// Reza Bayat Mokhtari 1,2,4 , Tina S. Homayouni 1 , Narges Baluch 3 , Evgeniya Morgatskaya 1 , Sushil Kumar 1 , Bikul Das 4 and Herman Yeger 1,2 1 Developmental and Stem Cell Biology, The Hospital for Sick Children, Toronto, Ontario, Canada 2 Department of Paediatric Laboratory Medicine, The Hospital for Sick Children and Institute of Medical Science, University of Toronto, Toronto, Ontario, Canada 3 Department of Pathology and Molecular Medicine, Queen’s University, Kingston, Ontario, Canada 4 Department of Immunology and Infectious Diseases, The Forsyth Institute, Cambridge, Massachusetts, USA Correspondence to: Herman Yeger, email: // Reza Bayat Mokhtari, email: // Keywords : Nrf2-Keap1, HIF-1alpha, carbonic anhydrase 9 (CAIX), histone deacetylase inhibitor (HDACi), carbonic anhydrase inhibitor (CAI) Received : October 19, 2016 Accepted : February 27, 2017 Published : March 30, 2017 Abstract Combination therapy, a treatment modality that combines two or more therapeutic agents, is a cornerstone of cancer therapy. The amalgamation of anti-cancer drugs enhances efficacy compared to the mono-therapy approach because it targets key pathways in a characteristically synergistic or an additive manner. This approach potentially reduces drug resistance, while simultaneously providing therapeutic anti-cancer benefits, such as reducing tumour growth and metastatic potential, arresting mitotically active cells, reducing cancer stem cell populations, and inducing apoptosis. The 5-year survival rates for most metastatic cancers are still quite low, and the process of developing a new anti-cancer drug is costly and extremely time-consuming. Therefore, new strategies that target the survival pathways that provide efficient and effective results at an affordable cost are being considered. One such approach incorporates repurposing therapeutic agents initially used for the treatment of different diseases other than cancer. This approach is effective primarily when the FDA-approved agent targets similar pathways found in cancer. Because one of the drugs used in combination therapy is already FDA-approved, overall costs of combination therapy research are reduced. This increases cost efficiency of therapy, thereby benefiting the “medically underserved”. In addition, an approach that combines repurposed pharmaceutical agents with other therapeutics has shown promising results in mitigating tumour burden. In this systematic review, we discuss important pathways commonly targeted in cancer therapy. Furthermore, we also review important repurposed or primary anti-cancer agents that have gained popularity in clinical trials and research since 2012.

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Combination therapy in combating cancer.

Stevia rebaudiana Bertoni is a shrub having a high content of sweet diterpenoid glycosides in its leaves, mainly stevioside and rebaudioside A, which are used as noncaloric, natural sweeteners. The aim of this study was to deepen the knowledge about the insulin-mimetic effect exerted by four different mixtures of steviol glycosides, rich in stevioside and rebaudioside A, in neonatal rat cardiac fibroblasts. The potential antioxidant activity of these steviol glycosides was also assessed, as oxidative stress is associated with diabetes. Likewise the insulin effect, steviol glycosides caused an increase in glucose uptake into rat fibroblasts by activating the PI3K/Akt pathway, thus inducing Glut4 translocation to the plasma membrane. The presence of S961, an insulin antagonist, completely abolished these effects, allowing to hypothesize that steviol glycosides could act as ligands of the same receptor engaged by insulin. Moreover, steviol glycosides counteracted oxidative stress by increasing reduced glutathione intracellular levels and upregulating expression and activity of the two antioxidant enzymes superoxide dismutase and catalase. The present work unravels the insulin-mimetic effect and the antioxidant property exerted by steviol glycosides, suggesting their potential beneficial role in the cotreatment of diabetes and in health maintenance.

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Glycosides from Stevia rebaudiana Bertoni Possess Insulin-Mimetic and Antioxidant Activities in Rat Cardiac Fibroblasts.

Glioblastoma multiforme (GBM) is a debilitating disease that is associated with poor prognosis, short median patient survival and a very limited response to therapies. GBM has a very complex pathogenesis that involves mutations and alterations of several key cellular pathways that are involved in cell proliferation, survival, migration and angiogenesis. Therefore, efforts that are directed toward better understanding of GBM pathogenesis are essential to the development of efficient therapies that provide hope and extent patient survival. In this review, we outline the alterations commonly associated with GBM pathogenesis and summarize therapeutic strategies that are aimed at targeting aberrant cellular pathways in GBM.

Targeting cellular pathways in glioblastoma multiforme

Gastric cancer is a heterogeneous cancer, making treatment responses difficult to predict. Here we show that we identify two distinct molecular subtypes, mesenchymal phenotype (MP) and epithelial phenotype (EP), by analyzing genomic and proteomic data. Molecularly, MP subtype tumors show high genomic integrity characterized by low mutation rates and microsatellite stability, whereas EP subtype tumors show low genomic integrity. Clinically, the MP subtype is associated with markedly poor survival and resistance to standard chemotherapy, whereas the EP subtype is associated with better survival rates and sensitivity to chemotherapy. Integrative analysis shows that signaling pathways driving epithelial-to-mesenchymal transition and insulin-like growth factor 1 (IGF1)/IGF1 receptor (IGF1R) pathway are highly activated in MP subtype tumors. Importantly, MP subtype cancer cells are more sensitive to inhibition of IGF1/IGF1R pathway than EP subtype. Detailed characterization of these two subtypes could identify novel therapeutic targets and useful biomarkers for prognosis and therapy response.

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Clinical and genomic landscape of gastric cancer with a mesenchymal phenotype

Regulation of the PI-3 kinase (PI3K)/Akt signalling pathway is essential for maintaining the integrity of fundamental cellular processes, cell growth, survival, death and metabolism, and dysregulation of this pathway is implicated in the development and progression of cancers. Receptor tyrosine kinases (RTKs) are major upstream regulators of PI3K/Akt signalling. The phosphatase and tensin homologue (PTEN), a well characterised tumour suppressor, is a prime antagonist of PI3K and therefore a negative regulator of this pathway. Loss or inactivation of PTEN, which occurs in many tumour types, leads to overactivation of RTK/PI3K/Akt signalling driving tumourigenesis. Cellular PTEN levels are tightly regulated by a number of transcriptional, post-transcriptional and post-translational regulatory mechanisms. Of particular interest, transcription of the PTEN pseudogene, PTENP1, produces sense and antisense transcripts that exhibit post-transcriptional and transcriptional modulation of PTEN expression respectively. These additional levels of regulatory complexity governing PTEN expression add to the overall intricacies of the regulation of RTK/PI-3 K/Akt signalling. This review will discuss the regulation of oncogenic PI3K signalling by PTEN (the regulator) with a focus on the modulatory effects of the sense and antisense transcripts of PTENP1 on PTEN expression, and will further explore the potential for new therapeutic opportunities in cancer treatment.

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PTEN/PTENP1: ‘Regulating the regulator of RTK-dependent PI3K/Akt signalling’, new targets for cancer therapy

Insulin and insulin-like growth factor (IGF) signaling system, commonly known for fine-tuning numerous biological processes, has lately made its mark as a much sought-after therapeutic targets for diabetes and cancer These receptors make an attractive anticancer target owing to their overexpression in variety of cancer especially in prostate and breast cancer Inhibitors of IGF signaling were subjected to clinical cancer trials with the main objective to confirm the effectiveness of these receptors as a therapeutic target However, the results that these trials produced proved to be disappointing as the role played by the cross talk between IGF and insulin receptor (IR) signaling pathways at the receptor level or at downstream signaling level became more lucid Therapeutic strategy for IGF-1R and IR inhibition mainly encompasses three main approaches namely receptor blockade with monoclonal antibodies, tyrosine kinase inhibition (ATP antagonist and non-ATP antagonist), and ligand neutralization via monoclonal antibodies targeted to ligand or recombinant IGF-binding proteins Other drug-discovery approaches are employed to target IGF-1R, and IR includes antisense oligonucleotides and recombinant IGF-binding proteins However, therapies with monoclonal antibodies and tyrosine kinase inhibition targeting the IGF-1R are not evidenced to be satisfactory as expected Factors that are duly held responsible for the unsuccessfulness of these therapies include (a) the existence of the IR isoform A overexpressed on a variety of cancers, enhancing the mitogenic signals to the nucleus leading to the endorsement of cell growth, (b) IGF-1R and IR that form hybrid receptors sensitive to the stimulation of all three IGF axis ligands, and (c) IGF-1R and IR that also have the potential to form hybrid receptors with other tyrosine kinase to potentiate the cellular transformation, tumorigenesis, and tumor vascularization This mini review is a concerted effort to explore and fathom the well-recognized roles of the IRA signaling system in human cancer phenotype and the main strategies that have been so far evaluated to target the IR and IGF-1R

Insulin receptor (IR) and insulin-like growth factor receptor 1 (IGF-1R) signaling systems: novel treatment strategies for cancer

The role played by overexpression of tyrosine kinase epidermal growth factor receptor (EGFR), the transmembrane receptor central to numerous cellular processes comprising cell migration, adhesion, apoptosis, and cell proliferation, has been highlighted in various cancers such as prostate, breast, lung, and ovarian cancers as well as in mutations in the EGFR kinase domain. Although many therapeutic approaches have targetted EGFR, the mutations occurring in the EGFR kinase domain including L858 EGFR and T790/L858R had led to the amplification of EGFR signals, consequently leading to increased cell proliferation and cell growth. The strategies involving the inhibition of EGFR L858 and T790M have been accredited with limited achievement in addition to being associated with unwanted adverse effects as a result of crosstalk of wild-type EGFR. All current EGFR tyrosine kinase inhibitors have been identified as ATP competitive inhibitors of wild-type EGFR possessing aniline and quinazoline moiety on the ligands skeleton. Our results obtained by performing molecular docking study on Maestro 9.3 molecular docking suite indicated that CID5280343 possesses better energy conformation against wild-type EGFR as well as two mutated EGFR. Moreover, it was discovered in this study that the natural compounds CID72276, CID5280445, CID441794, and CID72277 and InterBioScreen’s library STOCK1N-78657, STOCK1N-78976, and STOCK1N-78847 have better binding conformation against gatekeeper T790M mutated EGFR concluded to be brought about by means of flexible ligands/receptor-based molecular docking protocol. Miraculous features of these compounds are their various pharmacokinetic and pharmacodynamic parameters which were found to be satisfactory as drug-like molecules. This molecular docking study also summarizes docking free energy, protein–ligands interaction profile, and pharmacokinetic and pharmacodynamic parameter of lead molecules which were tremendously helpful in enhancing the activity of these natural compounds against EGFR.

In silico molecular docking study of natural compounds on wild and mutated epidermal growth factor receptor

Myricetin is a naturally omnipresent benzo-α-pyrone flavonoids derivative; has potent anticancer activity. Receptor tyrosine kinases family provides the decisive role in cancer initiation and progression. These receptors have recently caught the attention of the researchers as an attractive target to combat cancer, owing to the evidences endorsed their over-expression on cancer cells. This study is a concerted effort to explore the potent and specific multi-targeted inhibitor against RTKs and AR\ER employing molecular docking approach. IR, IGF1R, EGFR, VEGFR1, VEGFR2, and AR\ER were chosen as a protein and natural compounds as a ligand. Molecular docking procedure followed by using Maestro 9.6 (Schrodinger Inc). All natural compounds were docked with the X-ray crystal structures of selected proteins by employing grid-based ligand docking with energetics Maestro 9.6. IBS natural compounds docked with each selected protein molecules by using GLIDE high throughput virtual screening. On the basis of Gscore, we selected 20 compounds from IBS (50,000 compounds) along with 68 anticancer compounds from published literature for GLIDE extra precision molecular docking. Calculated docking free energy yielded the excellent dock score for the myricetin when docked with proteins EGFR, IR, and AR\ER. Protein-ligand interactions profile highlighted that the lipophilic, hydrogen bonding and π-π stacking interactions play a central role in protein-ligand interactions at the active site. The results of MTT assay reveal that the myricetin inhibit the viability and proliferation of cancer cells in a dose-dependent manner. Treatment with the myricetin led to down-regulation of mRNA expression of EGFR, IR, mTOR, and Bcl-2. Although, further in vitro and in vivo experimental studies are required for the experimental validation of our findings.

Screening and biological evaluation of myricetin as a multiple target inhibitor insulin, epidermal growth factor, and androgen receptor; in silico and in vitro.

Phosphoinositide-3 kinase (PI3K) signaling pathway comprises of a cornucopia of protein molecules capable of regulating numerous cellular events, including cell survival, cell cycle regulation, angiogenesis, and apoptosis. Deregulation of PI3K downstream signaling is a phenomenon commonly seen in various types of cancer and also held responsible for poor prognosis and resistance to chemotherapy. Targeting PI3K signaling pathway has become a new and promising strategy in combating cancer. In the present study, PI3K signaling components PI3K, PDK1, Akt, and mTOR were chosen and 51 natural compounds along with 17 reference compounds were selected as ligand with the aid of PubMed published literature search. Ligands were docked to protein molecules by using Maestro 9.3 (Schrodinger Inc.). It was discovered in this study that compounds myricetin, quercetin, morin, luteolin, and emodin yielded excellent dock score with the proteins concluded with the help of docking free energy. The remarkable feature of these compounds are their various pharmacodynamics and pharmacokinetic characteristics, many of which are in accordance with the “Lipinski’s Rule of five”. The docking study carried out is an endeavor to portray the docking of these compounds with the proteins, to summarize the various Gscore, hydrogen bond, electrostatic bond, and to chart out various factors that are decisive for and also govern the protein–ligand interactions.

Multitargeted molecular docking study of plant-derived natural products on phosphoinositide-3 kinase pathway components

Receptors for growth factors encompass within the superfamily of receptor tyrosine kinases and are known to regulate numerous biological processes including cellular growth, proliferation, metabolism, survival, cell differentiation and apoptosis. These receptors have recently caught the attention of the researchers as an attractive target to combat cancer owing to the evidence suggesting their over-expression in cancer cells. Therefore, we studied receptor-based molecular docking of IR (PDB; 3ETA), IGF1R (PDB; 1K3A), EGFR (PDB; 1M17), VEGFIR (PDB; 3HNG), and VEGFIIR (PDB; 2OH4) against natural compounds. Further, in vitro investigation of the biological effect of lead molecules in an array of cancer cell lines was done. All selected natural compounds were docked with the X-ray crystal structure of selected protein by employing GLIDE (Grid-based Ligand Docking with Energetics) Maestro 9.6. InterBioScreen natural compounds docked with each selected protein molecules by using GLIDE high throughput virtual screening. On the basis of Gscore, we select 20 compounds along with 68 anticancer compounds for GLIDE extra precision molecular docking. It was discovered in this study that compound epigallocatechin gallate (EGCG) yielded magnificent Gscore with IGF1R (PDB; 1K3A) and VEGFIIR (PDB; 2OH4), and protein-ligand interactions are chart out. Effect of EGCG on biological activity such as mRNA expression of selected protein, cell proliferation, oxidative stress, and cell migration was reported after the 48 h treatments in cancer cell lines. The RT-PCR densitometric bands analysis showed that compound EGCG reduced the mRNA expression of IGF1R, VEGFIIR, and mTOR at 80 μM concentration. Moreover, EGCG significantly reduced cell proliferation and ROS generation after 48 h treatments. Our result also indicated a reduction in the potential for cell migration that might show in vivo anti-metastasis activity of EGCG.

Screening of multi-targeted natural compounds for receptor tyrosine kinases inhibitors and biological evaluation on cancer cell lines, in silico and in vitro

Pushpendra Singh

Papers

Insulin receptor (IR) and insulin-like growth factor receptor 1 (IGF-1R) signaling systems: novel treatment strategies for cancer

In silico molecular docking study of natural compounds on wild and mutated epidermal growth factor receptor

Screening and biological evaluation of myricetin as a multiple target inhibitor insulin, epidermal growth factor, and androgen receptor; in silico and in vitro.

Multitargeted molecular docking study of plant-derived natural products on phosphoinositide-3 kinase pathway components

Screening of multi-targeted natural compounds for receptor tyrosine kinases inhibitors and biological evaluation on cancer cell lines, in silico and in vitro