New nanodrug design for cancer therapy: Its synthesis, formulation, in vitro and in silico evaluations.

The aim of this study was to develop a novel nanosize drug candidate for cancer therapy. For this purpose, (S)-methyl 2-[(7-hydroxy-2-oxo-4-phenyl-2H-chromen-8-yl)methyleneamino]-3-(1H-indol-3-yl)propanoate (ND3) was synthesized by the condensation reaction of 8-formyl-7-hydroxy-4-phenylcoumarin with l-tryptophan methyl ester. Its controlled release formulation was prepared and characterized by different spectroscopic and imaging methods. The cytotoxic effects of ND3 and its controlled release formulation were evaluated against MCF-7 and A549 cancer cell lines, and it was found that both of them have a toxic effect on cancer cells. For drug design and process development, the molecular docking analysis technique helps to clarify the effects of some DNA-targeted anticancer drugs to determine the interaction mechanisms of these drugs on DNA in a shorter time and at a lower cost. By using the molecular docking analysis and DNA binding assays, the interaction between the synthesized compound and DNA was elucidated and non-binding interactions were also determined. To predict the pharmacokinetics, and thereby accelerate drug discovery, the absorption, distribution, metabolism, excretion and toxicity values of the synthesized compound were determined by in silico methods.

Screening for antitumor activity of various plant extracts on HeLa and C 4-I cell lines.

PURPOSE: Cancer is a long process that leads the organism to death and is associated with the normal cells acquiring the ability to divide permanently. Nowadays, the use of natural products in cancer therapy has a great importance. In addition, working with plants that are endemic to Turkey and determining the biological activities of these plant extracts, is extremely important due to the potential for new drug development. There is no comparative study available in the literature on the antitumor effects of Colchicum sanguicolle, a new found species of the genus Colchicum in Turkey, Crateagus microphylla, of the genus Crateagus and Centaurea antiochia of the genus Centaurea. In this study, we tried to demonstrate the antitumor effect of these plant extracts on HeLa and C 4-1 cells. METHODS: Five different doses (0.001, 0.01, 0.05, 0.25 and 0.5 mg/ml) of the three plant types were prepared and applied for 24, 48 and 72 hrs on the cervical cancer derived cell lines. Subsequently, the growth rate was evaluated with the mitochondrial dehydrogenase enzyme method. RESULTS: Colchicum sanguicolle extracts showed the most effective antitumor activity. For the Colchicum sanguicolle extract, the IC50 dose for HeLa cells was 0.01 mg/ml at 48 hrs, while for the C-4 I cells it was 0.001 mg/ml at 48 hrs. These results showed that C-4 I cells were more sensitive to the Colchicum sanguicolle extracts. Conclus?on: The results of from this study regarding the antitumor effect of plant extracts of endemic varieties of Turkey may have an important place in design and development of anticancer drugs and would make contributions to other studies to be conducted in this area.

Targeting Periostin Expression Makes Pancreatic Cancer Spheroids More Vulnerable to Natural Killer Cells.

Pancreatic cancer (PaCa) characteristically has a dense tumor microenvironment, which results in poor patient prognosis. Pancreatic stellate cells (PSCs) are the most abundant cells in the PaCa microenvironment and the principal source of collagen. Periostin, a matricellular protein, is produced specifically by PSCs and promotes the aggressiveness of PaCa cells by facilitating extracellular collagen assembly. Here, we aimed to decrease extracellular collagen assembly by suppressing periostin, thereby increasing the cytotoxic activity of natural killer (NK) cells. Periostin expression was suppressed in PSCs (called PSC-P) using CRISPR-Cas9. PaCa cells (BxPC-3) were co-cultured with PSC and PSC-P cells in a 3D environment to form tumor spheroids mimicking the tumor microenvironment. The extracellular collagen production of spheroids was evaluated by Masson's trichrome staining. The cytotoxic activity of NK-92 cells was analyzed by flow cytometry and confocal microscopy via CD107a staining. Cell death in BxPC-3 cells was evaluated by measuring Annexin-V and PI positivity using flow cytometry. As a result, periostin suppression decreased extracellular collagen and increased the infiltration of NK-92 cells into spheroids, and induced cell death in PaCa cells. In conclusion, we suggest that periostin might be a therapeutic target for PaCa and further analysis is warranted using in vivo models for proof-of-concept.

Structural, spectroscopic, in silico, in vitro and DNA binding evaluations of tyrosyl-lysyl-threonine.

The main objective of the present study is to investigate the molecular structure and DNA binding interaction of the tyrosyl-lysyl-threonine (YKT) tripeptide, which has anticancer, antioxidant and analgesic properties, using various in silico (MD, QM, molecular docking), spectroscopic (UV, FT-IR, FTIR-ATR, Raman, gel electrophoresis) and in vitro (MCF-7 and HeLa cancer cell lines and BEAS-2B cell line) methods. The optimized geometry, vibrational wavenumbers, molecular electrostatic potential (MEP), natural bond orbital (NBO) and HOMO-LUMO (highest occupied molecular orbital- lowest unoccupied molecular orbital) calculations were carried out with Density Functional Theory (DFT) using B3LYP/6-311++G(d,p) basis set to indicate conformational, vibrational and intramolecular charge transfer characteristics. The assignment of all fundamental theoretical vibration wavenumbers was performed using potential energy distribution analysis (PED). DNA is a significant pharmacological target of drugs in several diseases such as cancer. For this reason, molecular docking calculation was used to elucidate the binding and interaction between YKT tripeptide and DNA at the atomic level. Also, the dynamic behaviors of YKT and DNA was examined using MD simulations. Besides, the interaction of YKT with DNA was experimentally examined by UV titration method and agarose gel electrophoresis method. Experimental results showed that YKT was intercalatively and electrostatically bound to CT-DNA (Calf thymus DNA) and cleavage pBR322 DNA in the presence of H2O2. The pharmacokinetic profile of YKT was also obtained. Cytotoxic effect of YKT was evaluated on MCF-7, HeLa and BEAS-2B cell lines. Hence, these studies about YKT tripeptide may pave the way for the development of various cancer drugs. Communicated by Ramaswamy H. Sarma.

Effects of hypobaric conditions on apoptosis signalling pathways in HeLa cells.

Nowadays increasing effectiveness in cancer therapy and investigation of formation of new strategies that enhance antiproliferative activity against target organs has become a subject of interest. Although the molecular mechanisms of apoptosis can not be fully explained, it is known that cell suicide program existing in their memory genetically is activated by pathophysiological conditions and events such as oxidative stress. Low pressure (hypobaric) conditions that create hypoxia promote apoptosis by inhibiting cell cycling. In this study, determination of the effects of fractional hypobaric applications at different times on HeLa cells at cellular and molecular levels were targeted. Experiments were carried out under hypobaric conditions (35.2 kPa) in a specially designed hypobaric cabin including 2% O2 and 98% N. Application of fractional hypobaric conditions was repeated two times for 3 hours with an interval of 24 hours. At the end of the implementation period cells were allowed to incubate for 24 hours for activation of repair mechanisms. Cell kinetic parameters such as growth rate (MTT) and apoptotic index were used in determination of the effect of hypobaric conditions on HeLa cells. Also in our study expression levels of the Bcl-2 gene family that have regulatory roles in apoptosis were determined by the RT-PCR technique to evaluate molecular mechanisms. The results showed that antiproliferative effect of hypobaric conditions on HeLa cells started three hours from the time of application and increased depending on the period of exposure. While there was a significant decrease in growth rate values, there was a significant increase in apoptotic index values (p<0.01). Also molecular studies showed that hypobaric conditions caused a significant increase in expression level of proapoptotic gene Bax and significant decrease in antiapoptotic Bfl-1. Consequently fractional application of hypobaric conditions on HeLa cell cultures increased both antiproliferative and apoptotic effects and these effects were triggered by the Bax gene.