Generation Of Anticancer Drugs Research Articles

Polyoxometalates as Potential Next-Generation Metallodrugs in the Combat Against Cancer.

Polyoxometalates (POMs) are an emerging class of inorganic metal oxides, which over the last decades demonstrated promising biological activities by the virtue of their great diversity in structures and properties. They possess high potential for the inhibition of various tumor types; however, their unspecific interactions with biomolecules and toxicity impede their clinical usage. The current focus of the field of biologically active POMs lies on organically functionalized and POM‐based nanocomposite structures as these hybrids show enhanced anticancer activity and significantly reduced toxicity towards normal cells in comparison to unmodified POMs. Although the antitumor activity of POMs is well documented, their mechanisms of action are still not well understood. In this Review, an overview is given of the cytotoxic effects of POMs with a special focus on POM‐based hybrid and nanocomposite structures. Furthermore, we aim to provide proposed mode of actions and to identify molecular targets. POMs are expected to develop into the next generation of anticancer drugs that selectively target cancer cells while sparing healthy cells.

Abstract 5021: Assessment of RAS-RAF-MAPK pathway perturbations and PD-L1 expression in an Isogenic 3D tissue-culture model of drug-resistant melanoma

Abstract Development of the next generation of anti-cancer drugs, biologics, and immunotherapies is currently hampered by extremely poor success rates of seemingly promising experimental therapies in human clinical trials. This poor success rate is partially due to a lack of biologically relevant cancer model systems. The 2D culture system, while relatively inexpensive and easy to use, may not always be adequately representative of the tumor microenvironment. Conversely, the use of animal models is costly and time consuming. The 3D culture system is able to represent biologically relevant complexities such as cell-cell communication, differential proliferation rates, and compound penetration. Combining 3D culture with CRISPR/Cas9 genome editing technologies can generate powerful cell-based model systems with both physiological relevance and well-controlled genetic and drug-susceptibility profiles. These new models will enable the investigation of specific molecular mechanisms, biofunctional outcomes of newly identified genetic alternations, and targeted therapeutic drug responses within a more biologically intricate context. In this study, we use CRISPR/Cas9 to generate isogenic drug-resistant melanoma models that can be used as either 2D or 3D cancer models, as well as for studies of the acquired drug resistance. Two different models were generated starting with the BRAF-inhibitor sensitive A375 melanoma cell line. We introduced either the NRASQ61K or the KRASG13D point mutations, both of which are known to confer BRAF inhibitor resistance and are commonly encountered in BRAF-resistant tumor samples. We then assessed the susceptibility of these new isogenic lines to BRAF inhibitors in both 2D and 3D model systems. We also determined the specific effect of these point mutations on the RAS-RAF-MAPK signaling pathway, a key component of cell-cycle escape and tumor proliferation. Furthermore, we assessed the impact of these mutations on the expression of Programmed Death-Ligand 1 (PD-L1), which recent advances in cancer immunology have directly linked to cancer immune evasion and poor clinical outcomes. Our results show that A375 melanoma isogenic cells carrying KRASG13D have dramatically increased EGFR expression levels, while isogenic cells carrying NRASQ61K have constant activation of the MEK-ERK pathway. We also found a significant increase in PD-L1 expression in KRASG13D, but not in NRASQ61K A375 cells, indicating that expression of PD-L1 is directly linked to KRAS mutation or the downstream effects thereof, rather than to a general increase in RAS-RAF-MAPK signaling. These results have direct implications for cancer immune checkpoint molecule studies, and highlight the utility of these isogenic melanoma models in both 2D and 3D applications in research and development of novel anti-cancer drugs and combination therapies. Citation Format: Elizabeth Turner, Luping Chen, Johnathan Haag, Alexei Miagkov, Lysa-Anne Volpe, Metewo Selase Enuameh, Robert Newman, Fang Tian. Assessment of RAS-RAF-MAPK pathway perturbations and PD-L1 expression in an Isogenic 3D tissue-culture model of drug-resistant melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5021.

Determining the Crystal Structures of Peptide Analogs of Boronic Acid in the Absence of Single Crystals: Intricate Motifs of Ixazomib Citrate Revealed by XRPD Guided by ss-NMR

Uncertainties in the structure determination of peptide analogs of boronic acid, exacerbated by the many coordination modes of boron, represent an obstacle in understanding their role in living organisms and thus also in developing the next generation of anticancer drugs. For that reason, we present here a general experimental-computational strategy allowing structure determination of complex boronic acid derivatives with extensive conformational variability. We demonstrate successful solution of the crystal structures of two nonsolvated polymorphs of ixazomib citrate directly from synchrotron powder diffraction data, which is challenging because the two molecules in the asymmetric unit cell that exhibit 32 degrees of conformational freedom push the limits of current solution procedures. We used a novel two-step Rietveld refinement based on DFT-D restraints to improve information quality derived from powder diffraction data to be comparable with that of single-crystal solutions. NMR crystallography was ap...

Hydroxycamptothecin mediates antiproliferative effects through apoptosis and autophagy in A549 cells.

Hydroxycamptothecin (HCPT) represents a new generation of anticancer drugs, with almost no side effects when used for the treatment of a number of types of cancer. Autophagy is becoming recognized as an important biological mechanism in human cancer, including lung cancer. However, the involvement of autophagy in the antiproliferative effects of HCPT on lung cancer remains unclear. In the present study, A549 cells, an accepted model of non-small cell lung cancer (NSCLC) cells, were employed. It was demonstrated that HCPT was able to suppress proliferation and induce apoptosis and autophagy in A549 cells. The molecular mechanism underlying HCPT-induced cell death was attributed to apoptosis and autophagy. Furthermore, it was demonstrated that an autophagy inhibitor, 3-methyladenine, accelerated HCPT-induced cell death in A549 cells. The results of the present study may lead to a deeper understanding of the molecular mechanism by which HCPT regulates NSCLC A549 cells. These results highlight the potential use of autophagy inhibitors in combination with traditional chemotherapy drugs for the treatment of lung cancer.

Structure-based pharmacophore design and virtual screening for novel potential inhibitors of epidermal growth factor receptor as an approach to breast cancer chemotherapy.

Cancer cells are described with features of uncontrolled growth, invasion and metastasis. The epidermal growth factor receptor subfamily of tyrosine kinases (EGFR-TK) plays a crucial regulatory role in the control of cellular proliferation and progression of various cancers. Therefore, its inhibition might lead to the discovery of a new generation of anticancer drugs. In the present study, structure-based pharmacophore modeling, molecular docking and molecular dynamics simulations were applied to identify potential hits, which exhibited good inhibition on the proliferation of MCF-7 breast cancer cell line and favorable binding interactions on EGFR-TK. Selected compounds were examined for their anticancer activity against the Michigan Cancer Foundation-7 (MCF-7) breast cancer cell line which overexpresses EGFR using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium reduction assay. Compounds 1 and 2, with an isoindoline-1-one core, induced significant inhibition of breast cancer cells proliferation with IC[Formula: see text] values 327 and 370 nM, respectively.

Regulation of cAMP and GSK3 signaling pathways contributes to the neuronal conversion of glioma.

Glioma is the most malignant type of primary central nervous system tumors, and has an extremely poor prognosis. One potential therapeutic approach is to induce the terminal differentiation of glioma through the forced expression of pro-neural factors. Our goal is to show the proof of concept of the neuronal conversion of C6 glioma through the combined action of small molecules. We investigated the various changes in gene expression, cell-specific marker expression, signaling pathways, physiological characteristics, and morphology in glioma after combination treatment with two small molecules (CHIR99021, a glycogen synthase kinase 3 [GSK3] inhibitor and forskolin, a cyclic adenosine monophosphate [cAMP] activator). Here, we show that the combined action of CHIR99021 and forskolin converted malignant glioma into fully differentiated neurons with no malignant characteristics; inhibited the proliferation of malignant glioma; and significantly down-regulated gene ontology and gene expression profiles related to cell division, gliogenesis, and angiogenesis in small molecule–induced neurons. In vivo, the combined action of CHIR99021 and forskolin markedly delayed neurological deficits and significantly reduced the tumor volume. We suggest that reprogramming technology may be a potential treatment strategy replacing the therapeutic paradigm of traditional treatment of malignant glioma, and a combination molecule comprising a GSK3 inhibitor and a cAMP inducer could be the next generation of anticancer drugs.

Raman scattering of Cisplatin near silver nanoparticles

The Raman scattering of Cisplatin (the first generation of anticancer drugs) has been studied. In the presence of silver nanoparticles, strong modifications of Raman spectra have been observed. The Raman frequencies have been shifted and the line profiles are broadened. We develop a theoretical model to explain the observed features of the Raman scattering. The model takes into account self-consistently the interaction of molecules with surface plasmonic waves excited in the silver nanoparticles, and it provides a qualitative agreement with the observed Raman spectra. We have demonstrated that the using silver nanoparticles can increase sensitivity of the technique, and potentially it has a broader range of applications to both spectroscopy and microscopy.

A theoretical investigation on bio-transformation of third generation anti-cancer drug Heptaplatin and its interaction with DNA purine bases

Heptaplatin is an approved platinum based cytostatic drug for the treatment of gastric cancers. The hydrolytic bio-transformation of Heptaplatin and the platination processes of guanine (G) and adenine (A) with resulting mono and di-aquated species of Heptaplatin have been investigated using density functional theory (DFT) combined with the conductor like dielectric continuum model (CPCM) approach, to spotlight the drug activation energy profiles and their binding mechanisms. The stationary points on the potential energy surfaces were fully optimized and characterized. The mono-functional binding of Heptaplatin, guanine as target over adenine due to electronic factors and more favorable hydrogen-bonds pattern.

Targeting energy metabolism of cancer cells: Combined administration of NCL-240 and 2-DG

Cancer cells increase their metabolism to produce the energy and biomolecules necessary for growth and proliferation. Thus, energy metabolism pathways may serve as targets for anti-cancer therapy. NCL-240 is a second generation anti-cancer drug belonging to the PITenins class of PI3K-Akt inhibitors. Our analysis suggested that NCL-240 caused disruptions in mitochondrial oxidative phosphorylation and up-regulated glycolysis, as evidenced by the loss of NMR peaks for the amino acid products derived from the TCA cycle along with presence of only lactate peaks and the loss of glucose peaks. NCL-240 was combined with 2-deoxy-d-glucose (2-DG) in early proof-of-concept studies on multiple cell lines. 2-DG enhanced cell death response to NCL-240 administration, with cytotoxicity results similar to those under hypoglycemic conditions. In further studies, NCL-240 encapsulated in phosphatidylcholine/cholesterol liposomes was combined with freely dissolved 2-DG. Cell cycle analysis of sensitive and resistant strains of A2780 cells treated with combinations of NCL-240/2-DG pointed to a G0/G1 phase arrest for 80–90% of the total, indicating an inability to grow and divide. Cytotoxicity studies with in vitro cancer cell monolayer models confirmed the results of cell cycle analysis. Significant improvements in cytotoxicity with combination treatments over control and individual treatments were seen in multiple cell lines. NCI/ADR-RES cancer cell spheroids further demonstrated the effectiveness of a NCL-240/2-DG combination.

Dianhydrogalactitol, a potential multitarget agent, inhibits glioblastoma migration, invasion, and angiogenesis

The complexity of cancer has led to single-target agents exhibiting lower-than-desired clinical efficacy. Drugs with multiple targets provide a feasible option for the treatment of complex tumors. Multitarget anti-angiogenesis agents are among the new generation of anticancer drugs and have shown favorable clinical efficacy. Dianhydrogalactitol (DAG) is a chemotherapeutic agent for chronic myeloid leukemia and lung cancer. Recently, it has been tested in phase II trials of glioblastoma treatment; however, mechanisms of DAG in glioblastoma have not been elucidated. Here we show that DAG could inhibit the migration and invasion of U251 cell line by inhibiting matrix metalloproteinase-2 (MMP2) expression. Furthermore, DAG could also inhibit tumor angiogenesis in vitro as well as in the zebrafish model. Mechanistic studies reveal that DAG inhibited both VEGFR2 and FGFR1 pathways. Our results suggest that DAG may be a potential multitarget agent that can inhibit tumor migration, invasion, and angiogenesis, and the anti-angiogenic effects may be involved in dual-suppression VEGF/VEGFR2 and FGF2/FGFR1 signal pathways.

Novel sulfonamide-containing 2-indolinones that selectively inhibit tumor-associated alpha carbonic anhydrases

Human carbonic anhydrases IX and XII are upregulated in many tumors and form a novel target for new generation anticancer drugs. Here we report the synthesis of novel 2-indolinone derivatives with the sulfonamide group as a zinc binding moiety. Enzyme inhibition assays confirmed that the compounds showed selectivity against hCA IX and XII over the widely distributed off-targets hCA I and II. Molecular modelling studies were performed to suggest modes of binding for these compounds.

AKT/mTOR signaling pathway is involved in salvianolic acid B-induced autophagy and apoptosis in hepatocellular carcinoma cells.

Chinese medicines are emerging as an attractive new generation of anticancer drugs. Here, we explored the impact of salvianolic acid B (SalB), the major water-soluble compounds of Danshen, on apoptosis and autophagy of human hepatocellular carcinoma cells (HCC). We also investigated the related molecular mechanisms. We found that SalB exhibits potent ability to inhibit HCC cells viability in a concentration-dependent manner, and to induce apoptosis via the mitochondrial apoptosis pathway. Additionally, SalB could also induce autophagy. Furthermore, pretreatment with the autophagy inhibitor chloroquine or 3-methyladenine showed the potential in attenuating the apoptosis rate induced by SalB. Mechanistically, SalB treatment inhibited the AKT/mTOR signaling cascade in vitro. Overexpression of AKT abolished the effects of SalB on HCC cells, suggesting a critical role of the AKT/mTOR signaling pathway in SalB-induced biological effects. Our results indicated that the mitochondrial pathway was involved in SalB-induced apoptosis of HCC cells. Moreover, the AKT/mTOR signaling pathway was involved in SalB-induced autophagy, which promoted apoptosis. This study may provide a promising strategy for using SalB as a chemotherapeutic agent for patients with HCC.

Selective killing of cancer cells by small molecules targeting heat shock stress response

HSF1 heat shock response has emerged as a valuable non-oncogenetic intervention point in targeted cancer therapy. Current reporter based high throughput screening has led to the discovery of several compounds or chemotypes that are effective in the growth inhibition of multiple cancer cell lines and relevant animal tumor models. However, some intrinsic limitations of reporter based assays can potentially lead to biased results. Using a previously validated high content image based assay, we performed a phenotypic screen targeting HSF1 heat shock pathway with a chemically diversified library of over 100,000 compounds. Several novel functional inhibitors of HSF1 pathway were identified with different chemotypes. Western blot analysis confirmed that selective compounds inhibit phosphorylation of HSF1, followed by reduced expression of HSP proteins. Moreover, HeLa cells stably transfected with HSF1 shRNA were more resistant to the compound treatment under lethal temperature than cells containing HSF1, validating HSF1 dependent mechanism of action. These compounds demonstrate nanomolar potency toward multiple cancer cell lines with relatively low cytotoxicity to normal cells. Further SAR and target identification study will pave the way for the potential development of next generation anticancer drugs.

Monofunctional Platinum (PtII) Compounds - Shifting the Paradigm in Designing New Pt-based Anticancer Agents.

Platinum (Pt)-based anticancer drugs, exemplified by cisplatin, are key components in combination chemotherapy. However, their effective use is hindered by toxicity and emergence of drug resistance. They bind to DNA and mainly form the Pt-GG diadduct, subsequently leading to apoptosis to mediate cell death. On the other hand, the Pt drug -proteins and -metabolites interactions, which involve the reaction between Pt and sulfur sites located in protein side chains and important bionucleophiles (e.g., glutathione), are responsible for the toxicity and drug resistance problem. Therefore, carefully designed coordinating ligands may provide the means of fine tuning the electronic environment around the core Pt atom and allow the resulting Pt compounds to bind with the DNA in a different manner. This may produce alternative cell death mechanisms in cancer cells, thereby circumventing Pt resistance. This article reviewed the recent development in monofunctional Pt complexes and their prospects in becoming a new generation of anticancer drugs.

Design of etoposide polymeric forms

The present work relates to the field of pharmacology and medicine, in particular, to a new generation of anticancer drugs based on biocompatible polymers containing etoposide as the drug substance. A polymeric composition containing a surfactant and cryoprotective agent in addition to polymer and drug substance was designed. The optimum formulation of the polymeric composition and process conditions for its preparation were selected. The samples of etoposide polymeric forms were found to exhibit in vitro cytotoxic activity against two human tumor cell lines, viz., MCF-7 breast adenocarcinoma and K562 myeloleukemia cell lines, which was either identical or higher than the activity of free etoposide. The samples of the PLGA-based polymeric form of etoposide exhibited the highest activity.

SKF-96365 activates cytoprotective autophagy to delay apoptosis in colorectal cancer cells through inhibition of the calcium/CaMKIIγ/AKT-mediated pathway

Store-operated Ca2+ entry (SOCE) inhibitors are emerging as an attractive new generation of anti-cancer drugs. Here, we report that SKF-96365, an SOCE inhibitor, exhibits potent anti-neoplastic activity by inducing cell-cycle arrest and apoptosis in colorectal cancer cells. In the meantime, SKF-96365 also induces cytoprotective autophagy to delay apoptosis by preventing the release of cytochrome c (cyt c) from the mitochondria into the cytoplasm. Mechanistically, SKF-96365 treatment inhibited the calcium/calmodulin-dependent protein kinase IIγ (CaMKIIγ)/AKT signaling cascade in vitro and in vivo. Overexpression of CaMKIIγ or AKT abolished the effects of SKF-96365 on cancer cells, suggesting a critical role of the CaMKIIγ/AKT signaling pathway in SFK-96365-induced biological effects. Moreover, Hydroxychloroquine (HCQ), an FDA-approved drug used to inhibit autophagy, could significantly augment the anti-cancer effect of SFK-96365 in a mouse xenograft model. To our best knowledge, this is the first report to demonstrate that calcium/CaMKIIγ/AKT signaling can regulate apoptosis and autophagy simultaneously in cancer cells, and the combination of the SOCE inhibitor SKF-96365 with autophagy inhibitors represents a promising strategy for treating patients with colorectal cancer.

Delivery of Constitutively Active Mutant MKK6(E) With TAT-OSBP Induces Apoptosis in Human Ovarian Carcinoma HO8910 Cells.

Biologically active peptides and proteins are novel agents that show promise in the development of anticancer drugs. Their relatively low cell permeability and poor tumor selectivity, however, impede their widespread applicability. In this study, we evaluated the tumor selectivity, cellular internalization, and biological activity of a cell-permeable ovarian cancer cell-specific therapeutic protein consisting of TAT-OSBP and constitutively active MKK6(E), an upstream kinase of the p38 signaling pathway that mediates cellular apoptosis. OSBP, a 7-amino-acid peptide with high affinity for human ovarian cancer HO8910 cells, was conjugated to the cell-penetrating peptide (TAT) to form a tumor-selective peptide (TAT-OSBP), which was further conjugated with EGFP or MKK6(E). Flow cytometry and fluorescent microscopy were performed to evaluate the tumor-targeted penetration of TAT-OSBP-EGFP. The inhibitory effects of TAT-OSBP-MKK6(E) were determined by cell proliferation and apoptosis assays. The internalization efficiency of TAT-OSBP-EGFP was significantly higher than that of TAT-EGFP. TAT-OSBP-EGFP selectively penetrated HO8910 cells. TAT-OSBP-MKK6(E) fusion protein inhibited cancer cell growth to varying degrees, with the highest level of inhibition in HO8910 cells. Moreover, TAT-OSBP-MKK6(E) significantly induced apoptosis of HO8910 cells. However, there was no significant difference in apoptosis in the normal ovarian epithelial cells treated with either TAT-OSBP-MKK6(E) or TAT-MKK6(E). Our results demonstrate that TAT-OSBP-MKK6(E) is a novel artificially designed molecule, which induces apoptosis and selectively targets human ovarian carcinoma HO8910 cells. Our study provides novel insights that may aid in the development of a new generation of anticancer drugs.

Are Synthetic Compounds that Silence the Liver-X-Receptor the Next Generation of Anti-cancer Drugs?

Glycolytic and lipogenic inhibitors have proven unsuccessful in cancer treatment strategies. In this issue of Cancer Cell, Flaveny and colleagues target the liver-X-receptor with an inverse agonist and show that key glycolytic and lipogenic genes are suppressed, leading to apoptosis of tumor cells without an effect on non-malignant cells.

New insights into the molecular and epigenetic effects of antitumor Pt(IV)-valproic acid conjugates in human ovarian cancer cells

Substitutionally inert Pt(IV) prodrugs, combining bioactive axial ligands with Pt(IV) derivatives of antitumor Pt(II) compounds, represent a new generation of anticancer drugs. The rationale behind these prodrugs is to release, by reductive elimination inside the cancer cell, an active Pt(II) drug which binds nuclear DNA as well as bioactive ligands that may potentiate toxic effects of the Pt(II) drugs by an independent pathway. Platinum prodrugs, such as Pt(IV) derivatives of cisplatin containing axial valproic acid (VPA) ligands, destroy cancer cells with greater efficacy than conventional cisplatin. These axial ligands were chosen because VPA inhibits histone deacetylase (HDAC) activity, thereby decondensing chromatin and subsequently increasing the accessibility of DNA within chromatin to DNA-binding agents. We examined the mechanism of cytotoxic activity of Pt(IV) derivatives of cisplatin with VPA axial ligands. Particular attention was paid to the role of the VPA ligand in these Pt(IV) prodrugs in the mechanism underlying their toxic effects in human ovarian tumor cells. We demonstrate that (i) treatment of the cells with these prodrugs resulted in enhanced histone H3 acetylation and decondensation of heterochromatin markedly more effectively than free VPA; (ii) of the total Pt inside the cells, a considerably higher fraction of Pt from the Pt(IV)-VPA conjugates is bound to DNA than from the conjugates with biologically inactive ligands. The results indicate that the enhanced cytotoxicity of the Pt(IV)-VPA conjugates is a consequence of several processes involving enhanced cellular accumulation, downregulation of HDACs and yet other biochemical processes (not involving HDACs) which may potentiate antitumor effects.

Abstract A57: Effects of pre-exposure to siRNA on silencing response: Do cells become resistant to siRNA silencing?

Abstract Development of resistance against cytotoxic effect of different generations of anticancer drugs is considered inevitable and remains a major concern for cancer therapy. An alternative therapeutic approach based on protein silencing via RNA interference (RNAi) have shown considerable promise; however, our information about the factors affecting siRNA efficiency and the limitations to their efficacy are limited. This study focused on determining the possibility of development of resistance against siRNA treatment as a result of repeated exposure to siRNA that could affect cellular internalization of the particles, silencing efficiency at mRNA level, and ultimately, the observed cellular response. Two approaches to siRNA exposure were undertaken: one involving a single high concentration exposure to eradicate siRNA-responsive cells, and one involving multiple exposures with a gradually increasing siRNA concentration to study the potential intracellular adaptations. Our results demonstrated a temporary decrease in silencing efficiency at mRNA level without significant alteration in siRNA cellular uptake. We conclude that cells would respond to repeated siRNA exposure in a similar fashion after a temporary initial alteration; however, we could not rule out the possibility of a change in phenotypical response (e.g., number of viable cells) as a result of adjustments in (or selection of cells reliant on) alternative pathways. Citation Format: Hamidreza Montazeri Aliabadi, Parvin Mahdipoor, Cezary Kucharsky, Nicole Chan, Hasan Uludag. Effects of pre-exposure to siRNA on silencing response: Do cells become resistant to siRNA silencing? [abstract]. In: Proceedings of the AACR Precision Medicine Series: Drug Sensitivity and Resistance: Improving Cancer Therapy; Jun 18-21, 2014; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(4 Suppl): Abstract nr A57.