Molecules For Cancer Therapy Research Articles

Magnetic Reactive Oxygen Species Nanoreactor for Switchable Magnetic Resonance Imaging Guided Cancer Therapy Based on pH-Sensitive Fe5C2@Fe3O4 Nanoparticles.

Reactive oxygen species (ROS) are crucial molecules in cancer therapy. Unfortunately, the therapeutic efficiency of ROS is unsatisfactory in clinic, primarily due to their rigorous production conditions. By taking advantage of the intrinsic acidity and overproduction of H2O2 in the tumor environment, we have reported an ROS nanoreactor based on core-shell-structured iron carbide (Fe5C2@Fe3O4) nanoparticles (NPs) through the catalysis of the Fenton reaction. These NPs are able to release ferrous ions in acidic environments to disproportionate H2O2 into •OH radicals, which effectively inhibits the proliferation of tumor cells both in vitro and in vivo. The high magnetization of Fe5C2@Fe3O4 NPs is favorable for both magnetic targeting and T2-weighted magnetic resonance imaging (MRI). Ionization of these NPs simultaneously decreases the T2 signal and enhances the T1 signal in MRI, and this T2/T1 switching process provides the visualization of ferrous ions release and ROS generation for the supervision of tumor curing. These Fe5C2@Fe3O4 NPs show great potential in endogenous environment-excited cancer therapy with high efficiency and tumor specificity and can be guided further by MRI.

Reduction of IL-2 fragmentation during manufacturing of a novel immunocytokine by DoE process optimization.

Interleukin-2 (IL-2) is a potent molecule in cancer therapy. Clinical application, however, is limited due to its strong side effects during the treatment. We developed an IL-2 variant (IL-2v) immunocytokine to circumvent the drawbacks of the current IL-2 therapy. During the production of the IL-2v immunocytokine in Chinese hamster ovary (CHO) cells, molecules with fragmented IL-2v and therefore reduced cytokine activity can be observed. To control product fragmentationdifferent production process conditions were investigated. By shifting temperature or pH after the cell growth phase to lower values, fragmented species can be reduced from 10% to 12% to about 4%. However, with the adopted process conditions, the effective titer is decreased concomitantly. Moreover, fermentation length and inoculation cell density are parameters to adjust fragmentation and effective titer. A suitable method for efficient process optimization is the design of experiment approach. With this procedure, novel optimal values for temperature, pH value, harvest day, and inoculation cell densities were proposed and tested subsequently. In comparison to the former process, the improved process reduces fragmentation by 66% while keeping the effective titer comparable. In summary, these findings will help to control fragmentation in CHO production processes of different IL-2v or IL-2 containing therapeutic proteins.

Targeting Immune-Related Molecules in Cancer Therapy: A Comprehensive In Vitro Analysis on Patient-Derived Tumor Models.

This study investigated the impact of immune-related pathway inhibition, among them indolamine 2,3-dioxygenase (IDO), alone and together with immune cells on growth and viability of colorectal cancer (CRC) cells. A panel of patient-derived CRC cell lines with different molecular characteristics (CpG island methylator phenotype, chromosomal, and microsatellite instability) was included. Initial phenotyping of CRC cell lines (n=17) revealed high abundance of immunosuppressive checkpoint-molecules in general, but an individual profile for IDO. Presence of immune-related molecules was independent of the molecular subtype. Selective treatment of CRC cell lines showing high or low IDO expression (n=2 cell lines each) was performed with single agents and combinations of Indoximod, Curcumin, and Gemcitabine with and without the addition of peripheral blood lymphocytes (PBL) in an allogeneic setting. All substances affected CRC cell growth in a cell line specific manner. The combination of Curcumin and Gemcitabine proved to be most effective in tumor cell elimination. Functional read-out analyses identified cellular senescence, after both single and combined treatment. Curcumin alone exerted strong cytotoxic effects by inducing early and late apoptosis. Necrosis was not detectable at all. Addition of lymphocytes generally boosted antitumoral effects of all IDO-inhibitors, with up to 80 % cytotoxicity for the Curcumin treatment. Here, no obvious differences became apparent between individual cell lines. Combined application of Curcumin and low-dose chemotherapy is a promising strategy to kill tumor target cells and to stimulate antitumoral immune responses.

Platinum and ruthenium complexes as promising molecules in cancer therapy

Cancer is one of the most common fatal diseases in humans nowadays. About 20 million new cancer cases are expected in the next two decades worldwide. The development of new chemotherapeutic agents with improved properties is presently the main challenge in the medicinal chemistry. Cisplatin was introduced to oncology in 1978 as first chemotherapeutic agent regarding its specific interaction with DNA, leading to its damage and causing the cell death. Since the first application of cisplatin in cancer therapy, there has been a growing interest in new metal-based compounds, in particular platinum and ruthenium complexes, with better anticancer activity and less side-effects compared to cisplatin. Carboplatin and oxaliplatin have shown promising action against some types of cancer, which are resistant to cisplatin. With the aim to overcome cross-resistance to these Pt(II) drugs, bioavailable platinum complexes (satraplatin and picoplatin) firstly found application as orally administered drugs, as well as some combined therapies of Pt(II) drugs (cisplatin, picoplatin) with specific resistant modulators. In recent years, novel polymer and liposomal formulations of platinum drugs (prolindac, lipoplatin, lipoxal, aroplatin) have been designed with strategy to improve drug delivery to target cancer cells and reduce toxicity. Complexes based on ruthenium have great potential to become leading candidates for the medical use in anticancer therapy. Some of these compounds have shown good anticancer activity, both in vitro and in vivo and two of them (KP1019 and NAMI-A) have passed clinical trials and given promising results.

Neoantigens in tumor immunotherapy

Malignant tumors are known to have complex mutational profiles and harbor concurrent alterations in many somatic genes. The inher ent genetic heterogeneity is a critical determinant of cancer cells. The term of neoantigens was introduce to emphasize that these antigens are specific for cancer cells and are absent in normal tissue. Neoantigenes are highly immunogenic and at present are considered to be the target molecules in cancer therapy. Although the neoantigens have been known for a long time, their study and use became possible with the increase in the availability of mass cluster sequencing to detect all mutations in tumors and bioinformatic algorithms predicting what mutated peptides will be highly affine to human leukocyte antigen autologous molecules with subsequent activation of the immune response.

Forskolin improves sensitivity to doxorubicin of triple negative breast cancer cells via Protein Kinase A-mediated ERK1/2 inhibition

Triple negative breast cancer (TNBC) is an invasive, metastatic, highly aggressive tumor. Cytotoxic chemotherapy represents the current treatment for TNBC. However, relapse and chemo-resistance are very frequent. Therefore, new therapeutic approaches that are able to increase the sensitivity to cytotoxic drugs are needed. Forskolin, a natural cAMP elevating agent, has been used for several centuries in medicine and its safeness has also been demonstrated in modern studies. Recently, forskolin is emerging as a possible novel molecule for cancer therapy. Here, we investigate the effects of forskolin on the sensitivity of MDA-MB-231 and MDA-MB-468 TNBC cells to doxorubicin through MTT assay, flow cytometry-based assays (cell-cycle progression and cell death), cell number counting and immunoblotting experiments. We demonstrate that forskolin strongly enhances doxorubicin-induced antiproliferative effects by cell death induction. Similar effects are observed with IBMX and isoproterenol cAMP elevating agents and 8-Br-cAMP analog, but not by using 8-pCPT-2′-O-Me-cAMP Epac activator. It is important to note that the forskolin-induced potentiation of sensitivity to doxorubicin is accompanied by a strong inhibition of ERK1/2 phosphorylation, is mimicked by ERK inhibitor PD98059 and is prevented by pre-treatment with Protein Kinase A (PKA) and adenylate cyclase inhibitors. Altogether, our data indicate that forskolin sensitizes TNBC cells to doxorubicin via a mechanism depending on the cAMP/PKA-mediated ERK inhibition. Our findings sustain the evidence of anticancer activity mediated by forskolin and encourage the design of future in-vivo/clinical studies in order to explore forskolin as a doxorubicin sensitizer for possible use in TNBC patients.

Isolation and characterization of anti ROR1 single chain fragment variable antibodies using phage display technique.

Receptor tyrosine kinase-like orphan receptor (ROR1) belongs to one of the families of receptor tyrosine kinases (RTKs). RTKs are involved in the various physiologic cellular functions including proliferation, migration, survival, signaling and differentiation. Several RTKs are deregulated in various cancers implying the targeting potential of these molecules in cancer therapy. ROR1 has recently been shown to be expressed in various types of cancer cells but not in normal adult cells. Hence a molecular inhibitor of extracellular domain of ROR1 that inhibits ROR1-cell surface interaction is of great therapeutic importance. In an attempt to develop molecular inhibitors of ROR1, we screened single chain variable fragment (scFv) phage display libraries, Tomlinson I + J, against one specific synthetic oligopeptide from extracellular domain of ROR1 and selected scFvs were characterized using various immunological techniques. Several ROR1 specific scFvs were selected following five rounds of panning procedure. The scFvs showed specific binding to ROR1 using immunological techniques. Our results demonstrate successful isolation and characterization of specific ROR1 scFvs that may have great therapeutic potential in cancer immunotherapy.

Multi-Target Directed Indole Based Hybrid Molecules in Cancer Therapy : An Up-To-Date Evidence-Based Review.

Cancer is a multifactorial disease and most of its types still remain incurable, in spite of enormous efforts to explicate various tumor pathophysiology. The anti-cancer drug discovery paradigm "one-compound-one-target" has failed and subsequently shifted to two-drug cocktail and recently the "multi-target approach" in order to design and develop agents able to act simultaneously on multiple intracellular constituents and signaling pathways. Novel hybrid compounds are now designed by incorporating two covalently linked independently acting pharmacores, each efficient at combating cancer. They can deliver synergistic effects from the dual action of both independently acting moieties by interacting with multiple targets. These composite molecules are also less prone to drug resistance, leading to an improved pharmacological potency than each individual moiety. As indole nucleus is a central component of many natural and synthetic molecules with extensive biological activity, this review incorporates a variety of such hybrid compounds with indole moiety as one of the active units, where better therapeutic effect has been successfully achieved, by either simultaneous or sequential action of individual functional pharmacore. The current limitations and challenges encountered in the development of these hybrid agents are also discussed.

Multiple-Sized Amphiphilic Janus Gold Nanoparticles by Ligand Exchange at Toluene/Water Interface

Amphiphilic Janus gold nanoparticles (GNPs) are prospected to encapsulate drug molecules in cancer therapy and to serve as heterogeneous catalysts at oil/water interfaces, where Janus GNPs with different sizes are required. In this work, multiple-sized precursor GNPs were synthesized by seeded growth method protected with tris(hydroxymethyl)phosphine oxide (THPO) ligand molecule, and a ligand exchange reaction with triphenylphosphine (PPh3) at the toluene/water interface was employed to prepare amphiphilic Janus GNPs. UV-vis and transmission electron microscopy (TEM) analyses indicate that the as-prepared GNPs are nanocrystals with average diameters of 2.3 nm, 9.5 nm, 16.1 nm and 18.8 nm, respectively. Contact angle, Raman and X-ray photonic spectroscopy (XPS) analyses reveal that the self-assembled GNP films exhibit hydrophilic on one side and hydrophobic on another, owing to the adsorption of hydrophilic ligands (THPO and THP) and a similar amount of hydrophobic ligands (PPh3 and PPh3O). Angle-resolved XPS analysis further demonstrates that the individual GNPs actually possess hydrophilic and hydrophobic compartments on the surface, which regularly packed by supramolecular interactions at toluene/water interface to form the self-assembled GNP films.

MiRNAs: Novel molecules in cancer therapy

MiRNAs: Novel molecules in cancer therapy

Reduced graphene oxide-silver nanoparticle nanocomposite: a potential anticancer nanotherapy.

BackgroundGraphene and graphene-based nanocomposites are used in various research areas including sensing, energy storage, and catalysis. The mechanical, thermal, electrical, and biological properties render graphene-based nanocomposites of metallic nanoparticles useful for several biomedical applications. Epithelial ovarian carcinoma is the fifth most deadly cancer in women; most tumors initially respond to chemotherapy, but eventually acquire chemoresistance. Consequently, the development of novel molecules for cancer therapy is essential. This study was designed to develop a simple, non-toxic, environmentally friendly method for the synthesis of reduced graphene oxide–silver (rGO–Ag) nanoparticle nanocomposites using Tilia amurensis plant extracts as reducing and stabilizing agents. The anticancer properties of rGO–Ag were evaluated in ovarian cancer cells.MethodsThe synthesized rGO–Ag nanocomposite was characterized using various analytical techniques. The anticancer properties of the rGO–Ag nanocomposite were evaluated using a series of assays such as cell viability, lactate dehydrogenase leakage, reactive oxygen species generation, cellular levels of malonaldehyde and glutathione, caspase-3 activity, and DNA fragmentation in ovarian cancer cells (A2780).ResultsAgNPs with an average size of 20 nm were uniformly dispersed on graphene sheets. The data obtained from the biochemical assays indicate that the rGO–Ag nanocomposite significantly inhibited cell viability in A2780 ovarian cancer cells and increased lactate dehydrogenase leakage, reactive oxygen species generation, caspase-3 activity, and DNA fragmentation compared with other tested nanomaterials such as graphene oxide, rGO, and AgNPs.ConclusionT. amurensis plant extract-mediated rGO–Ag nanocomposites could facilitate the large-scale production of graphene-based nanocomposites; rGO–Ag showed a significant inhibiting effect on cell viability compared to graphene oxide, rGO, and silver nanoparticles. The nanocomposites could be effective non-toxic therapeutic agents for the treatment of both cancer and cancer stem cells.

Comparative Aspects of BRAF Mutations in Canine Cancers.

Activating mutations of the BRAF gene lead to constitutive activation of the MAPK pathway. The characterization and discovery of BRAF mutations in a variety of human cancers has led to the development of specific inhibitors targeting the BRAF/MAPK pathway and dramatically changed clinical outcomes in BRAF-mutant melanoma patients. Recent discovery of BRAF mutation in canine cancers underscores the importance of MAPK pathway activation as an oncogenic molecular alteration evolutionarily conserved between species. A comparative approach using the domestic dog as a spontaneous cancer model will provide new insights into the dysregulation of BRAF/MAPK pathway in carcinogenesis and facilitate in vivo studies to evaluate therapeutic strategies targeting this pathway’s molecules for cancer therapy. The BRAF mutation in canine cancers may also represent a molecular marker and therapeutic target in veterinary oncology. This review article summarizes the current knowledge on BRAF mutations in human and canine cancers and discusses the potential applications of this abnormality in veterinary oncology.

Targeting IP3‐dependent Calcium Signaling Enhances Sorafenib Lethality for Hepatoma via ER Stress Related Apoptosis

IRE1, hallmark of endoplasmic reticular (ER) stress, has been previously shown as a prosurvival molecule in cancer therapy by multikinase inhibitor sorafenib. However, given that the detail mechanism by which trigger IRE1 to impair the sorafenib lethality remains unclear, the present study was designed to clarify whether calcium signaling was involved in this pharmacologic process in HCC. Firstly, under confocal microscopy, sorafenib (100μM) was found to drive a dramatic intracellular calcium mobilization in HepG2 cells. Secondly, western blotting shew that IRE1 was significantly increased in cells either in the presence of sorafenib (20μM) or directly addition of low dose of CaCl2(0.4mM) for 24h. Interestingly, while using 2‐Aminoethyl diphenylborinate (2‐APB) (75μM), the inhibitor of IP3 receptor, to blunt the sorafenib induced‐calcium release in HepG2 cells, the increment of IRE1 and its activity evoked by sorafenib were substantially attenuated in these cells. Meanwhile, flow cytometry shew that 2‐APB and sorafenib(20μM) amazingly caused a synergistic effect versus sorafenib alone with about 50‐60% augmentation of apoptosis. Taken together, IP3‐elicited Ca2+ releaselikely plays a critical role in the sorafenib up‐regulating IRE1 activity and that is emerging as a novel potential target for anti‐HCC therapeutic strategy in the future.(Supported by NSFC Grants 881172260, 81360077 to G.Z. and 81102495 to L.C.)

Novel Ubiquitin-derived High Affinity Binding Proteins with Tumor Targeting Properties

Targeting effector molecules to tumor cells is a promising mode of action for cancer therapy and diagnostics. Binding proteins with high affinity and specificity for a tumor target that carry effector molecules such as toxins, cytokines, or radiolabels to their intended site of action are required for these applications. In order to yield high tumor accumulation while maintaining low levels in healthy tissues and blood, the half-life of such conjugates needs to be in an optimal range. Scaffold-based binding molecules are small proteins with high affinity and short systemic circulation. Due to their low molecular complexity, they are well suited for combination with effector molecules as well as half-life extension technologies yielding therapeutics with half-lives adapted to the specific therapy. We have identified ubiquitin as an ideal scaffold protein due to its outstanding biophysical and biochemical properties. Based on a dimeric ubiquitin library, high affinity and specific binding molecules, so-called Affilin® molecules, have been selected against the extradomain B of fibronectin, a target almost exclusively expressed in tumor tissues. Extradomain B-binding molecules feature high thermal and serum stability as well as strong in vitro target binding and in vivo tumor accumulation. Application of several half-life extension technologies results in molecules of largely unaffected affinity but significantly prolonged in vivo half-life and tumor retention. Our results demonstrate the utility of ubiquitin as a scaffold for the generation of high affinity binders in a modular fashion, which can be combined with effector molecules and half-life extension technologies.

Organo-Metallic Compounds: Novel Molecules in Cancer Therapy

The discovery of cis-platin in the treatment of cancer there has been a considerable exploration on the antitumoral activity of other transition metal complexes. One of the main problems about the application of transition metal complexes for chemotherapy is their potential toxicity. Recently the attention has been focused on titanium based complexes, which could have significant potential effect against solid tumor. The advantage of Ti (IV) complexes is their relative biological compatibility, which mostly leads to mild and revisable side effects. However, the hydrolytic instability of known Ti(IV) complexes and formation of different species upon water addition makes their therapeutic application problematic, and raises a strong interest in the development of relatively stable Ti(IV) complexes with well defined hydrolytic behavior that demonstrate appreciable cytotoxic activity. Strong ligand binding is also of interest to avoid complete ligand stripping by transferrin, so that the ligand may be used as a target for structure–activity relationship investigations. Titanocene dichloride (Cp2TiCl2) shows an average antiproliferative activity in vitro and promising result in vivo. Recent work has been performed in developing therapeutic analogues of Cp2TiCl2 by varying the central metal, the labile ligands (Cl) and the biscyclopentadienyl moiety. In particular, small changes to the Cp ligand can strongly affect the hydrolytic stability and water solubility properties of the metallocenes and have an impact on the cytotoxic activity. In this review we want summarize the importance of different organo-mettalic compounds in cancer therapy with focus on possible structure modification.

Withanone as an inhibitor of survivin: A potential drug candidate for cancer therapy

Survivin, the smallest inhibitor of apoptosis protein, which has been reported to be highly expressed in almost all known cancers, plays a dual role in survival as well as the proliferation of cancer cells. It inhibits apoptosis by inhibiting caspases as well as facilitating mitosis by becoming a part of chromosomal passenger complex through its BIR5 domain. Docking studies carried out with herbal ligand withanone derived from roots of Withania somnifera have shown strong binding affinity of −19.1088kJ/mol with BIR5 domain of survivin and in turn interferes with inhibitory action against caspases and may lead to apoptosis. Binding of withanone at BIR5 domain of survivin may also interfere with chromosomal passenger complex and lead to halt the mitotic process within the cancer cell. Docking studies support various experimental outcomes and suggest withanone as a potential candidate molecule in cancer therapy.

Abstract 2103: XB05: a promising small molecule for cancer therapy.

Abstract XB05 is synthetic small molecule which was serendipitously discovered to have potent anticancer properties. XB05 displays selectivity for malignant cells over non-malignant cells. Human myeloid leukemia cells (U937) and human lung cancer cells (A549) treated with XB05 show an increase in oxidative stress, DNA damage and a mixed apoptotic and necrotic mechanism of cell death. We conducted a COMPARE analysis to search for correlations between XB05 response and gene expression using publicly available microarray studies of the National Cancer Institute 60 cell line panel. This analysis uncovered a correlation between cell death in response to XB05 and expression of the transcription factor, SOX9. We find that SOX9 is differentially expressed across a panel of human lung cancer cell lines. MTT proliferation and clonogenic cell survival assays suggest that SOX9 expression positively correlates with the sensitivity of lung cancer cells to XB05. Furthermore, we show that siRNA knockdown of SOX9 recapitulates the phenotype of XB05-induced cell death, and that SOX9 may be downregulated at the protein level by XB05 treatment. Overall, these data demonstrate potent activity of XB05 against human cancer cells via a novel mechanism of action, and a potential role for SOX9 as a biomarker for selection and monitoring of therapy with XB05. Citation Format: Francesca R. Salipur, Elsa M. Reyes Reyes, Bo Xu, Ned Smith, Jian Cai, Gerald B. Hammond, Paula J. Bates. XB05: a promising small molecule for cancer therapy. [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 2103. doi:10.1158/1538-7445.AM2013-2103

APPLICATIONS OF SUPPORT VECTOR MACHINES AS A ROBUST TOOL IN HIGH THROUGHPUT VIRTUAL SCREENING

Chemical space is enormously huge but not all of it is pertinent for the drug designing. Virtual screening methods act as knowledge-based filters to discover the coveted novel lead molecules possessing desired pharmacological properties. Support Vector Machines (SVM) is a reliable virtual screening tool for prioritizing molecules with the required biological activity and minimum toxicity. It has to its credit inherent advantages such as support for noisy data mainly coming from varied high-throughput biological assays, high sensitivity, specificity, prediction accuracy and reduction in false positives. SVM-based classification methods can efficiently discriminate inhibitors from non-inhibitors, actives from inactives, toxic from non-toxic and promiscuous from non-promiscuous molecules. As the principles of drug design are also applicable for agrochemicals, SVM methods are being applied for virtual screening for pesticides too. The current review discusses the basic kernels and models used for binary discrimination and also features used for developing SVM-based scoring functions, which will enhance our understanding of molecular interactions. SVM modeling has also been compared by many researchers with other statistical methods such as Artificial Neural Networks, k-nearest neighbour (kNN), decision trees, partial least squares, etc. Such studies have also been discussed in this review. Moreover, a case study involving the use of SVM method for screening molecules for cancer therapy has been carried out and the preliminary results presented here indicate that the SVM is an excellent classifier for screening the molecules.

The HVEM network: new directions in targeting novel costimulatory/co-inhibitory molecules for cancer therapy

The regulation of the immune system is controlled by many cell surface receptors. A prominent representative is the 'molecular switch' HVEM (herpes virus entry mediator) that can activate either proinflammatory or inhibitory signaling pathways. HVEM ligands belong to two distinct families: the TNF-related cytokines LIGHT and lymphotoxin-α, and the Ig-related membrane proteins BTLA and CD160. HVEM and its ligands have been involved in the pathogenesis of various autoimmune and inflammatory diseases, but recent reports indicate that this network may also be involved in tumor progression and resistance to immune response. Here we summarize the recent advances made regarding the knowledge on HVEM and its ligands in cancer cells, and their potential roles in tumor progression and escape to immune responses. Blockade or enhancement of these pathways may help improving cancer therapy.

Effects of STAT3 Gene Silencing and Rapamycin on Apoptosis in Hepatocarcinoma Cells

The PI3K/Akt/mTOR and JAK/STAT3 signaling pathways are important for regulating apoptosis, and are frequently activated in cancers. In this study, we targeted STAT3 and mTOR in human hepatocellular carcinoma Bel-7402 cells and examined the subsequent alterations in cellular apoptosis. The expression of STAT3 was silenced with small interfering RNA (siRNA)-expressing plasmid. The activity of mTOR was inhibited using rapamycin. Following treatment, Annexin V/propidium iodide staining followed by flow cytometry and Hoechst33258 immunofluorescence staining was used to examine cellular apoptosis. JC-1 staining was used to monitor depolarization of mitochondrial membrane (ΔΨm). Furthermore, the expression of activated caspase 3 protein was analyzed by Western blotting. Compared to non-treated or control siRNA-transfected cells, significantly higher levels of apoptosis were detected in siSTAT3-transfected or rapamycin-treated cells (P < 0.05), which was further enhanced in cells targeted for both molecules (P < 0.05). The pro-apoptotic effects were accompanied with concomitant depolarization of mitochondrial membrane and up-regulation of activated caspase 3. Combined treatments using rapamycin and STAT3 gene silencing significantly increases apoptosis in Bel-7402 cells, displaying more dramatic effect than any single treatment. This study provides evidence for targeting multiple molecules in cancer therapy.