Multifaceted Effects of Kinase Inhibitors on Pancreatic Cancer Cells Reveals Pivotal Entities with Therapeutic Implications
Pancreatic cancer is one of the most aggressive forms of cancer and is the seventh leading cause of cancer deaths worldwide. Pancreatic ductal adenocarcinoma (PDAC) accounts for over 90% of pancreatic cancers. Most pancreatic cancers are recalcitrant to radiation, chemotherapy, and immunotherapy, highlighting the urgent need for novel treatment options for this deadly disease. To this end, we screened a library of kinase inhibitors in the PDAC cell lines PANC-1 and BxPC-3 and identified two highly potent molecules: Aurora kinase inhibitor AT 9283 (AT) and EGFR kinase inhibitor WZ 3146 (WZ). Both AT and WZ exhibited a dose-dependent inhibition of viability in both cell lines. Thus, we conducted an in-depth multilevel (cellular, molecular, and proteomic) analysis with AT and WZ in PANC-1 cells, which harbor KRAS mutation and exhibit quasimesenchymal properties representing pancreatic cancer cells as having intrinsic chemoresistance and the potential for differential response to therapy. Elucidation of the molecular mechanism of action of AT and WZ revealed an impact on the programmed cell death pathway with an increase in apoptotic, multicaspase, and caspase 3/7 positive cells. Additionally, the key survival molecule Bcl-2 was impacted. Moreover, cell cycle arrest was observed with both kinase inhibitors. Additionally, an increase in superoxide radicals was observed in the AT-treated group. Importantly, proteomic profiling revealed differentially regulated key entities with multifaceted effects, which could have a deleterious impact on PDAC. These findings suggest potential targets for efficacious treatment, including a possible increase in the efficacy of immunotherapy using PD-L1 antibody due to the upregulation of lactoferrin and radixin. Furthermore, combination therapy outcomes with gemcitabine/platinum drugs may also be more effective due to an increase in the NADH dehydrogenase complex. Notably, protein–protein interaction analysis (STRING) revealed possible enrichment of reactome pathway entities. Additionally, novel therapy options, such as vimentin-antibody–drug conjugates, could be explored. Therefore, future studies with the two kinases as monotherapy/combination therapy are warranted.
Disruption of Glioblastoma Multiforme Cell Circuits with Cinnamaldehyde Highlights Potential Targets with Implications for Novel Therapeutic Strategies
Casticin Impacts Key Signaling Pathways in Colorectal Cancer Cells Leading to Cell Death with Therapeutic Implications
Disruption of Colorectal Cancer Network by Polyphyllins Reveals Pivotal Entities with Implications for Chemoimmunotherapy
The prevalence of colorectal cancer has increased world-wide with high rates of mortality and morbidity. In the absence of efficacious drugs to treat this neoplasia, there is an imminent need to discover molecules with multifaceted effects. To this end, we opted to study the effect of steroidal saponins such as Polyphyllins. We performed anticancer activity studies with three analogs of Polyphyllins: Polyphyllin D (PD), Polyphyllin II (PII) and Polyphyllin G (PG). Here we show the potent effect of PD, PII (IC50 of 0.5−1 µM) and PG (IC50 of 3 µM) in inhibiting the viability of colorectal adenocarcinoma cells (DLD-1) and colorectal carcinoma cells (HCT116). PD and PII also showed inhibition of cell proliferation and sustained response upon withdrawal of the compounds when assessed by clonogenic assays in both the cell lines. Elucidation of the molecular mode of action revealed impact on the programmed cell death pathway. Additionally, proteomic profiling of DLD-1 revealed pivotal proteins differentially regulated by PD and PII, including a downregulated peroxiredoxin-1 which is considered as one of the novel targets to combat colorectal cancers and an upregulated elongation factor 2 (EF2), one of the key molecules considered as a tumor associated antigen (TAA) in colon cancer. Entities of cell metabolic pathways including downregulation of the key enzyme Phosphoglycerate kinase 1 of the glycolytic pathway was also observed. Importantly, the fold changes per se of the key components has led to the loss of viability of the colorectal cancer cells. We envision that the multifaceted function of PD and PII against the proliferation of colorectal carcinoma cells could have potential for novel treatments such as chemoimmunotherapy for colorectal adenocarcinomas. Future studies to develop these compounds as potent anti-colorectal cancer agents are warranted.
Computational Identification of Stearic Acid as a Potential PDK1 Inhibitor and In Vitro Validation of Stearic Acid as Colon Cancer Therapeutic in Combination with 5-Fluorouracil
Background: Colorectal cancer is the third largest cause of cancer-related mortality worldwide. Although current treatments with chemotherapeutics have allowed for management of colorectal cancer, additional novel treatments are essential. Intervening with the metabolic reprogramming observed in cancers called “Warburg Effect,” is one of the novel strategies considered to combat cancers. In the metabolic reprogramming pathway, pyruvate dehydrogenase kinase (PDK1) plays a pivotal role. Identification and characterization of a PDK1 inhibitor is of paramount importance. Further, for efficacious treatment of colorectal cancers, combinatorial regimens are essential. To this end, we opted to identify a PDK1 inhibitor using computational structure-based drug design FINDSITEcomb and perform combinatorial studies with 5-FU for efficacious treatment of colorectal cancers.
Methods: Using computational structure-based drug design FINDSITEcomb, stearic acid (SA) was identified as a possible PDK1 inhibitor. Elucidation of the mechanism of action of SA was performed using flow cytometry, clonogenic assays.
Results: When the growth inhibitory potential of SA was tested on colorectal adenocarcinoma (DLD-1) cells, a 50% inhibitory concentration (IC50) of 60 µM was recorded. Moreover, SA inhibited the proliferation potential of DLD-1 cells as shown by the clonogenic assay and there was a sustained response even after withdrawal of the compound. Elucidation of the mechanism of action revealed, that the inhibitory effect of SA was through the programmed cell death pathway. There was increase in the number of apoptotic and multicaspase positive cells. SA also impacted the levels of the cell survival protein Bcl-2. With the aim of achieving improved treatment for colorectal cancer, we opted to combine 5-fluorouracil (5-FU), the currently used drug in the clinic, with SA. Combining SA with 5-FU, revealed a synergistic effect in which the IC50 of 5-FU decreased from 25 to 6 µM upon combination with 60 µM SA. Further, SA did not inhibit non-tumorigenic NIH-3T3 proliferation.
Conclusions: We envision that this significant decrease in the IC50 of 5-FU could translate into less side effects of 5-FU and increase the efficacy of the treatment due to the multifaceted action of SA. The data generated from the current studies on the inhibition of colorectal adenocarcinoma by SA discovered by the use of the computational program as well as synergistic action with 5-FU should open up novel therapeutic options for the management of colorectal adenocarcinomas.
Interception of Signaling Circuits of Esophageal Adenocarcinoma Cells by Resveratrol Reveals Molecular and Immunomodulatory Signatures
Cellular molecular and proteomic profiling deciphers the SIRT1 controlled cell death pathways in esophageal adenocarcinoma cells
Worldwide prevalence of esophageal adenocarcinomas with high rates of mortality coupled with increased mutations in esophageal cells warrants investigation to understand deregulation of cell signaling pathways leading to cancer. To this end, the current study was undertaken to unravel the cell death signatures using the model human esophageal adenocarcinoma cell line-OE33. The strategy involved targeting the key epigenetic modulator SIRT1, a histone deacetylase by a small molecule inhibitor – sirtinol. Sirtinol induced a dose-dependent inhibition of cell viability under both normoxic and hypoxic conditions with long term impact on proliferation as shown by clonogenic assays. Signature apoptotic signaling pathways including caspase activation and decreased Bcl-2 were observed. Proteomic analysis highlighted an array of entities affected including molecules involved in replication, transcription, protein synthesis, cell division control, stress-related proteins, spliceosome components, protein processing and cell detoxification/degradation systems. Importantly, the stoichiometry of the fold changes of the affected proteins per se could govern the cell death phenotype by sirtinol. Sirtinol could also potentially curb resistant and recurrent tumors that reside in hypoxic environments. Overall, in addition to unraveling the cellular, molecular and proteomics basis of SIRT1 inhibition, the findings open up avenues for designing novel strategies against esophageal adenocarcinoma.
Inhibition of Prostate Cancer Cells by 4,5-Dicaffeoylquinic Acid through Cell Cycle Arrest
Prostate cancer is a major cause of cancer-related mortality in men. Even though current therapeutic management has contributed to reducing mortality, additional intervention strategies are warranted to further improve the outcomes. To this end, we have investigated the efficacy of dicaffeoylquinic acids, ingredients in Yerba Mate (Ilex paraguariensis), an evergreen cultivated in South America, the leaves of which are used to prepare a tea/coffee-like drink. Of the various analogs tested, 4,5-dicaffeoylquinic acid (4,5-diCQA) was the most active molecule against DU-145 prostate cancer cells with a 50% inhibitory concentration (IC50) of 5 μM. 4,5-diCQA was active both under normoxic and hypoxic conditions. The effect of 72-hour treatment on DU-145 cells persisted for an extended time period as assessed by clonogenic assay. Mechanistic studies revealed that the toxic effect was not due to induction of programmed cell death but through cell cycle arrest at S phase. Additionally, 4,5-diCQA did not impact PI3K/MAPK signaling pathway nor did it affect the depolarization of the mitochondrial membrane. 4,5-diCQA-induced accumulation of cells in the S-phase also seems to negatively impact Bcl-2 expression. 4,5-diCQA also exhibited inhibitory activity on LNCaP and PC-3 prostate cancer cells suggesting that it has therapeutic potential on a broad range of prostate cancers. Taken together, the novel inhibitory activity and mechanism of action of 4,5-diCQA opens up potential therapeutic options for using this molecule as monotherapy as well as in combinatorial therapies for the clinical management of prostate cancer.
—23 May 2019
“Recently, I’ve made cancer the focus of Georgia Tech’s Quantitative Engineering Physiology Course. The global impact of this disease is clear and the recently launched “Cancer Moonshot” initiative makes it a timely topic. We challenge our students to own their laboratory experiences and engineer strategies to interfere with the physiological processes that lead to cancer. Student teams develop their own comprehensive research proposals, which then become the foundation for experimental validation in the lab and employing relevant techniques.”