Our c-MET/CD3 bispecific antibody showed potent T-cell mediated tumor cells killing in vitro. Meanwhile, this antibody inhibited the phosphorylation of c-MET and downstream signal transduction in tumor cells due to its monovalent nature. In c-MET-overexpressing lung cancer and ovarian cancer xenograft models, c-MET/CD3 bispecific antibody inhibited tumor growth and increased the proportion of activated CD56+ tumor infiltrating lymphocytes. Moreover, in vivo combination therapy with FDA-proved PD-1/PD-L1 antibodies showed higher tumor inhibition than monotherapy in above two tumor models. Collectively, the novel c-MET/CD3 bispecific antibody can provide immunotherapy for c-MET-overexpressing tumors which conventional antibody or small molecular inhibitor might not. These findings also support the immunotherapeutic effects on combination of T-cell dependent bispecific antibody and immune checkpoint blockade. Citation Format: Lei Huang, Kun Xie, Ruiqin Wang, Hua Gu, Jianmin Fang. T-cell recruitment tumor lysis via a novel c-MET/CD3 bispecific antibody [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 535.

Suppression of c-Met-Overexpressing Tumors by a Novel c-Met/CD3 Bispecific Antibody.

IntroductionOverexpression of c-Met, or hepatocyte growth factor (HGF) receptor, is commonly observed in tumor biopsies and often associated with poor patient survival, which makes HGF/c-Met pathway an attractive molecular target for cancer therapy. A number of antibody-based therapeutic strategies have been explored to block c-Met or HGF in cancers; however, clinical efficacy has been very limited, indicating that blockade of c-Met signal alone is not sufficient. Thus, an alternative approach is to develop an immunotherapy strategy for c-Met-overexpressing cancers. c-Met/CD3 bispecific antibody (BsAb) could bridge CD3-positive T lymphocytes and tumor cells to result in potent tumor cell killing.Materials and MethodsA bispecific antibody, BS001, which binds both c-Met and CD3, was generated using a novel BsAb platform. Western blotting and T cells-mediated killing assays were utilized to evaluate the BsAb’s effects on cell proliferation, survival and signal transduction in tumor cells. Subcutaneous tumor mouse models were used to analyze the in vivo anti-tumor effects of the bispecific antibody and its combination therapy with PD-L1 antibody.ResultsBS001 showed potent T-cell mediated tumor cells killing in vitro. Furthermore, BS001 inhibited phosphorylation of c-Met and downstream signal transduction in tumor cells. In A549 lung cancer xenograft model, BS001 inhibited tumor growth and increased the proportion of activated CD56+ tumor infiltrating lymphocytes. In vivo combination therapy of BS001 with Atezolizumab (an anti-programmed cell death protein1-ligand (PD-L1) antibody) showed more potent tumor inhibition than monotherapies. Similarly, in SKOV3 xenograft model, BS001 showed a significant efficacy in tumor growth inhibition and tumor recurrence was not observed in more than half of mice treated with a combination of BS001 and Pembrolizumab.Conclusionc-Met/CD3 bispecific antibody BS001 exhibited potent anti-tumor activities in vitro and in vivo, which was achieved through two distinguished mechanisms: through antibody-mediated tumor cell killing by T cells and through inhibition of c-Met signal transduction.

Overexpression of LGR-5 as a Predictor of Poor Outcome in Patients with Hepatocellular Carcinoma.

Hepatocarcinogenesis and distant metastasis pose major challenges for physicians. They are regulated by several genes, such as AKT, JUK, Wnt, and P53, and their expression activates several important processes such as cell proliferation, migration, motility, and interaction in the microenvironment. The leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR-5) is a novel biomarker, particularly in stem cells, and is involved in embryogenesis, tumor development, and tumor cell signal transduction. Here, we investigated LGR-5 expression using immunohistochemistry and analyzed the correlation between clinical features and prognosis in patients with hepatocellular carcinoma (HCC). We found that LGR-5 expression was higher in tumor tissues than in normal liver tissues, and that high LGR-5 expression possibly favored poor outcomes in HCC, especially in well/moderate differentiation grade, hepatitis C virus (HCV)-negative, and hepatitis B virus (HBV)-positive groups. Thus, the LGR-5 marker is suggested to be a routine biomarker for poor prognosis, thereby providing a platform for anti-LGR-5-targeted therapy in the future.

Beta 2-microglobulin and tumors

Beta 2-microglobulin (β2-MG) has low expression levels in tissues and cells of normal organisms, but its expression levels is increasing in tumor. The main function of β2-MG is to regulate signal transduction in tumor cells and promote tumor cell growth and angiogenesis. High level expression of β2-MG is associated with the occurrence, progression, metastasis and prognosis of human tumors and can serve as an independent indicator of tumor prognosis. It has been confirmed in a variety of tumor cells that agent targeting β2-MG is expected to become a new generation of targeted drugs. Key words: Beta 2-microglobulin; Neoplasms; Prognosis

Therapeutic effects of tyroservatide on metastasis of lung cancer and its mechanism affecting integrin-focal adhesion kinase signal transduction.

Tyroservatide (YSV) can inhibit the growth and metastasis of mouse lung cancer significantly. This study investigated the therapeutic effects of tripeptide YSV on metastasis of human lung cancer cells and explored its possible mechanism that affects integrin–focal adhesion kinase (FAK) signal transduction in tumor cells. YSV significantly inhibited the adhesion and the invasion of highly metastatic human lung cancer cell lines 95D, A549, and NCI-H1299. In addition, YSV significantly inhibited phosphorylation of FAK Tyr397 and FAK Tyr576/577 in the 95D, A549, and NCI-H1299 human lung cancer cells in vitro. And the mRNA level and protein expression of FAK in these human lung cancer cells decreased at the same time. YSV also significantly inhibited mRNA and protein levels of integrin β1 and integrin β3 in the 95D, A549, and NCI-H1299 human lung cancer cells. Our research showed that YSV inhibited adhesion and invasion of human lung cancer cells and exhibited therapeutic effects on metastasis of lung cancer.

Abstract 1213: Comparing protein pathway activation mapping portraits between gliobastoma patient-matched primary tumor, xenografts and neurospheres: implications for precision medicine

Abstract Background: Most current targeted therapeutics are directed against the activated protein kinase-driven signaling pathway architecture. While guiding therapy selection based on protein phosphorylation is biochemically rational given these proteins represent the direct drug targets it is not certain which protein networks/signaling protein targets are the causal driver of any patient's tumor. Thus, the availability of a system that recapitulates the human in vivo setting and allows for credentialing and identification of which activated signaling networks are causally necessary for tumor viability would be extremely important. Methods: To test two such systems and determine how they resemble the cell signaling architecture of the human primary tumor we compared eight snap-frozen laser capture microdissected (LCM) primary glioblastoma multiforme (GBM) samples with tumor-matching LCM mouse xenografts as well as matching neurospheres (NP). Reverse phase protein microarrays were utilized wherein the total level or phosphorylated level of 70 key signal transduction proteins known to be the direct targets for a number of important targeted therapies and implicated in GBM tumor biology were measured. Results: Overall, the signaling architecture of the xenografts more closely resembled the matched primary LCM tumors in this study set. However, unsupervised hierarchical clustering of the data revealed instances where pathway information such as total c-MET, phospho RAF (S259), phospho GSK3αβ (S21/9) and total EGFR was retained across matched LCM primary tumor, xenografts and NP. In addition there were a number of instances where the signaling architecture of the primary human tumor did not resemble the matched NP and xenograft signature. Conclusion: These pilot data indicate that 3D cell culture conditions may effectively recapitulate in vivo signaling network activation in certain instances, although these preliminary results also suggest that xenografts more closely resemble the cell architecture of matching primary GBM tumors. These data highlight the importance and impact of the microenvironment and culture conditions on tumor cell signal transduction network activation. However, two caveats to this approach remain: (a) drug studies in GBM xenografts and NP are limited to those targeting signaling pathways that are faithfully reproduced from the human in vivo setting and (b) the time required to develop a patient matched xenograft model for individualized drug testing may be too long for a fast-progressing disease such as GBM. Citation Format: Claudius Mueller, Ana C. deCarvalho, Tom Mikkelsen, Laila Poisson, Valerie Calvert, Andrew Borgman, David M. Cherba, Mary E. Winn, Emanuel F. Petricoin. Comparing protein pathway activation mapping portraits between gliobastoma patient-matched primary tumor, xenografts and neurospheres: implications for precision medicine. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1213. doi:10.1158/1538-7445.AM2014-1213

Quantitative proteomic profiling of tumor cell response to telomere dysfunction using isotope-coded protein labeling (ICPL) reveals interaction network of candidate senescence markers

Telomerase inhibition causes progressive telomere shortening and cellular senescence, which constitutes a universal barrier to tumor growth and therefore an attractive target for tumor therapy. To expand our previous studies, we investigated the global effects of telomere dysfunction on the proteome of tumor cells in order to find novel senescence biomarkers. Telomerase-deficient HCT-116 cell clones were analyzed by a quantitative proteomic approach using isotope-coded protein labeling (ICPL) and nanoflow-HPLC-MS/MS. Stringent reduction of the extensive proteomic data from this tumor cell model revealed a list of 59 markers including proteins identified in our former studies and a number of novel proteins involved in tumorigenesis and metastasis such as SFN, S100A4, ANXA2, and LGALS1. A loss of the chromatin protein HMGB2 was demonstrated not only in various telomerase-inhibited clones of different tumor cell lines, but also in normal human fibroblasts undergoing replicative senescence and in aging telomerase knockout mice. Impressively, a coherent and dense network of protein-protein interactions for the bulk of the markers and their implementation in signaling pathways involving key regulators for tumorigenesis were revealed. These results have an impact on the understanding of telomere- and senescence-related signal transduction in tumor cells in consideration of the general lack of senescence markers. Induction of cellular senescence constitutes a potent concept for tumor therapy which interferes with immortalization and additional hallmarks of cancer. The application of a powerful quantitative proteomic approach using isotope-coded protein labeling to an approved model for senescence represented by telomerase inhibited tumor cells led to the identification of novel candidate biomarkers for telomere dysfunction and replicative senescence. Thereby, the identified markers not only fit in the context of the investigated processes with a relevance for additional hallmarks of cancer but are also involved in a strong interaction network and integrated in canonical pathways centered around key cancer-relevant proteins. These potential markers alone or in combination will significantly extend the view on telomere-associated signal transduction in tumor cells and contribute to the field of cellular senescence and aging in consideration of the general lack of biomarkers in this regard.

Abstract 3498: Functional profiling of Champions TumorGraft™ models from metastatic melanoma patients.

Abstract Introduction: Molecularly targeted agents, such as the BRAF inhibitor vemurafenib, may produce short-term responses in some patients; however, most patients are intrinsically resistant, or develop resistance through restructuring of signal transduction pathways. SnapPath™ is a live-cell-processing platform that utilizes ex vivo signal transduction modulation of live tumor samples to produce Functional Signaling Profiles (FSPs). Application of this technology to Champions TumorGraft models may provide novel insights to guide oncology drug development as these models preserve the biological properties of the original human tumor. Methods: Fresh melanoma tumor specimens were collected from patients and implanted into immunodeficient mice. Fine needle aspiration biopsies were performed on each melanoma TumorGraft model and processed on the SnapPath™ platform (BioMarker Strategies) to modulate tumor cell signal transduction networks through brief ex vivo exposure to the vemurafenib tool compound PLX-4720. Cell lysates were then analyzed using a multiplexed immunoassay to assess the inhibition of the downstream MAPK markers pMEK1 and pERK-1/2. FSPs were then created for each TumorGraft model based on baseline and modulated levels of each phosphoprotein. In parallel, the in vivo sensitivity to vemurafenib and BRAF mutation status was evaluated in each Champions TumorGraft model. FSPs were then compared with in vivo efficacy, gene expression and genotype data. Results: Functional profiling stratified the TumorGraft models into two distinct groups upon ex vivo exposure to a BRAF inhibitor: 1) MAPK markers suppressed and 2) MAPK markers not suppressed. As anticipated, TumorGraft models that showed resistance to ex vivo BRAF inhibition demonstrated vemurafenib resistance in vivo and were BRAF wild type. There were other models that displayed MAPK suppression with ex vivo BRAF inhibition and vemurafenib sensitivity in vivo or MAPK suppression with ex vivo BRAF inhibition but demonstrated vemurafenib resistance in vivo. One of these TumorGrafts contained a BRAF V600E mutation, suggesting the activation of an alternate pathway that conferred resistance. The other TumorGraft contained a novel BRAF insertion. The functional profiling suggests that this insertion may activate BRAF and is susceptible to vemurafenib inhibition, but the tumor may contain an alternate pathway that confers resistance. Analysis of gene expression data demonstrated hierarchical clustering of BRAF mutated TumorGraft models. Conclusions: These results demonstrate the capability of the SnapPath™ platform to generate FSPs from FNAs of Champions melanoma TumorGraft models. Overall, the combination of Champions TumorGraft models with functional profiling represents a powerful tool for pharmacodynamic assessment of targeted therapeutics in clinically relevant models and has the potential to guide oncology therapy. Citation Format: Elizabeth M. Bruckheimer, Adam Schayowitz, Kala Barnes, Greg Bertenshaw, Tin Khor, James Cotton, Jay Friedman, Dhanrajan Tiruchinapalli, Douglas P. Clark. Functional profiling of Champions TumorGraft™ models from metastatic melanoma patients. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3498. doi:10.1158/1538-7445.AM2013-3498

Carboplatin, paclitaxel and sorafenib in advanced melanoma – what have we learned?

Sorafenib is a small orally available multikinase inhibitor which is approved for the treatment of advanced renal cancer and hepatocellular carcinoma and is one of the first agents in molecular targeted therapy that was brought into clinical trials for metastatic melanoma patients. It blocks tumor proliferation by targeting the vascular endothelial growth factor receptors, platelet-derived growth factor receptors, RAF1, BRAF and probably other receptors involved in signal transduction in tumor cells (1). It is not exactly understood which targets of the above are the most relevant ones in the therapy of advanced melanoma.

Curcumin targets fibroblast–tumor cell interactions in oral squamous cell carcinoma

Co-culture of periodontal ligament fibroblasts (PDLs) and SCC-25 oral squamous carcinoma cells (OSCC) results in conversion of PDLs into carcinoma-associated fibroblasts (CAFs) and induces epithelial-to mesenchymal transition (EMT) of OSCC tumor cells. We hypothesized that Curcumin targets this dynamic mutual interaction between CAFs and tumor cells. Normal and 2μM Curcumin-treated co-culture were performed for 4 days, followed by analysis of tumor cell invasivity, mRNA/protein expression of EMT-markers and mediators, activity measure of matrix metalloproteinase 9 (MMP-9), and western blot analysis of signal transduction in tumor cells and fibroblasts. In Curcumin-treated co-culture, in tumor cells, the levels of nuclear factor κB (NFκBα) and early response kinase (ERK)—decreased, in fibroblasts, integrin αv protein synthesis decreased compared to corresponding cells in normal co-culture. The signal modulatory changes induced by Curcumin caused decreased release of EMT-mediators in CAFs and reversal of EMT in tumor cells, which was associated with decreased invasion. These data confirm the palliative potential of Curcumin in clinical application.

Abstract 3192: Translational research platform (SnapPath™) for enabling drug development and personalized medicine in metastatic melanoma

Abstract Introduction: The approved BRAF inhibitor, vemurafenib, produces short-term responses in many metastatic melanoma patients; however, most patients develop resistance through reactivation of signal transduction pathways. More sophisticated tools for the analysis of signal transduction in preclinical models and clinical samples would vastly improve drug development and personalized cancer therapy. SnapPath™ is a live-cell-processing platform that utilizes ex vivo signal transduction modulation of tumor samples to produce Functional Signaling Profiles (FSPs), which can be used in drug development and clinical studies. Methods: Melanoma cell line xenografts (SK-Mel-28, SK-Mel-3, Colo829, MALME3M and A2058) were generated in athymic nude mice. Fine needle aspiration biopsies (FNA) were performed on each tumor type when tumor volumes reached 500mm3. One FNA sample consisted of 4 passes of a 23G, 1′’ needle on a 10cc syringe. The live cell biopsies were processed on the SnapPath™ live-tumor-cell platform (BioMarker Strategies, LLC) to modulate tumor cell signal transduction networks through brief exposure to the BRAF inhibitor PLX-4720 (3uM) or IGF1 (100ng/ml). The resulting cell lysates were then analyzed using the Bio-Plex multiplexed immunoassay system for the following phosphoproteins: pIGF-1R, pMEK1, pErk-1/2, pAkt, pGSK3B, pp70S6 kinase and pStat3. Functional signaling profiles (FSP) were then created for each tumor based on baseline and modulated levels of each phosphoprotein. FSPs were then compared with known targeted drug sensitivity profiles. Results: Reproducible FSPs were obtained from FNA samples of each xenograft model. The FSPs obtained from FNA samples were similar to those derived from 2-dimensional in vitro cultures of the corresponding cell lines. Exposure of samples to the BRAF inhibitor PLX-4720 resulted in modulation of the MAPK pathway that corresponded to the anticipated activity of the drug. Responses of the xenograft samples to IGF1 growth factor stimulation differed between cell lines and may correlate with resistance mechanisms to BRAF inhibitors via PI3K, mTOR, or AKT, and may also correlate with responses to combination drug therapy. Conclusions: These results demonstrate the capability of the SnapPath™ platform to generate functional signaling profiles from fine needle aspiration biopsy samples of xenografted melanoma tumors. The derived FSPs correspond to anticipated drug effects on signal transduction and consequently represent a valuable tool for pharmacodynamic studies in preclinical xenograft and primary human tumorgraft models as well extension into the clinical setting. The elucidation of bypass pathways by SnapPath™ represents an important new tool for the analysis of resistance mechanisms in melanoma. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3192. doi:1538-7445.AM2012-3192

Logical network of genotoxic stress-induced NF-kappaB signal transduction predicts putative target structures for therapeutic intervention strategies

Genotoxic stress is induced by a broad range of DNA-damaging agents and could lead to a variety of human diseases including cancer. DNA damage is also therapeutically induced for cancer treatment with the aim to eliminate tumor cells. However, the effectiveness of radio- and chemotherapy is strongly hampered by tumor cell resistance. A major reason for radio- and chemotherapeutic resistances is the simultaneous activation of cell survival pathways resulting in the activation of the transcription factor nuclear factor-kappa B (NF-κB). Here, we present a Boolean network model of the NF-κB signal transduction induced by genotoxic stress in epithelial cells. For the representation and analysis of the model, we used the formalism of logical interaction hypergraphs. Model reconstruction was based on a careful meta-analysis of published data. By calculating minimal intervention sets, we identified p53-induced protein with a death domain (PIDD), receptor-interacting protein 1 (RIP1), and protein inhibitor of activated STAT y (PIASy) as putative therapeutic targets to abrogate NF-κB activation resulting in apoptosis. Targeting these structures therapeutically may potentiate the effectiveness of radio-and chemotherapy. Thus, the presented model allows a better understanding of the signal transduction in tumor cells and provides candidates as new therapeutic target structures.

Phase I study of sorafenib in Japanese patients with hepatocellular carcinoma

Sorafenib is an orally active multikinase inhibitor that targets serine and threonine, and tyrosine kinases that are involved in tumor-cell signal transduction and tumor angiogenesis. This phase I trial was conducted to evaluate the pharmacokinetics (PK), safety, and preliminary efficacy of sorafenib in Japanese patients with hepatocellular carcinoma (HCC) with underlying liver dysfunction. Patients with unresectable HCC, Child-Pugh status A or B, and adequate organ functions were treated. A single dose of sorafenib was administered, followed by a 7-day wash-out period, after which patients received either sorafenib 200 mg (cohort 1) or 400 mg (cohort 2) twice daily. The PK were investigated after a single dose and during steady state. The efficacy was evaluated using the Response Evaluation Criteria in Solid Tumors. A total of 27 patients were evaluated for PK, safety, and efficacy. Although both area under the concentration-time curve for 0-12 h and maximal concentration at steady state were slightly lower in Child-Pugh B patients than in Child-Pugh A patients, the difference was not considered to be clinically relevant. Common adverse drug events included elevated lipase, amylase, rash or desquamation, diarrhea, and hand-foot skin reaction. A dose-limiting toxicity of hand-foot skin reaction was observed in one patient (cohort 2). Among the 24 patients evaluable for tumor response, one patient (4%) achieved a partial response, 20 (83%) had stable disease, and three (13%) had progressive disease. Sorafenib demonstrated a favorable tolerability and safety profile in Japanese HCC patients. Moreover, promising preliminary antitumor activity has been observed. Finally, there were no clinically relevant differences in PK between Child-Pugh A and B patients.