LEP (-2548G>A LEP) and LEPR (223Gln>Arg, 109Lys>Arg) polymorphisms as breast cancer risk factors in the Polish female population.

On a global scale, breast cancer is the most common type of cancer in women, and it is still a growing problem. Therefore, new prognostic or diagnostic markers are required that would facilitate the assessment of patients or provide more efficient therapy, respectively. In these studies, we analyzed the contribution of LEP (2548G>A) and LEPR (109 Lys>Arg and 223Gln>Arg) genes polymorphisms to the risk of breast cancer development. The study involved 209 women aged 59.6 ± 11 years diagnosed with breast cancer and 202 healthy women aged 57.8 ± 8.2 years, who were blood donors. Polymorphism were evaluated by PCR-RFLP reaction followed by the verification of part of the samples by sequencing. The results of the study confirmed obesity as a significant breast cancer development risk factor in Polish women. However, no significant association between the studied polymorphisms and breast cancer risk or severity of the neoplastic disease was found. Interestingly, it was shown that wild type 223Gln>Gln leptin receptor (LEPR) was statistically more common in women with human epidermal growth factor receptor 2 negative (HER2-) than human epidermal groth factor receptor 2 positive (HER2+) breast cancer and wild type form of 2548G>A LEP was more common in women with progesterone receptor positive (PR+) than progesterone receptor negative (PR-) breast cancer. Studied polymorphisms of the LEP and LEPR genes do not increase breast cancer risk in the population of Polish women. However, they can affect PR an HER receptors expression and thus the severity of the disease. Noteworthy, this interesting correlation is being reported for the first time and might constitute an essential contribution to the identification of molecular mechanisms of carcinogenesis.

Effect of 3-O-acetylaleuritolic acid from in vitro-cultured Drosera spatulata on cancer cells survival and migration.

BACKGROUND: Drosera spatulata is a source of many compounds such as naphthoquinones, phenolic acids, flavonoids, anthocyanins, and naphthalene derivatives. Unfortunately, the information regarding the biological activity and chemical profile of those compounds is still incomplete. Herein, we investigated the biological activity of 3-O-acetylaleuritolic acid (3-O-AAA) in cancer cell lines. METHODS: The cell viability of HeLa, HT-29, MCF7, and MCF12A cells was assessed using MTT assay. Proliferation potential was assessed using the clonogenic assay and flow cytometry. Migration modulation was tested using a scratch assay. Protein expression was analyzed by immunoblotting. RESULTS: 3-O-AAA significantly inhibited the growth of all tested tumor cells. The results of the colony formation assay suggested cytostatic properties of the studied compound. The scratch assay showed that 3-O-AAA was an efficient migration inhibitor in a dose-dependent manner. Moreover, it caused modulation of mTOR, beclin1, and Atg5 proteins suggesting a possible role of the compound in autophagy induction. CONCLUSION: Collectively, these results demonstrated that 3-O-AAA inhibited the proliferation and migration of cancer cell lines as well as contributed to autophagy induction showing some anticancer properties.

Telomerase Inhibitor TMPyP4 Alters Adhesion and Migration of Breast-Cancer Cells MCF7 and MDA-MB-231.

Human telomeres were one of the first discovered and characterized sequences forming quadruplex structures. Association of these structures with oncogenic and tumor suppressor proteins suggests their important role in cancer development and therapy efficacy. Since cationic porphyrin TMPyP4 is known as G-quadruplex stabilizer and telomerase inhibitor, the aim of the study was to analyze the anticancer properties of this compound in two different human breast-cancer MCF7 and MDA-MB-231 cell lines. The cytotoxicity of TMPyP4 alone or in combination with doxorubicin was measured by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromid) and clonogenic assays, and the cell-cycle alterations were analyzed by flow cytometry. Telomerase expression and activity were evaluated using qPCR and telomeric repeat amplification protocol (TRAP) assays, respectively. The contribution of G-quadruplex inhibitor to protein pathways engaged in cell survival, DNA repair, adhesion, and migration was performed using immunodetection. Scratch assay and functional assessment of migration and cell adhesion were also performed. Consequently, it was revealed that in the short term, TMPyP4 neither revealed cytotoxic effect nor sensitized MCF7 and MDA-MB-231 to doxorubicin, but altered breast-cancer cell adhesion and migration. It suggests that TMPyP4 might substantially contribute to a significant decrease in cancer cell dissemination and, consequently, cancer cell survival reduction. Importantly, this effect might not be associated with telomeres or telomerase.

The non-canonical functions of telomerase: to turn off or not to turn off.

Telomerase is perceived as an immortality enzyme that enables passing the Hayflick limit. Its main function is telomere restoration but only in a limited group of cells, including cancer cells. Since it is found in a vast majority of cancer cells, it became a natural target for cancer therapy. However, it has much more functions than just altering the metabolism of telomeres-it also reveals numerous so-called non-canonical functions. Thus, a question arises whether it is always beneficial to turn it off when planning a cancer strategy and considering potential side effects? The purpose of this review is to discuss some of the recent discoveries about telomere-independent functions of telomerase in the context of cancer therapy and potential side effects.

Comparison of bioactive compounds content in leaf extracts of Passiflora incarnata, P. caerulea and P. alata and in vitro cytotoxic potential on leukemia cell lines

ABSTRACT Passiflora caerulea L., P. alata Curtis and P. incarnata L. (synonym for P. edulis Sims), are the most popular representatives of the Passiflora genus in South America. In recent years, a growing attention is paid to the biological activity and phytochemical profiles of crude extracts from various species of Passiflora in worldwide. The aim of this study was to evaluate and to compare of anti-leukemic activity of the dry crude extracts from leaves of three Passiflora species from greenhouse of Poland in two human acute lymphoblastic leukemia cell lines: CCRF-CEM and its multidrug resistant variant. Two systems of liquid chromatography in order to assessment of phytochemical composition of extracts were applied. Extracts of P. alata and P. incarnata showed the potent inhibitory activity against human acute lymphoblastic leukemia CCRF-CEM, while P. caerulea not showed activity (or activity was poor). Despite similarities in quality phytochemical profile of extracts from P. caerulea and P. incarnata, differences in quantity of chemical compounds may determine their various pharmacological potency. For the activity of P. alata extract the highest content of terpenoids and a lack of flavones C-glycosides are believed to be crucial. Summarizing, the crude extract from P. alata leaves may be considered as a substance for complementary therapy for cancer patients.

Cytotoxic activity of stigmasteryl esters and products of their thermo-oxidative degradation against drug sensitive and drug resistant human acute lymphoblastic leukemia cells.

BACKGROUND: Phytosterols are mainly known as a cholesterol-lowering factor, although they form oxidation products during food storage and processing. Moreover, phytosterol oxidation products (POP) can be ab- sorbed and found in human serum, so there is the need to investigate their impact on different kinds of cells. METHODS: Esters of fatty acids (oleic, linoleic and linolenic) with stigmasterol were synthetized and heated at 180°C, for 1–12 hours. The cytotoxic effect on the leukemic cells of unheated stigmasteryl esters and the mixture of compounds after heating was determined using MTT assays. POP were identified using GC-MS. The total number of POP was analysed by SPE fractionation and GC-FID separation. Dimers, trimers and oligomers in non-polar fraction were determined by gel permeation chromatography with refractive index detection. RESULTS: After heating, stigmasterol oxidation products were formed (up to 1.1 mg/g ester). The heating increased the potency of the compounds to reduce cell population and form POPs and oligomers in a time-dependent manner. CONCLUSIONS: The cytotoxicity depends on the kind of ester, dose and time. The strongest cytotoxic effect was found after 72 hours of cell treatment. Among the three stigmasteryl esters tested the most cytotoxic effect was caused by stigmasteryl linoleate.

Telomerase and drug resistance in cancer.

It is well known that a decreased expression or inhibited activity of telomerase in cancer cells is accompanied by an increased sensitivity to some drugs (e.g., doxorubicin, cisplatin, or 5-fluorouracil). However, the mechanism of the resistance resulting from telomerase alteration remains elusive. There are theories claiming that it might be associated with telomere shortening, genome instability, hTERT translocation, mitochondria functioning modulation, or even alterations in ABC family gene expression. However, association of those mechanisms, i.e., drug resistance and telomerase alterations, is not fully understood yet. We review the current theories on the aspect of the role of telomerase in cancer cells resistance to therapy. We believe that revealing/unravelling this correlation might significantly contribute to an increased efficiency of cancer cells elimination, especially the most difficult ones, i.e., drug resistant.

Semisynthetic oleanane triterpenoids inhibit migration and invasion of human breast cancer cells through downregulated expression of the ITGB1/PTK2/PXN pathway.

This paper reports a study on the role of two synthetic derivatives of oleanolic acid (OA), HIMOXOL and Br-HIMOLID, in the regulation of cell migration and invasion and the underlying molecular mechanisms of breast cancer cells. The effect of the compounds on four breast cancer cell lines (MCF7, MDA-MB-231, MDA-MB-468, and T-47D) and also on noncancerous breast cells, MCF-12A, was reported. The compounds had no effect on the migration of MCF-12A cells. However, both the derivatives revealed a higher cytotoxicity than the maternal compound OA, and in sub-cytotoxic concentrations, they decreased the migration of MCF7, MDA-MB-231, and MDA-MB-468 breast cancer cells and also the invasion of MCF7 and MDA-MB-231 cells; although, the derivatives had no effect on the migration and invasion of T-47D cells. Both the derivatives of OA inhibited the cell migratory and invasive abilities of breast cancer cells by downregulating the expressions of ITGB1, PTK2, and PXN genes and by decreasing the phosphorylation status and the level of its respective proteins (integrin ß1, FAK, and paxillin, respectively). This study is the first to report the antimigratory and anti-invasive activities of HIMOXOL and Br-HIMOLID in breast cancer cells.

Nanoscale size effect in in situ titanium based composites with cell viability and cytocompatibility studies.

Novel in situ Metal Matrix Nanocomposite (MMNC) materials based on titanium and boron, revealed their new properties in the nanoscale range. In situ nanocomposites, obtained through mechanical alloying and traditional powder metallurgy compaction and sintering, show obvious differences to their microstructural analogue. A unique microstructure connected with good mechanical properties reliant on the processing conditions favour the nanoscale range of results of the Ti-TiB in situ MMNC example. The data summarised in this work, support and extend the knowledge boundaries of the nanoscale size effect that influence not only the mechanical properties but also the studies on the cell viability and cytocompatibility. Prepared in the same bulk, in situ MMNC, based on titanium and boron, could be considered as a possible candidate for dental implants and other medical applications. The observed relations and research conclusions are transferable to the in situ MMNC material group. Aside from all the discussed relations, the increasing share of these composites in the ever-growing material markets, heavily depends on the attractiveness and a possible wider application of these composites as well as their operational simplicity presented in this work.

In vitro biocompatibility of titanium after plasma surface alloying with boron.

Recently, the effect of different sizes of precursor powders during surface plasma alloying modification on the properties of titanium surface was studied. In this work we show in vitro test results of the titanium (α-Ti) after plasma surface alloying with boron (B). Ti-B nanopowders with 2 and 10wt% B were deposited onto microcrystalline Ti substrate. The in vitro cytocompatibility of these biomaterials was evaluated and compared with a conventional microcrystalline Ti. During the studies, established cell line of human gingival fibroblasts and osteoblasts were cultured in the presence of tested materials, and its survival rate and proliferation activity were examined. For this purpose, MTT assay, flow cytometric and fluorescent microscopic evaluation were made. Biocompatibility tests carried out indicate that the Ti after plasma surface alloying with B could be a possible candidate for dental implants and other medicinal applications. Plasma alloying is a promising method for improving the properties of titanium, thus increasing the field of its applications.

Methyl 3-hydroxyimino-11-oxoolean-12-en-28-oate (HIMOXOL), a synthetic oleanolic acid derivative, induces both apoptosis and autophagy in MDA-MB-231 breast cancer cells.

HIMOXOL (methyl 3-hydroxyimino-11-oxoolean-12-en-28-oate) is a synthetic derivative of oleanolic acid (OA). HIMOXOL revealed the highest cytotoxic effect among tested synthetic OA analogs. In this study we focused on elucidating the cytotoxic mechanism of HIMOXOL in MDA-MB-231 breast cancer cells. HIMOXOL reduced MDA-MB-231 cell viability with an IC50 value of 21.08±0.24µM. In contrast to OA, the tested compound induced cell death by activating apoptosis and the autophagy pathways. More specifically, we found that HIMOXOL was able to activate the extrinsic apoptotic pathway, which was proven by observation of caspase-8, caspase-3 and PARP-1 protein activation in Western blot analysis. An increase in the ratio of Bax/Bcl-2 protein levels was also detected. Moreover, HIMOXOL triggered microtubule-associated protein LC3-II expression and upregulated beclin 1. This observed compound activity was modulated by mitogen-activated protein kinases and NFκB/p53 signaling pathways. Together, these data suggest that HIMOXOL, a synthetic oleanolic acid derivative which activates dual cell death machineries, could be a potential and novel chemotherapeutic agent.

Telomerase as a useful target in cancer fighting-the breast cancer case.

Telomerase was initially considered as a relevant factor distinguishing cancer from normal cells. During detailed studies, it appeared that its expression and activity is not only limited to cancer cells however, but in this particular cells, the telomerase is much more abundant. Thus, it has become a very promising target for an anticancer therapy. It was revealed in many studies that regulation of telomerase is a multifactorial process in mammalian cells, involving regulation of expression of telomerase subunits coding genes, post-translational protein-protein interactions, and protein phosphorylation. Numerous proto-oncogenes and tumor suppressor genes are engaged in this mechanism, and the complexity of telomerase control is studied in the context of tumor development as well as aging. Additionally, since numerous studies reveal a correlation between short telomeres and increased genome instability or cell mortality, the telomerase control appears to be one of the crucial factors to study in order to improve the cancer diagnostics and therapy or prevention. Interestingly, almost 100 % of adenocarcinoma, including breast cancer cells, expresses telomerase which makes it a good target for telomerase-related therapy. Additionally, telomerase is also supposed to be associated with drug resistance. Thus, targeting the enzyme might result in attenuation of this phenomenon. Moreover, since stem cells existence was reported, it must be considered whether targeting telomerase can bring some serious side effects and result in stem cells viability or their regenerative potential decrease. Thus, we review some molecular mechanisms engaged in therapy based on targeting telomerase in breast cancer cells.