Tested Cancer Cell Research Articles

New alkaloids from the stem bark of Cinnamomum bejolghota

Three new alkaloids, 3,4-bis(3,4-dimethoxyphenyl) pyridine (1), 1-(4-hydroxybenzyl)− 6-hydroxyisoquinoline) (2), and 4-(3,4-dimethoxyphenyl)− 2-methyl pyridine (3) along with six known compounds, spathulenol (4), 5,7-di-O-methyl-3′,4′-methylenedioxyflavan-3-ol (5), 3,4-dimethoxycinnamyl alcohol (6), 3,4-dimethoxycinnamaldehyde (7), veratric acid (8) and veratraldehyde (9), were isolated from the stem bark of Cinnamomum bejolghota (Buch.-Ham.) Sweet. Their structures were elucidated based on the interpretation of spectrometric and spectroscopic data (HRESI-MS, 1D- and 2D NMR) and by comparison with literature data for known compounds. Compounds 1–5 were tested for their cytotoxic activities against four human cancer cell lines (KB, MCF-7, HepG-2, and SK-LU1). Compound 1 showed moderate activity against KB, MCF7, HepG2 and SK-LU-1 cell lines with IC50 values ranging from 31.1 to 45.9 µM, while compounds 2–4 were inactive on all tested cancer cell lines up to a concentration of 128 μM and flavanol 5 showed a very weak cytotoxic activity.

Molecular mechanisms underlying the clinical efficacy of panobinostat involve Stochasticity of epigenetic signaling, sensitization to anticancer drugs, and induction of cellular cell death related to cellular stresses

Panobinostat, also known as Farydak®, LBH589, PNB, or panobinostat lactate, is a hydroxamic acid that has been approved by the Food and Drug Administration (FDA) for its anti-cancer properties. This orally bioavailable drug is classified as a non-selective histone deacetylase inhibitor (pan-HDACi) that inhibits class I, II, and IV HDACs at nanomolar levels due to its significant histone modifications and epigenetic mechanisms. A mismatch between histone acetyltransferases (HATs) and HDACs can negatively affect the regulation of the genes concerned, which in turn can contribute to tumorigenesis. Indeed, panobinostat inhibits HDACs, potentially leading to acetylated histone accumulation, re-establishing normal gene expression in cancer cells, and helping to drive multiple signaling pathways. These pathways include induction of histone acetylation and cytotoxicity for the majority of tested cancer cell lines, increased levels of p21 cell cycle proteins, enhanced amounts of pro-apoptotic factors (such as caspase-3/7 activity and cleaved poly (ADP-ribose) polymerase (PARP)) associated with decreased levels of anti-apoptotic factors [B-cell lymphoma 2 (Bcl-2) and B-cell lymphoma-extra-large (Bcl-XL)], as well as regulation of immune response [upregulated programmed death-ligand 1 (PD-L1) and interferon gamma receptor 1 (IFN-γR1) expression] and other events. The therapeutic outcome of panobinostat is therefore mediated by sub-pathways involving proteasome and/or aggresome degradation, endoplasmic reticulum, cell cycle arrest, promotion of extrinsic and intrinsic processes of apoptosis, tumor microenvironment remodeling, and angiogenesis inhibition. In this investigation, we aimed to pinpoint the precise molecular mechanism underlying panobinostat's HDAC inhibitory effect. A more thorough understanding of these mechanisms will greatly advance our knowledge of cancer cell aberrations and, as a result, provide an opportunity for the discovery of significant new therapeutic perspectives through cancer therapeutics.

Novel topoisomerase II/EGFR dual inhibitors: design, synthesis and docking studies of naphtho[2′,3′:4,5]thiazolo[3,2-a]pyrimidine hybrids as potential anticancer agents with apoptosis inducing activity

Topoisomerases II are ubiquitous enzymes with significant genotoxic effects in many critical DNA processes. Additionally, epidermal growth factor receptor (EGFR) plays pivotal role in tumour growth and angiogenesis. A novel series of naphtho[2',3':4,5]thiazolo[3,2- a ]pyrimidine hybrids have been designed, synthesised and evaluated for their topo IIα/EGFR inhibitory and apoptotic inducer activities. Cytotoxicity of the synthesised hybrids was evaluated against MCF-7, A549 and HCT-116 cell lines. Of the synthesised hybrids, 6i, 6a and 6c experienced superior cytotoxic activity compared to doxorubicin and erlotinib against the tested cancer cells. The molecular mechanism of these hybrids revealed their ability to successfully inhibit topo IIα and EGFR activities in micromolar concentration and may serve as topo II catalytic inhibitor. Moreover, these hybrids significantly arrested cell cycle at G2/M phase together with increased p53, caspae-7, caspase-9 levels and Bax/Bcl-2 ratio. The synthesised hybrids showed efficient binding pattern in molecular docking study and have acceptable drug likeness characters.

Discovery of 1H-indazole-6-amine Derivatives as Anticancer Agents: Simple But Effective

Background: Indazole is a promising structure present in various biological activity compounds; in particular, many 6-aminoindazole-containing compounds demonstrated anticancer activity. In our previous research, we discovered some of the 6-aminoindazole derivatives with excellent cytotoxicity in the human colorectal cancer cell line (HCT116). Objective: In this study, a series of 6-substituted amino-1H-indazole derivatives were designed and synthesized through simple and well-known chemical reactions, which were evaluated for anti-proliferative activity in four human cancer cell lines. Method: The title compounds were designed based on the structures of potential anticancer candidates in our previous report. The synthesis of 6-aminoindazole derivatives through acetylation and reductive amination with 6-amininoindazole as the starting material. Sulforhodamin B (SRB) assay was used for in vitro biological evaluation of synthesized compounds. Various physicochemical properties of them were predicted by online site Molinspiration. Results: Seven out of eight synthesized compounds showed growth inhibitory activity with IC50 values from 2.9 to 59.0 μM range in all four tested cancer cell lines. Of them, the compound N-(4-fluorobenzyl)- 1H-indazol-6-amine (9f) exhibited a potent anti-proliferative activity, with an IC50 value of 14.3±4.4 μM in the human colorectal cancer cell (HCT116) and non-cytotoxicity in the normal cell (lung fibroblast cells, MRC5, IC50 >100 μM). Conclusion: The bioactivity result and conformance of the physicochemical properties of the synthesized compounds to the "rule of three" for hit-like compounds suggested that 9f was effective and could be used as a hit for the development of novel anticancer agents.

Anti-cancer activity, and molecular docking of novel hybrid heterocyclic steroids revealed promising anti-hepatocellular carcinoma agent: Implication of cyclin dependent kinase-2 pathway.

To identify new steroidal agents with potential biological activities, we synthesized hybrid steroids containing thiazole, pyrazole, isoxazole, thiophene or phthalazine moiety. Epi-androsterone 1 reacted with phenylthiosemicarbazide to afford the corresponding androstane-4-phenyl-3-thiosemicarbazone derivative 2. The latter product was used in the synthesis of a series of annulated steroid derivatives. Also, Epi-androsterone 1 reacted with the thienopyridazine derivative 16 to afford the thieno[3,4-d]pyridazino-N-ylidenoandrostane derivative 17. Compound 17 reacted readily with electron-poor olefins to yield the corresponding phthalazine steroid derivatives. Detailed experimental and spectroscopic evidences for the structures of the newly synthesized compounds are explained. Compounds 3, 7, 8a, 12a, 14, 17 and 21a, were investigated individually as anticancer agents on different panel of human malignant cell lines. Moreover, a computer modelling investigation was performed to speculate the macromolecular targets for the most promising candidate. The results revealed a concentration-dependent reduction in the number of viable cells in all cancer cell lines. Most notably, compound 7 was the most effective compound against all tested cancer cell lines, especially against HepG2 cell line; therefore, the mode of action of this compound against HCC was investigated. Compound 7 was able to induce cell cycle arrest, and DNA fragmentation in HepG2 cells. Moreover, compound 7 induced apoptosis via upregulating the expression of caspase-3, -8, -9, P53, Bax and inhibiting the expression of BCL2, and CDK2 genes. Our results highlighted compound 7 as a promising anti-hepatocellular carcinoma agent, with theoretical, and practical potential binding affinity with CDK2; therefore, more investigations are required to elucidate its chemotherapeutic value as anti-HCC agent.

7-Geranyloxycinnamic Acid Isolated from Melicope lunu-ankenda Leaves Perturbs Colon Cancer and Breast Cancer Cell Lines' Growth via Induction of Apoptotic Pathway.

Globally, breast cancer is the most prevalent form of cancer in women and there is a need for alternative therapies such as plant-derived compounds with low systemic toxicity and selective toxicity to cancer cells. The aim of this study is to assess the cytotoxicity effects of 7-geranyloxycinnamic acid isolated from leaves of Melicope lunu-ankenda, a traditional medicinal plant, on the human breast cancer cell lines. Dried leaf powder was used for the preparation of different crude extracts using different solvents of increasing order of polarity. The structure of the isolated compound from the petroleum ether extract was elucidated by 1H and 13C NMR, LC-MS, and DIP-MS spectroscopy. The cytotoxic activity of the crude extract and 7-geranyloxycinnamic acid analyzed using MTT assay. Apoptotic analysis was evaluated using Annexin V-PI staining, AO/PI staining, intracellular ROS measurement, and measurement of activities of caspases 3/7, 8, and 9. Crude extracts and the isolated pure compound showed significant cytotoxicity against tested cancer cell lines. 7-geranyloxycinnamic acid was found to exert significant cytotoxic effects against breast cancer cell lines such as the MCF-7 and MDA-MB-231 cell lines. The cytotoxic effects are attributed to its ability to induce apoptosis via accumulation of ROS and activation of caspases in both breast cancer cell lines. The pure compound, 7-geranyloxycinnamic acid isolated from the leaves of M. lunu-ankenda, can exert significant cytotoxic effects against breast cancer cell lines without affecting the normal cells.

Synthesis, characterization, and biological activity of sterically hindered fluorine-containing platinum(II) complexes

Picoplatin is a sterically hindered antitumor compound with unique chemical and DNA binding properties, but its curative effect is not as good as other platinum drugs. Therefore, a series of sterically hindered fluorine-containing platinum(II) complexes Y1–Y5 were synthesized and characterized by introducing fluorine atoms at the 2-position of the pyridine group and different leaving group ligands. In vitro antiproliferative activity assessments of these complexes have been performed against A549, MCF-7, SW480, HCT116, HepG2 human cancer cell lines, and LO2 normal cells. The results showed that Y1 with 2-fluoropyridine coordination and chloride ion as the leaving group exhibited stronger antiproliferative activity and selectivity in the tested cancer cell lines, especially for A549 cancer cells. DNA binding constants were determined by UV absorption and fluorescence titration, and the binding ability of Y1 to DNA is much stronger than that of the others, which shows Y1 has better DNA damage ability. For the determination of apoptosis by the complex, the results were consistent with those of the MTT assay, and the apoptosis rate of Y1 reached 38.9%. Therefore, our work offers an advantageous method for the development of new platinum-based antitumor drugs.

Prenylated Flavonoids with Selective Toxicity against Human Cancers.

The antiproliferative activity of xanthohumol (1), a major prenylated chalcone naturally occurring in hops, and its aurone type derivative (Z)-6,4'-dihydroxy-4-methoxy-7-prenylaurone (2) were investigated. Both flavonoids, as well as cisplatin as a reference anticancer drug, were tested in vivo against ten human cancer cell lines (breast cancer (MCF-7, SK-BR-3, T47D), colon cancer (HT-29, LoVo, LoVo/Dx), prostate cancer (PC-3, Du145), lung cancer (A549) and leukemia (MV-4-11) and two normal cell lines (human lung microvascular endothelial (HLMEC)) and murine embryonic fibroblasts (BALB/3T3). Chalcone 1 and aurone 2 demonstrated potent to moderate anticancer activity against nine tested cancer cell lines (including drug-resistant ones). The antiproliferative activity of all the tested compounds against cancer and the normal cell lines was compared to determine their selectivity of action. Prenylated flavonoids, especially the semisynthetic derivative of xanthohumol (1), aurone 2, were found as selective antiproliferative agents in most of the used cancer cell lines, whereas the reference drug, cisplatin, acted non-selectively. Our findings suggest that the tested flavonoids can be considered strong potential candidates for further studies in the search for effective anticancer drugs.

Determination of Some Isoquinoline Alkaloids in Extracts Obtained from Selected Plants of the Ranunculaceae, Papaveraceae and Fumarioideae Families by Liquid Chromatography and In Vitro and In Vivo Investigations of Their Cytotoxic Activity.

Alkaloids are heterocyclic bases with widespread occurrence in nature. Plants are rich and easily accessible sources of them. Most isoquinoline alkaloids have cytotoxic activity for different types of cancer, including malignant melanoma, the most aggressive type of skin cancer. The morbidity of melanoma has increased worldwide every year. For that reason, developing new candidates for anti-melanoma drugs is highly needed. The aim of this study was to investigate the alkaloid compositions of plant extracts obtained from Macleaya cordata root, stem and leaves, Pseudofumaria lutea root and herb, Lamprocapnos spectabilis root and herb, Fumaria officinalis whole plant, Thalictrum foetidum root and herb, and Meconopsis cambrica root and herb by HPLC-DAD and LC-MS/MS. For determination of cytotoxic properties, human malignant melanoma cell line A375, human Caucasian malignant melanoma cell line G-361, and human malignant melanoma cell line SK-MEL-3 were exposed in vitro to the tested plant extracts. Based on the in vitro experiments, Lamprocapnos spectabilis herb extract was selected for further, in vivo research. The toxicity of the extract obtained from Lamprocapnos spectabilis herb was tested using an animal zebrafish model in the fish embryo toxicity test (FET) for determination of the LC50 value and non-toxic doses. Determination of the influence of the investigated extract on the number of cancer cells in a living organism was performed using a zebrafish xenograft model. Determination of the contents of selected alkaloids in different plant extracts was performed using high performance liquid chromatography (HPLC) in a reverse-phase system (RP) on a Polar RP column with a mobile phase containing acetonitrile, water and ionic liquid. The presence of these alkaloids in plant extracts was confirmed by LC-MS/MS. Preliminary cytotoxic activity of all prepared plant extracts and selected alkaloid standards was examined using human skin cancer cell lines A375, G-361, and SK-MEL-3. The cytotoxicity of the investigated extract was determined in vitro by cell viability assays (MTT). For in vivo determination of investigated extract cytotoxicity, a Danio rerio larvae xenograft model was used. All investigated plant extracts in in vitro experiments exhibited high cytotoxic activity against the tested cancer cell lines. The results obtained using the Danio rerio larvae xenograft model confirmed the anticancer activity of the extract obtained from Lamprocapnos spectabilis herb. The conducted research provides a basis for future investigations of these plant extracts for potential use in the treatment of malignant melanoma.

Design, synthesis, anticancer, and antibacterial evaluation of some quinazolinone-based derivatives as DHFR inhibitors.

Two series of quinazolinone derivatives were designed and synthesized as dihydrofolate reductase (DHFR) inhibitors. All compounds were evaluated for their antibacterial and antitumor activities. Antibacterial activity was evaluated against three strains of Gram-positive and Gram-negative bacteria. Compound 3d exhibited the highest inhibitory activity against Staphylococcus aureus DHFR (SaDHFR) with IC50 of 0.769 ± 0.04 μM compared to 0.255 ± 0.014 μM for trimethoprim. Compound 3e was also more potent than trimethoprim against Escherichia coli DHFR (EcDHFR) with IC50 of 0.158 ± 0.01 μM and 0.226 ± 0.014 μM, respectively. Compound 3e exhibited a promising antiproliferative effect against most of the tested cancer cells. It also showed potent activity against leukemia (CCRF-CEM, and RPMI-8226); lung NCI-H522, and CNS U251 with GI% of 65.2, 63.22, 73.28, and 97.22, respectively. The cytotoxic activity of compound 3e was almost half the activity of doxorubicin against CCRF-CEM cell line with IC50 of 1.569 ± 0.06 μM and 0.822 ± 0.03 µM, respectively. In addition, compound 3e inhibited human DHFR with IC50 value of 0.527 ± 0.028 µM in comparison to methotrexate (IC50 = 0.118 ± 0.006 µM). Compound 3e caused an arrest of the cell cycle mainly at the S phase and caused a rise in the overall apoptotic percentage from 2.03% to 48.51%. (23.89-fold). Treatment of CCRF-CEM cells with compound 3e produced a significant increase in the active caspase-3 level by 6.25-fold compared to untreated cells. Molecular modeling studies were performed to evaluate the binding pattern of the most active compounds in the bacterial and human DHFR.

Mannose Inhibits the Pentose Phosphate Pathway in Colorectal Cancer and Enhances Sensitivity to 5-Fluorouracil Therapy

Colorectal cancer (CRC) is one of the leading cancers and causes of death in patients. 5-fluorouracil (5-FU) is the therapy of choice for CRC, but it exhibits high toxicity and drug resistance. Tumorigenesis is characterized by a deregulated metabolism, which promotes cancer cell growth and survival. The pentose phosphate pathway (PPP) is required for the synthesis of ribonucleotides and the regulation of reactive oxygen species and is upregulated in CRC. Mannose was recently reported to halt tumor growth and impair the PPP. Mannose inhibitory effects on tumor growth are inversely related to the levels of phosphomannose isomerase (PMI). An in silico analysis showed low PMI levels in human CRC tissues. We, therefore, investigated the effect of mannose alone or in combination with 5-FU in human CRC cell lines with different p53 and 5-FU resistance statuses. Mannose resulted in a dose-dependent inhibition of cell growth and synergized with 5-FU treatment in all tested cancer cell lines. Mannose alone or in combination with 5-FU reduced the total dehydrogenase activity of key PPP enzymes, enhanced oxidative stress, and induced DNA damage in CRC cells. Importantly, single mannose or combination treatments with 5-FU were well tolerated and reduced tumor volumes in a mouse xenograft model. In summary, mannose alone or in combination with 5-FU may represent a novel therapeutic strategy in CRC.

Abstract 6312: Targeting HSPG is more efficient than using soluble heparin for interfering with cancer cell adhesion and migration

Abstract Given the relevance of heparan sulfate proteoglycans (HSPG) and of heparin binding ligands in the regulation of cancer cell adhesion, migration and invasiveness, drugs that may interfere with HSPG functions have been studied for decades for their possible use in oncology. In this view heparin analogues have been tested for their effectiveness not only in cancer associated thromboembolism but also for their potential effects on cancer growth and metastasis. Despite a number of promising preclinical results and clinical hypothesis, many different clinical trials on the use of heparin analogues in different cancers reported no significant benefits in tumor progression and overall survival, indicating that using heparin as a soluble decoy for the many HSPG ligands does not appear to be the best possible approach (1,2). Taking advantage of the tetra-branched peptide NT4, which is a specific ligand of HSPG sulfated glycosaminoglycan (GAG) chains (3, 4), we tested the effectiveness of targeting cell membrane HSPG, as a potential alternative approach for interfering with HSPG functions in cancer cell invasiveness. We had demonstrated that NT4 can either inhibit or increase oriented migration in respectively PANC-1 pancreas adenocarcinoma and TE671 rhabdomyosarcoma human cancer cells, indicating a crucial but diverse role of HSPGs in oriented cell migration in different cancer cells. NT4 and heparin were here compared for their effect on adhesion and migration of different cancer cell lines on extracellular matrix (ECM) supports, as well as for their effect on proliferation and in vitro invasiveness of the same cell lines. We found that inhibition of cancer cells adhesion to ECM proteins by NT4 varies in different cancer cell lines, ranging from 90% inhibition to as low as 20%, whereas heparin has no effect. Analogous results were obtained when testing cancer cell migration. NT4 inhibited migration and in vitro invasiveness of different human cancer cell lines, with the sole exception of TE671 cells. Heparin did not modify cell migration in any of the tested cancer cell lines. NT4 had no effect on cell proliferation, whereas, notably, heparin resulted to stimulate cell proliferation in some cancer cell lines. Differently from using heparin as an unspecific soluble decoy, targeting HSPG with well-defined ligands can result in an efficient tumor-specific treatment. Nevertheless, as for most cancer cellular targets, a detailed definition of cellular mechanisms and a related personalized approach is mandatory. 1) Schünemann HJ et al. Lancet Haematol. 2020;7(10):e746. 2) Posch F et al. ibid:e703. 3) Brunetti J et al. Sci. Rep. 2016,6,27174. 4) Depau L et al. J Med Chem 2020;63(24):15997. Citation Format: Lorenzo Depau, Elisabetta Mandarini, Jlenia Brunetti, Marta Zanchi, Chiara Falciani, Luisa Bracci. Targeting HSPG is more efficient than using soluble heparin for interfering with cancer cell adhesion and migration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6312.

Abstract 3341: Preliminary clinical validation of COLO eDX, a novel plasma circulating tumor DNA methylation assay for detecting colorectal cancer

Abstract Introduction: Liquid biopsies with cell-free DNA (cfDNA) from plasma, are rapidly emerging as an important and minimally invasive approach for the early diagnosis of cancer. Here we present COLO eDX, as a novel methylation-specific droplet digital PCR (ddPCR)-based colorectal cancer (CRC) diagnostic tool. Methods: During the discovery phase, we analyzed DNA methylome profiles of more than 10,000 samples from CRC, normal colorectal mucosa, and other cancer types (the TCGA and GEO public databases). Through the multi-step biomarker discovery studies, we selected 14 CRC-specific hypermethylated CpG sites. Among them, 2 candidate sites were uniquely methylated in a CRC cell line among tested cancer cell lines from various organs including CRC (HCT-15), 2 hepatocellular carcinomas (Huh7 and SK-HEP-1), 2 lung cancers (A549 and HCC827), bladder cancer (HT-1376), prostate cancer (LNCap clone FGC), and breast cancer (T-47D). Through the methylation-specific ddPCR assay of paired tissue samples from the patients with CRC, we identified that single candidate was specifically hypermethylated in the most tumor tissues regardless of cancer stage. Copy numbers of the target region (TC) were divided by the copy numbers of the internal control (IC) to normalize. Results: We prospectively collected post-operative plasma samples from patients with CRC (N=23) who underwent surgery at Seoul National University Boramae Medical Center. According to the AJCC 8th staging system and risk groups allocation, 3 patients had standard risk and 8 patients had high risk for recurrence, among stage II patients. Among stage III patients, 3 patients had low risk and 6 patients had high risk for recurrence. 3 patients were stage IV at initial presentation. IC value in the stage IV patients (R2 resected) were significantly higher than that other groups (R0 resected), while there was no significant difference among stage II and stage III patients. Recurrence free survival for patients TC or IC value equal or higher than the median and those lower than the median were not statistically different. Since 3 patients in high risk stage III and 4 patients in high risk stage II patients did not receive the recommended adjuvant chemotherapy, comparison of RFS should be interpreted with caution. Conclusion: We have developed and validated the qualitative detection of methylation status of the bisulfite-converted target region in cfDNA. Further studies are warranted to evaluate the impact of COLO eDx on recurrence in a larger cohort of CRC patients. Citation Format: Jin-Soo Kim, Yun Young Lee, Seung Yeon Jung, Mi Young Kim, Rumi Shin, Jin Hyun Park, Joon An, Jinil Han, Seung Chul Heo, Youngho Moon. Preliminary clinical validation of COLO eDX, a novel plasma circulating tumor DNA methylation assay for detecting colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3341.

Phenolic composition, antioxidant, cytotoxic activities and cardioprotective effect of hydroalcoholic extract from aerial-parts of Hypericum attenuatum Fisch. ex Choisy

This study investigated phenolic metabolites, antioxidant, cytotoxic and cardioprotective effects of the hydroalcoholic extract from the aerial parts of Hypericum attenuatum Fisch. ex Choisy. The total phenolic and flavonoid contents of the extract were 132.40 ± 2.06 mg GAE/g and 101.46 ± 1.47 mg QE/g respectively. The extract exhibited antioxidant activities with an EC50 value against DPPH radical of 0.099 ± 0.03 mg/mL and a FRAP value of 1.22 ± 0.086 mmol/L Fe2+. The extract could protect H9c2 cardiomyoblasts from the injury of H2O2, while it restored the H9c2 cell viability to 82.69 ± 2.33% at 100 μg/mL. The extract possessed cytotoxicity on MGC803, C666-1 and SW620 cells with IC50 values of 69.77 ± 2.43 μg/mL, 74.97 ± 1.08 μg/mL and 58.91 ± 1.81 μg/mL, respectively. Moreover, it could promote apoptosis of the tested cancer cells. This research provided useful information for the utilization of H. attenuatum as herbal medicine.

Inhibition Effect of Cell-Free Supernatants of Environmental Acanthamoeba Strains on the Viability of Metastatic Cell-Lines

It is known that some of the therapeutic agents against cancer cells are isolated from natural sources such as plants and animals. However, due to increasing drug resistance, studies on the discovery of new sources are needed. In this study, it was aimed to investigate the inhibition effects of four native Acanthamoeba strains on different cancer cell lines (MDA-MB-231, PC3, MAT-LyLu). 3T3 cells were used as normal cell line. All strains were recultured by using non-nutrient agar spread by heat-inactivated Escherichia coli ATCC 25922. A.castellanii ATCC 50373 was used as the standard strain. Molecular identification of the native Acanthamoeba isolates was done by polymerase chain reaction (PCR) and DNA sequence analysis using specific primer pairs (P-FLA-F, P-FLA-R, JDP-F, JDP-R). Axenic cultures of all strains were obtained in 25 cm2 tissue culture flasks and in peptone yeast extract glucose (PYG) medium. In order to investigate the effect of cell-free supernatants obtained from axenic cultures on cancer cell lines and 3T3 cell viability, MTT method was applied using different concentrations of cell-free supernatants (1%, 2%, 5%, 10%, 15%). It was determined that the viability of 3T3 cells was not affected by any Acanthamoeba cell-free supernatants (p≤ 0.05). All of the samples tested were found to have a significant inhibitory effect (p<0.05) on the viability of PC3 and MAT-LyLu cells (human and rat prostate cancer cell line). However, none of the samples had an inhibitory effect on the viability of MDA-MB-231 (breast cancer cell-line). Two native Acanthamoeba cell-free supernatants showed higher inhibitory potency (28% and 21.9%) at 2% concentration against PC3 cells compared to the reference strain (16%). Similarly, the same Acanthamoeba samples were also shown to have a better inhibition potential on the viability of MAT-LyLu cells than the reference strain. It was found that the inhibitory potential of Acanthamoeba cell-free supernatants may not be related to proteins and proteases. The results obtained from this study showed that Acanthamoeba species living in the aquatic environment isolated from our country have a potential inhibitory effect against the tested cancer cell lines. In addition to plants and animals, Acanthamoeba cell-free supernatants can also be a source for natural therapeutic substances that act against cancer cells. However, it is necessary to carry out new studies using more strains in order to detect strains with higher inhibitory effects.

Actinobacteria from Arctic and Atlantic deep-sea sediments-Biodiversity and bioactive potential.

The deep-sea covers over 70% of the Earth's surface and harbors predominantly uncharacterized bacterial communities. Actinobacteria are the major prokaryotic source of bioactive natural products that find their way into drug discovery programs, and the deep-sea is a promising source of biotechnologically relevant actinobacteria. Previous studies on actinobacteria in deep-sea sediments were either regionally restricted or did not combine a community characterization with the analysis of their bioactive potential. Here we characterized the actinobacterial communities of upper layers of deep-sea sediments from the Arctic and the Atlantic (Azores and Madeira) ocean basins, employing 16S rRNA metabarcoding, and studied the biosynthetic potential of cultivable actinobacteria retrieved from those samples. Metabarcoding analysis showed that the actinobacterial composition varied between the sampled regions, with higher abundance in the Arctic samples but higher diversity in the Atlantic ones. Twenty actinobacterial genera were detected using metabarcoding, as a culture-independent method, while culture-dependent methods only allowed the identification of nine genera. Isolation of actinobacteria resulted on the retrieval of 44 isolates, mainly associated with Brachybacterium, Microbacterium, and Brevibacterium genera. Some of these isolates were only identified on a specific sampled region. Chemical extracts of the actinobacterial isolates were subsequently screened for their antimicrobial, anticancer and anti-inflammatory activities. Extracts from two Streptomyces strains demonstrated activity against Candida albicans. Additionally, eight extracts (obtained from Brachybacterium, Brevibacterium, Microbacterium, Rhodococcus, and Streptomyces isolates) showed significant activity against at least one of the tested cancer cell lines (HepG2 and T-47D). Furthermore, 15 actinobacterial extracts showed anti-inflammatory potential in the RAW 264.4 cell model assay, with no concomitant cytotoxic response. Dereplication and molecular networking analysis of the bioactive actinobacterial extracts showed the presence of some metabolites associated with known natural products, but one of the analyzed clusters did not show any match with the natural products described as responsible for these bioactivities. Overall, we were able to recover taxonomically diverse actinobacteria with different bioactivities from the studied deep-sea samples. The conjugation of culture-dependent and -independent methods allows a better understanding of the actinobacterial diversity of deep-sea environments, which is important for the optimization of approaches to obtain novel chemically-rich isolates.

Anticancer activities and DNA/BSA interactions for five Cu(II) compounds with substituted terpyridine ligands

The reactions of copper chloride and 4′-(4′-substituted-phenyl)-2,2′:6′,2′′-terpyridine ligands bearing nitro (L1 ), methyl (L2 ), phenyl (L3 ), tolyl (L4 ) and carboxyl (L5 ) gave five copper compounds (1–5) which have mononuclear structures with their Cu(II) ions five-coordinate in the irregular square pyramidal geometries. Their in vitro biological properties were evaluated by CCK-8 assay, which show remarkable proliferation inhibitive properties on the cancer cell lines including A549, Eca-109, and Bel-7402, and two human normal cell lines (HL-7702 and HLF). The values of IC50 indicate better anticancer activities of these compounds than that of cisplatin. Compound 2 with methyl group exhibits the best anticancer activity, with IC50 value < 2.5 µM on all the tested cancer cells. The interactions of the compounds with DNA were evaluated using UV–Vis absorption titrations, circular dichroism spectroscopy, and DNA viscosity. The complexes show significant insertion binding with DNA sequences and strong intercalator binding mode with DNA sequences. Compounds 1–5 interact with the functional groups of DNA through π···π stacking, van der Waals interactions, hydrophobic interactions, and hydrogen bonds as determined by molecular docking. In addition, strong binding interactions between the five copper(II) complexes and bovine serum albumin (BSA) were studied by fluorescence titration and circular dichroism spectroscopy.

Synthesis, antitumor evaluation and computational study of thiazolidinone derivatives of dehydroabietic acid-based B ring-fused-thiazole.

In an attempt to search for new natural product-based antitumor agents, a series of novel thiazolidinone derivatives of dehydroabietic acid-based B ring-fused-thiazole were designed and synthesized. The primary antitumor tests showed that compounds 5m exhibited almost the best inhibitory activity against the tested cancer cells. The computational study suggested NOTCH1, IGF1R, TLR4, and KDR were the core targets of the title compounds, and the IC50 of SCC9 and Cal27 is strong correlation with the binding ability of TLR4 and compounds.

Conversion of antibacterial quinolone drug levofloxacin to potent cytotoxic agents.

Levofloxacin, the optical S-(-) isomer of ofloxacin, is a broad-spectrum antibacterial agent widely used to control various infections caused by Gram-positive and Gram-negative bacteria. While the COOH group is necessary for antibacterial activity, its modification can offer anticancer activity to the fluoroquinolone framework. Therefore, several levofloxacin carboxamides 11a-j and 12 containing 5-substituted-1,3,4-thiadiazole residue were synthesized and screened in vitro for their anticancer activity. The in vitro MTT viability assay revealed that the most compounds had significant activity against cancer cells MCF-7, A549, and SKOV3. In particular, the 3-chloro- and 4-fluoro- benzyl derivatives (11b and 11h), with IC50 values of 1.69-4.76 μM were as potent as or better than doxorubicin. It should be noted that the mother quinolone levofloxacin showed no activity on the tested cancer cell lines. The SAR analysis demonstrated that the 3-chloro or 4-fluoro substituent on the S-benzyl moiety had positive effect on the activity. Further in vitro evaluations of the most promising compounds 11b and 11h by flow cytometric analysis and comet test revealed the ability of compounds in the induction of apoptosis and blockage of the cell proliferation at the G1-phase by nuclear fragmentation and DNA degradation in cancer cells. The obtained results demonstrated that the alteration of 6-COOH functional group in the levofloxacin structure and conjugation with a proper heterocyclic pharmacophore is a good strategy to obtain new anticancer agents.

Chemical and Biological Studies of Endophytes Isolated from Marchantia polymorpha.

Natural bioresources, predominantly plants, have always been regarded as the richest source of drugs for diseases threatening humanity. Additionally, microorganism-originating metabolites have been extensively explored as weapons against bacterial, fungal, and viral infections. However, the biological potential of metabolites produced by plant endophytes still remains understudied, despite significant efforts reflected in recently published papers. Thus, our goal was to evaluate the metabolites produced by endophytes isolated from Marchantia polymorpha and to study their biological properties, namely anticancer and antiviral potential. The cytotoxicity and anticancer potential were assessed using the microculture tetrazolium technique (MTT) against non-cancerous VERO cells and cancer cells-namely the HeLa, RKO, and FaDu cell lines. The antiviral potential was tested against the human herpesvirus type-1 replicating in VERO cells by observing the influence of the extract on the virus-infected cells and measuring the viral infectious titer and viral load. The most characteristic metabolites identified in the ethyl acetate extract and fractions obtained by use of centrifugal partition chromatography (CPC) were volatile cyclic dipeptides, cyclo(l-phenylalanyl-l-prolyl), cyclo(l-leucyl-l-prolyl), and their stereoisomers. In addition to the diketopiperazine derivatives, this liverwort endophyte also produced arylethylamides and fatty acids amides. The presence of N-phenethylacetamide and oleic acid amide was confirmed. The endophyte extract and isolated fractions showed a potential selective anticancer influence on all tested cancer cell lines. Moreover, the extract and the first separated fraction noticeably diminished the formation of the HHV-1-induced cytopathic effect and reduced the virus infectious titer by 0.61-1.16 log and the viral load by 0.93-1.03 log. Endophytic organisms produced metabolites with potential anticancer and antiviral activity; thus, future studies should aim to isolate pure compounds and evaluate their biological activities.