Anti-Cancer Agents in Medicinal Chemistry (Formerly Current Medicinal Chemistry - Anti-Cancer Agents) - Volume 19, Issue 6, 2019

Background: Cancer, as one of the most dangerous disease, causes millions of deaths every year. The main reason is the absence of an effective and thorough treatment. Drug delivery systems have significantly reduced the side-effect of chemotherapy. Combined with nanotechnology, smart drug delivery systems including many different nanoparticles can reduce the side-effect of chemotherapy better than traditional drug delivery systems. Methods: In this article, we will describe in detail the different kinds of nanoparticles and their mechanisms emphasizing the triggering factors in drug delivery. Besides, the application of smart drug delivery systems in imaging will be introduced. Results: Combined with nanotechnology, smart drug delivery systems including many different nanoparticles can reduce the side-effect of chemotherapy better than traditional drug delivery systems. Conclusion: Despite considerable progress in nanoparticle research over the past decade, such as smart drug delivery systems for the treatment of cancer, molecular imaging probes and the like. The range of nanoparticles used in multifunction systems for imaging and drug delivery continues to grow and we expect this dilatation to continue. But to make nanoparticles truly a series of clinical products to complement and replace current tools, constant exploration efforts and time are required. Overall, the future looks really bright.

Background: Pemetrexed (PMT) is a multitargeted antifolate agent that is used for treating patients with Non-Small Cell Lung Cancer (NSCLC). However, patients have presented clinical responses of drug resistance to PMT. Objective: This study aimed to explore the underlying mechanisms of PMT resistance in NSCLC cells. Methods: PMT-resistant NCI-H460/PMT cells were established by treating with PMT in a concentrationescalation manner. MTT assay and colony formation were performed to detect cell proliferation. Immunofluorescence was used to detect the expression of Ki-67. Transwell assay was performed to measure cell migration ability. qPCR and Western blot were used to detect the mRNA and protein expression levels of indicated genes. Small interfering RNAs (siRNA) were used to knockdown ATP binding cassette subfamily B member 1 (ABCB1) and Thymidylate Synthase (TYMS). Results: This study showed that compared with the parental cells, the NCI-H460/PMT cells displayed weakened proliferation and enhanced cell mobility. In addition, the NCI-H460/PMT cells demonstrated cellular senescence, which might result in PMT resistance. The NCI-H460/PMT cells exhibited cross-resistance to other chemotherapeutics, including fluorouracil, paclitaxel, doxorubicin, etoposide and gemcitabine, possibly because of the upregulated expression of ABCB1. However, the ABCB1 knockdown by siRNA failed to eradicate PMT resistance. Moreover, TYMS, a target of PMT, was obviously upregulated in the resistant cells. The genetic silence of TYMS partially abrogated PMT resistance, suggesting that the overexpression of TYMS was a key resistant mechanism of PMT. Conclusion: The overexpression of TYMS was an important resistance mechanism of PMT for KRAS-mutated NCI-H460 cells. Cross-resistance to other chemotherapeutics should be considered in addressing PMT resistance.

Background: Multi Drug Resistance (MDR) is the phenomenon that cancers develop resistance to majority of chemotherapy drugs and is a serious obstacle to the treatment for Hepatocellular Carcinoma (HCC). Polo-Like Kinase 1 (PLK1) is a serine/threonine kinase associated with tumor growth and clinical prognosis in HCC and BI2536 is its potent inhibitor with IC50 of 0.83nM. Aims: To test whether the down-regulation of PLK1 by its inhibitor BI2536 would have beneficial effects on the reversal of MDR in HCC cells. Methods: The CCK-8 assay was used to determine the viability of HepG2/ADM and SMMC7721/ADM cells and their parental cells treated with BI2536. Then animal model studies were performed. Cell invasion assay and wound healing assay were used to determine the invasion ability and motility. Flow cytometric was used to test the apoptosis induced by BI2536. Western blot and quantitative real-time PCR were performed to test the change of expression of MDR and apoptosis-related gene. Results: BI2536 down-regulated the expression of PLK1 protein and mRNA specifically. BI2536 can significantly reduce IC50 for ADM and other drugs in ADM-resistant HCC cells. Meanwhile, it inhibited cell viability, proliferation, and invasion, and induced cell cycle arrest and apoptosis in HCC cells with MDR. Conclusion: Our results suggest that PLK1 inhibitor BI2536 can re-sensitize HCC cancer cell with MDR through induction of apoptosis. Thus, PLK1 inhibitor BI2536 may act as an effective chemotherapeutic drug in the clinical treatment of HCC patients with MDR.

Background: Histone deacetylase inhibitors (HDACIs) have got immense importance as promising drugs for cancer treatment as these inhibitors regulate cellular differentiation, gene expression, cell cycle arrest and apoptosis. The current study investigates the effect of the hybrid-polar HDACI m-carboxycinnamic acid bishydroxyamide (CBHA) on the growth of human pancreatic adenocarcinoma cells, using the cell line MIA PaCa- 2 as an in vitro model. Methods: Following CBHA treatment of the MIA PaCa-2 cells, we characterized the effect of CBHA by in vitro cytotoxicity evaluation, clonogenic assay, cell cycle analysis, immunoblotting for soluble and insoluble fractions of tubulin, immunofluorescence and caspase-3 assay. Results: We observed that the histone deacetylase inhibitor CBHA markedly impaired growth of the pancreatic cancer cells by resulting in dose-dependent G2/M arrest, disruption of microtubule organization, induction of caspase-mediated apoptosis and in vitro suppression of HDAC6. Our study also shows that inhibition of HDAC6 by CBHA induced acetylation of α-tubulin. Conclusion: Together, our findings show that CBHA can be a potential plausible therapeutic that could be exploited for pancreatic cancer therapy.

Background: Some reports have demonstrated the role of the G Protein-coupled Estrogen Receptor (GPER) in growth and proliferation of breast cancer cells. Objective: In an effort to develop new therapeutic strategies against breast cancer, we employed an in silico study to explore the binding modes of tetrahydroquinoline 2 and 4 to be compared with the reported ligands G1 and G1PABA. Methods: This study aimed to design and filter ligands by in silico studies determining their Lipinski's rule, toxicity and binding properties with GPER to achieve experimental assays as anti-proliferative compounds of breast cancer cell lines. Results: In silico studies suggest as promissory two tetrahydroquinoline 2 and 4 which contain a carboxyl group instead of the acetyl group (as is needed for G1 synthesis), which add low (2) and high hindrance (4) chemical moieties to explore the polar, hydrophobic and hindrance effects. Docking and molecular dynamics simulations of the target compounds were performed with GPER to explore their binding mode and free energy values. In addition, the target small molecules were synthesized and assayed in vitro using breast cancer cells (MCF-7 and MDA-MB-231). Experimental assays showed that compound 2 decreased cell proliferation, showing IC50 values of 50μM and 25μM after 72h of treatment of MCF-7 and MDA-MB-231 cell lines, respectively. Importantly, compound 2 showed a similar inhibitory effect on proliferation as G1 compound in MDA-MB-231 cells, suggesting that both ligands reach the GPER-binding site in a similar way, as was demonstrated through in silico studies. Conclusion: A concentration-dependent inhibition of cell proliferation occurred with compound 2 in the two cell lines regardless of GPER.

Background: DNA methylation, which acts as an expression regulator for multiple Tumor Suppressor Genes (TSGs), is believed to play an important role in Nasopharyngeal Carcinoma (NPC) development. Methods: We compared the effects of 5-aza-2-deoxycytidine (decitabine, DAC) on gene expression using RNA sequencing in NPC cells. Results: We analyzed Differentially Expressed Genes (DEGs) in NPC cells using DAC demethylation treatment and found that 2182 genes were significantly upregulated (≥ 2-fold change), suggesting that they may play a key role in cell growth, proliferation, development, and death. For data analysis, we used the Gene Ontology database and pathway enrichment analysis of the DEGs to discover differential patterns of DNA methylation associated with changes in gene expression. Furthermore, we evaluated 74 methylated candidate TSGs from the DEGs in NPC cells and summarized these genes in several important signaling pathways frequently disrupted by promoter methylation in NPC tumorigenesis. Conclusion: Our study analyzes the DEGs and identifies a set of genes whose promoter methylation in NPC cells is reversed by DAC. These genes are potential substrates of DNMT inhibitors and may serve as tumor suppressors in NPC cells.

Background: The effect of estrogen has been traditionally studied through the modulation of its alpha and beta nuclear receptors; however, the G Protein-Coupled Estrogen Receptor (GPER) has been recently involved in the pathology of numerous tumors. Although the study of GPER in cervical cancer has begun, its contribution still remains to be completely evaluated. Objective: The purpose of this work was to determine the expression of this receptor in different degrees of cervical lesions and whether the stimulation with its specific agonist (G-1) modulated mechanisms of cell survival or cell death in cervical cancer cells. Methods: Sections of 44 formalin-fixed paraffin-embedded blocks from patients were analyzed by automated immunohistochemistry. After the stimulation with G-1, proliferation was evaluated by the xCELLigence technology, the integrity of the mitochondrial membrane permeability by MitoCaptureTM fluorescence staining, apoptosis by flow cytometry, and senescence by the senescence-associated β-galactosidase kit. Results: GPER was widely expressed in cervical cancer but not in its precursor lesions. The staining was predominantly cytoplasmic, although it was also important in the nucleus of the epithelial cells. G-1 inhibited proliferation, decreased the mitochondrial permeability, and increased the percentage of apoptosis in SiHa, HeLa, and C-33A. Only in C-33A, an increase of the cells in necrosis was observed, whereas SiHa was the only cell line in which senescence was evidenced. Conclusion: GPER is a receptor associated with cervical cancer that inhibits the growth and induces different mechanisms of death in cells derived from uterine cervical cancer. It suggests that GPER can be considered a pharmacological target that prevents the development of cervical carcinogenesis.

Background: Astaxanthin (AST) shows a large range of beneficial effects together with anti-cancer and antioxidation properties. Human Serum Albumin (HSA) is the most abundant protein in blood plasma which plays the role of a depot and transport protein for many exogenous compounds. However, whether HSA could enhance AST-induced cytotoxic effects in human ovarian cancer cells has not been examined to date. Objective: This study aims to explore the anticancer effect and the molecular mechanism of AST combine with HSA induced cytotoxicity in ovarian cancer SKOV3 cells. Methods: The ovarian cancer SKOV3 cells were treated by AST combined with HSA to study the effects of cell proliferation, cell morphology, cell cycle arrest, related protein expression, nuclear transfer, cell migration, and drug-resistant. Results: Our data confirmed that AST+HSA treatment enhanced the anticancer effects of AST, arrested G1 phase cell cycle and induced apoptosis in SKOV3 cells. AST+HSA induced apoptosis via mitochondrial apoptotic pathways was related to the increased ratio of Bcl-2/Bax and activation of caspase-3. Besides, exposure of cells to AST+HSA triggered the inactivation of NF-ΚB and activation p53 and MAPKs signaling pathways. Furthermore, AST+HSA significantly overcome the drug-resistant and inhibited the migration of SKOV3 cells. Conclusion: AST combined treatment with HSA considerably inhibited NF-ΚB expression and translocation to nucleus, thereby improving the AST-induced cytotoxic effect on SKOV3 cells. These findings may provide rationale to combine AST with HSA for the treatment of ovarian cancer.

Background: Traditional chemotherapeutics of low-molecular weight diffuse passively across intact membrane structures of normal healthy cells found in tissues and organ systems in a non-specific unrestricted manner which largely accounts for the induction of most sequelae which restrict dosage, administration frequency, and duration of therapeutic intervention. Molecular strategies that offer enhanced levels of potency, greater efficacy and broader margins-of-safety include the discovery of alternative candidate therapeutics and development of methodologies capable of mediating properties of selective “targeted” delivery. Materials and Methods: The covalent immunopharmaceutical, dexamethasone-(C21-phosphoramidate)-[anti- EGFR] was synthesized utilizing organic chemistry reactions that comprised a multi-stage synthesis regimen. Multiple forms of analysis were implemented to vadliate the successful synthesis (UV spectrophotometric absorbance), purity and molar-incorporation-index (UV spectrophotometric absorbance, chemical-based protein determination), absence of fragmentation/polymerization (SDS-PAGE/chemiluminescent autoradiography), retained selective binding-avidity of IgG-immunoglobulin (cell-ELISA); and selectively “targeted” antineoplastic cytotoxicity (biochemistry-based cell vitality/viability assay). Results: The botanicals carnosic acid, ginkgolide-B and tangeretin, each individually exerted maximum antineoplastic cytotoxicity levels of 58.1%, 5.3%, and 41.1% respectively against pulmonary adenocarcinoma (A549) populations. Dexamethasone-(C21-phosphoramidate)-[anti-EGFR] formulated at corticosteroid/ glucocorticoid equivalent concentrations produced anti-neoplastic cytotoxicity at levels of 7.7% (10-9 M), 26.9% (10-8 M), 64.9% (10-7 M), 69.9% (10-6 M) and 73.0% (10-5 M). Ccarnosic acid, ginkgolide-B and tangeretin in simultaneous dual-combination with dexamethasone-(C21-phosphoramidate)-[anti-EGFR] exerted maximum anti-neoplastic cytotoxicity levels of 70.5%, 58.6%, and 69.7% respectively. Discussion: Carnosic acid, ginkgolide-B and tangeretin botanicals exerted anti-neoplastic cytotoxicity against pulmonary adenocarcinoma (A549) which additively contributed to the anti-neoplastic cytotoxic potency of the covalent immunopharmaceutical, dexamethasone-(C21-phosphoramidate)-[anti-EGFR]. Carnosic acid and tangeretin were most potent in this regard both individually and in dual-combination with dexamethasone-(C21- phosphoramidate)-[anti-EGFR]. Advantages and attributes of carnosic acid and tangeretin as potential monotherapeutics are a wider margin-of-safety of conventional chemotherapeutics which would readily complement the selective “targeted” delivery properties of dexamethasone-(C21-phosphoramidate)-[anti-EGFR] and possibly other covalent immunopharmaceuticals in addition to providing opportunities for the discovery of combination therapies that provide heightened levels of anti-neoplastic efficacy.

Background: Cancer is one of the most fatal diseases across the world and it was reported that 90% of cancer fatality depends on its angiogenesis potential. Black seed or Nigella sativa L. is a medicinal plant native to southwest Asia. N. sativa has been used for medicinal purposes for centuries and predominantly has bioactive components like Thymoquinone, which is used as a candidate for anti-cancer and anti-angiogenesis drugs. Methods: Callus was induced from leaf tissue, after that alcoholic extracts were prepared from three-month-old calluses. Thymoquinone content was measured by HPLC methods. AGS cell line was cultured and treated with standard Thymoquinone and extracts from callus. Then, cell proliferation, expression of angiogenic factor (VEGF-A gene), and apoptosis test were done by MTT assay, real-time PCR and Annexin-v kit, respectively. Results: HPLC found the maximum amount of Thymoquinone in the extract of leaf calluses, which were grown in the dark. MTT assay revealed that particular doses of extracts reduced cell proliferation. Real-time and Fluorescence- Activated Cell Sorting (FACS) results demonstrated that standard Thymoquinone and callus extracts down-regulated the VEGF-A gene expression, and all three induced apoptosis in the AGS cell line. Conclusion: It has been shown that TQ has pro-apoptotic and anti-metastatic effects on stomach cancer cell line, and these properties can introduce it as an anti-cancer drug in the near future.

Background: Triple Negative Breast Cancer (TNBC) tends to be more aggressive than other types of breast cancer. Resistance to chemotherapy is a major obstacle hence there is a significant need for new antineoplastic drugs with multi-target potency. Numerous Benzoisoxazole moieties have been found to possess a broad spectrum of pharmacological activities. In the present study, we have synthesized 9 novel derivatives of Benzisoxazole 7(a-i) and screened them for their biological potential. Methods: Chemical synthesis, Mass spectrometry (HRMS), cell proliferation and cytotoxicity assay, wound healing assay, flow cytometry and nuclear staining. Angio-inhibitory activity assessed by corneal micropocket assay and in vivo peritoneal angiogenesis assay. Results: The Benzisoxazole derivatives 7(a-i) were synthesized and screened for their biological potency by both in vitro and in vivo experimental models. Among the series, compound 3-(1-((3-(3(Benzyloxy)-4-methoxyphenyl)- 4,5-dihydroisoxazole-5-yl)methyl)piperidine-4-yl)6-fluorobenzo[d] isoxazole (7e) was found to be most promising, with an average IC50 value of 50.36 ± 1.7 μM in MTT assay and showed 81.3% cell death. The compound 7e also showed 60-70% inhibition on a recombinant Metastasis-Associated protein (MTA1) induced proliferation and cell migration in MDAMB-231 cells, which is known to play a major role in angiogenesis. The anti-tumour studies inferred the regression of tumour activity. This was due to inhibition of neovascularization and evoking apoptosis process as assessed by corneal vascularization, peritoneal angiogenesis and apoptotic hallmarks in 7e treated cells. Conclusion: These findings not only show the biological efficacy of compound 7e but it is also an effective beginning to explore the mechanism of metastasis and cancer therapy strategy targeting MTA1. The observed biological activity makes compound 7e an attractive drug candidate.