Melatonin and Metformin Failed to Modify the Effect of Dacarbazine in Melanoma.

LESSONS LEARNED: Melatonin did not increase the efficacy of systemic chemotherapy in melanoma. Metformin did not increase the efficacy of systemic chemotherapy in melanoma. BACKGROUND: Current data support the possibility of antitumor activity of melatonin and metformin. METHODS: From March 2014 to December 2016, 57 patients with disseminated melanoma received dacarbazine (DTIC) 1,000 mg/m2 on day 1 of a 28-day cycle, either as monotherapy (first group) or in combination with melatonin 3 mg p.o. daily (second group) or metformin 850 mg two times a day p.o. daily (third group) as the first-line of chemotherapy. The primary endpoint was objective response rate (ORR). Secondary endpoints were time to progression (TTP), overall survival (OS), immunologic biomarkers, and quality of life. RESULTS: ORR was 7% and did not differ among the treatment groups. Median TTP was 57, 57, and 47 days, respectively, in the first, second, and third groups (р = .362). Median OS was 236, 422, and 419 days, respectively (p = .712). Two patients from the combinations groups showed delayed response to therapy. The increase of CD3+ CD4+ HLA-DR+ lymphocytes (p = .003), CD3+ CD8+ HLA-DR+ (p = .045), CD3+ CD8+ lymphocytes (p = .012), CD4+ CD25high CD127low lymphocytes (p = .029), and overall quantity of lymphocytes (p = .021) was observed in patients with clinical benefit. CONCLUSION: No benefit was found in either combination over DTIC monotherapy. Delayed responses in melatonin and metformin combination groups were registered. The increase of lymphocyte subpopulations responsible for antitumor immune response demonstrates the immune system's potential involvement in clinical activity.

Metformin Restores the Drug Sensitivity of MCF-7 Cells Resistant Derivates via the Cooperative Modulation of Growth and Apoptotic-Related Pathways.

The phenomenon of the primary or acquired resistance of cancer cells to antitumor drugs is among the key problems of oncology. For breast cancer, the phenomenon of the resistance to hormonal or target therapy may be based on the numerous mechanisms including the loss or mutation of estrogen receptor, alterations of antiapoptotic pathways, overexpression of growth-related signaling proteins, etc. The perspective approaches for overcoming the resistance may be based on the usage of compounds such as inhibitors of the cell energetic metabolism. Among the latter, the antidiabetic drug metformin exerts antitumor activity via the activation of AMPK and the subsequent inhibition of mTOR signaling. The experiments were performed on the ERα-positive MCF-7 breast cancer cells, the MCF-7 sublines resistant to tamoxifen (MCF-7/T) and rapamycin (MCF-7/Rap), and on triple-negative MDA-MB-231 breast cancer cells. We have demonstrated metformin's ability to enhance the cytostatic activity of the tamoxifen and rapamycin on both parent MCF-7 cells and MCF-7-resistant derivates mediated via the suppression of mTOR signaling and growth-related transcriptional factors. The cooperative effect of metformin and tested drugs was realized in an estrogen-independent manner, and, in the case of tamoxifen, was associated with the activation of apoptotic cell death. Similarly, the stimulation of apoptosis under metformin/tamoxifen co-treatment was shown to occur in the MCF-7 cells after steroid depletion as well as in the ERα-negative MDA-MB-231 cells. We conclude that metformin co-treatment may be used for the increase and partial restoration of the cancer cell sensitivity to hormonal and target drugs. Moreover, the combination of metformin with tamoxifen induces the apoptotic death in the ERα-negative breast cancer cells opening the additional perspectives in the treatment of estrogen-independent breast tumors.

Upregulation of Akt/Raptor signaling is associated with rapamycin resistance of breast cancer cells.

mTOR inhibitors are considered today to be one of the most promising anticancer drugs. Here to study the mechanism of the acquired resistance of MCF-7 breast cancer cells to mTOR inhibitors two different models of the cell resistance were used: rapamycin-resistant MCF-7/Rap subline developed under long-term rapamycin treatment, and metformin-resistant MCF-7/M subline obtained by long-term metformin treatment. We have found that both resistant sublines were characterized by common features: increased expression of mTOR-interacting Raptor protein, increased phosphorylation of Akt, and activation of growth-related transcriptional factor AP-1. Cell response to mTOR inhibitors was partially restored under treatment with PI3K inhibitor wortmannin supporting the direct connection between Akt activation and poor cell response to therapeutic drugs. Transfection of mir-181c, one of the positive regulators of Akt and mTOR, led to an increase in the cell resistance to both mTOR inhibitors, rapamycin and metformin, which correlated with Raptor overexpression and activation of Akt/AP-1 signaling. In general, the effect of Raptor overexpression in the resistant cells, as well as the ability of mir-181c to modulate the Raptor expression, can open novel perspectives in the treatment of rapalogues-resistant cancers, based on the drugs design targeting mir-181c/Raptor axis.

Dissimilar associations of same metabolic parameters with main chronic noncommunicable diseases (cancer vs some other NCDs).

Hormone-dependent tissues' cancers (mainly breast and endometrial and several others) are among the most frequent malignancies in adults and are often discussed in context of their correlation with other chronic noncommunicable diseases (NCDs), for example, cardiovascular and cerebrovascular conditions, and their risk factors, which may also be hormone metabolic. An idea that is often expressed delineates common factors leading to NCDs of malignant and nonmalignant nature. However, this idea is not always confirmed by study results. The reasons for this discrepancy are not clear and require further analysis. This editorial tries to show the importance of this problem with a few examples (in particular, by attracting information on the role of birthweight, adult height and family history of diabetes) which may help us understand some mechanisms behind interconnections of major NCDs, including cancer.

Endocrinology of obese and nonobese endometrial cancer patients: is there role of tumor molecular-biological type?

Aim: To compare endocrine characteristics of  endometrial cancer (EC) patients based on recent molecular EC types classification. Materials & methods: A total of 234 treatment-naive EC patients as well their tumors were studied. Results: Patients with POLE mutations demonstrated tendency to lower body mass index (BMI) and higher serum estradiol. Patients with p53 overexpression were older and had higher diabetes incidence. In the without characteristic molecular profile group there was no difference in fasting serum insulin, estradiol and testosterone levels between women with BMI ≥30.0 and <30.0. The mismatch repair deficient group patients had a tendency toward later menarche compared with the without characteristic molecular profile group one. Conclusion: Studied endocrine characteristics are associated with BMI or tumor molecular-biological type that might be relevant to EC genesis, course and prognosis.

Endometrial cancer evolution: new molecular-biologic types and hormonal-metabolic shifts.

The question hidden in the title of this manuscript (whether the topic develops or remains constant) is important for all areas of science. It is also a serious problem for endometrial cancer (EC) study. In recent times the incidence of EC gradually increases in parallel with obesity epidemics. The main point of this review was evaluation of changes in EC area in last few decades, which are not only seen in tumor incidence, but also in its biology, hormonal-metabolic characteristics of patients and in the ratio of risk and anti-risk factors. One can hope that data accumulated recently and summarized here under the notion of EC evolution will find its use for advancement of EC prevention and treatment.

Features of omental adipose tissue in endometrial cancer patients with 'standard' or 'metabolically healthy' obesity: associations with tumor process characteristics.

PURPOSE: Adipose tissue products may contribute to endometrial cancer (EC) initiation and further growth that encourages the analysis of this issue in patients with different obesity phenotypes. METHODS/PATIENTS: Omental fat depot characteristics were studied in EC patients (n = 57) with "standard" (SO) or "metabolically healthy" (MHO) obesity. Collected omental samples were evaluated by immunohistochemistry /IHC/ for brown fat marker UCP1, CYP19 (aromatase) and macrophage infiltration markers (CD68, CD163, crown-like structures/CLS) expression. Total RNA extracted from the same samples was investigated for UCP1, CYP19, PTEN and adipokine omentin mRNA. RESULTS: Immunohistochemistry data revealed a statistically significant increase in aromatase and CD68 expression and tendency to increase of UCP1 expression in SO patients' omental fat compared to samples obtained from MHO patients. Additionally, positive correlation of EC clinical stage with UCP1 protein and its mRNA content in omental fat was pronounced in MHO as well as SO group, while with omentin mRNA it was discovered only in patients with SO. An inclination to the correlation with better tumor differentiation was seen for UCP1 and CD68 protein expression in patients with MHO and with worse (high grade) differentiation-for CD68 expression in the group with SO. CONCLUSIONS: In aggregate, this suggests that obesity phenotype has significant impact on omental fat tissue characteristics which is related to the clinical course of EC and may have practical consequences.

Plasma exosomal miR-21 and miR-181a differentiates follicular from papillary thyroid cancer.

Thyroid cancer (TC) is the most common endocrine malignancy and its incidence has increased over the last few decades. As has been revealed by a number of studies, TC tissue's micro-RNA (miRNA) profile may reflect histological features and the clinical behavior of tumor. However, alteration of the miRNA profile of plasma exosomes associated with TC development has to date not been explored. We isolated exosomes from plasma and assayed their characteristics using laser diffraction particle size analysis, atomic force microscopy, and western blotting. Next, we profiled cancer-associated miRNAs in plasma exosomes obtained from papillary TC patients, before and after surgical removal of the tumor. The diagnostic value of selected miRNAs was evaluated in a large cohort of patients displaying different statuses of thyroid nodule disease. MiRNA assessment was performed by RT-qPCR. In total, 60 patients with different types of thyroid nodal pathology were included in the study. Our results revealed that the development of papillary TC is associated with specific changes in exosomal miRNA profiles; this phenomenon can be used for differential diagnostics. MiRNA-31 was found to be over-represented in the plasma exosomes of patients with papillary TC vs. benign tumors, while miRNA-21 helped to distinguish between benign tumors and follicular TC. MiRNA-21 and MiRNA-181a-5p were found to be expressed reciprocally in the exosomes of patients with papillary and follicular TC, and their comparative assessment may help to distinguish between these types of TC with 100 % sensitivity and 77 % specificity.

The phenomenon of acquired resistance to metformin in breast cancer cells: The interaction of growth pathways and estrogen receptor signaling.

Metformin, a biguanide antidiabetic drug, is used to decrease hyperglycemia in patients with type 2 diabetes. Recently, the epidemiological studies revealed the potential of metformin as an anti-tumor drug for several types of cancer, including breast cancer. Anti-tumor metformin action was found to be mediated, at least in part, via activation of adenosine monophosphate-activated protein kinase (AMPK)-intracellular energy sensor, which inhibits the mammalian target of rapamycin (mTOR) and some other signaling pathways. Nevertheless, some patients can be non-sensitive or resistant to metformin action. Here we analyzed the mechanism of the formation of metformin-resistant phenotype in breast cancer cells and its role in estrogen receptor (ER) regulation. The experiments were performed on the ER-positive MCF-7 breast cancer cells and metformin-resistant MCF-7 subline (MCF-7/M) developed due to long-term metformin treatment. The transcriptional activity of NF-κB and ER was measured by the luciferase reporter gene analysis. The protein expression was determined by immunoblotting (Snail1, (phospho)AMPK, (phospho)IκBα, (phospho)mTOR, cyclin D1, (phospho)Akt and ERα) and immunohistochemical analysis (E-cadherin). We have found that: 1) metformin treatment of MCF-7 cells is accompanied with the stimulation of AMPK and inhibition of growth-related proteins including IκBα, NF-κB, cyclin D1 and ERα; 2) long-term metformin treatment lead to the appearance and progression of cross-resistance to metformin and tamoxifen; the resistant cells are characterized with the unaffected AMPK activity, but the irreversible ER suppression and constitutive activation of Akt/Snail1 signaling; 3) Akt/Snail1 signaling is involved into progression of metformin resistance. The results presented may be considered as the first evidence of the progression of cross-resistance to metformin and tamoxifen in breast cancer cells. Importantly, the acquired resistance to both drugs is based on the constitutive activation of Akt/Snail1/E-cadherin signaling that opens new perspectives to overcome the metformin/tamoxifen resistance of breast cancer.

Insulinemia, heterogeneity of obesity and the risk of different types of endometrial cancer: existing evidence.

Due to a number of reasons, endometrial cancer is a point of interest not only for oncologists, but also for a variety of specialists - especially endocrinologists. The endocrinology of endometrial cancer can be firmly divided into two categories - steroid and non-steroid. The steroid approach dominated during several decades due to hyperestrogenization signs observed in some patients. The balance was only regained in the last 15 years, when the role of diabetes and insulin resistance began to draw attention. This review aims to provide an update on connections between insulinemia (insulin resistance) and different obesity phenotypes as well to discuss their relation to development of endometrial cancer, its clinical-morphological features and the increasing number of its molecular-biological subtypes.

Lectin-induced agglutination method of urinary exosomes isolation followed by mi-RNA analysis: Application for prostate cancer diagnostic.

BACKGROUND: Prostate cancer is the most common cancer in men. Prostate-specific antigen has, however, insufficient diagnostic specificity. Novel complementary diagnostic approaches are greatly needed. MiRNAs are small regulatory RNAs which play an important role in tumorogenesis and are being investigated as a cancer biomarker. In addition to their intracellular regulatory functions, miRNAs are secreted into the extracellular space and can be found in various body fluids, including urine. The stability of extracellular miRNAs is defined by association with proteins, lipoprotein particles, and membrane vesicles. Among the known forms of miRNA packaging, tumour-derived exosome-enclosed miRNAs is thought to reflect the vital activity of cancer cells. The assessment of the exosomal fraction of urinary miRNA may present a new and highly specific method for prostate cancer diagnostics; however, this is challenged by the absence of reliable and inexpensive methods for isolation of exosomes. METHODS: Prostate cancer (PC) cell lines and urine samples collected from 35 PC patients and 35 healthy donors were used in the study. Lectins, phytohemagglutinin, and concanavalin A were used to induce agglutination of exosomes. The efficiency of isolation process was evaluated by AFM and DLS assays. The protein content of isolated exosomes was analysed by western blotting. Exosomal RNA was assayed by automated electrophoresis and expression level of selected miRNAs was evaluated by RT-qPCR. The diagnostic potency of the urinary exosomal miRNA assessment was estimated by the ROC method. RESULTS: The formation of multi-vesicular agglutinates in urine can be induced by incubation with lectin at a final concentration of 2 mg/ml. These agglutinates contain urinary exosomes and may be pelleted by centrifugation with a relatively low G-force. The analysis of PC-related miRNA in urinary exosomes revealed significant up-regulation of miR-574-3p, miR-141-5p, and miR-21-5p associated with PC. CONCLUSIONS: Lectin-induced aggregation is a low-cost and easily performed method for isolation of exosomes from urine. Isolated exosomes can be further analysed in terms of miRNA content. The miRNA profile of urinary exosomes reflects development of prostate cancer and may present a promising diagnostic tool.

Potential and real 'antineoplastic' and metabolic effect of metformin in diabetic and nondiabetic postmenopausal females.

AIM: The goal of this study was to determine if the single nucleotide polymorphisms marking potential sensitivity to metformin (MF) correlate with hormone-metabolic status as well as with actual response to MF in postmenopausal cancer patients with or without Type 2 diabetes mellitus and in diabetics without cancer. PATIENTS & METHODS: The carriage of ten different SNPs was evaluated in all patients by PCR, and hormone-metabolic status was estimated by anthropometry, ELISA and enzyme colorimetric assays. The response to daily 1-1.7 g of MF was studied based on hormone-metabolic parameters and indirect end points (endometrium thickness, mammographic breast density). RESULTS & CONCLUSION: The changes in evaluated 'antineoplastic' and metabolic response marker values were seen in 33.3 and 61.8% of the cases, respectively. Several genetic markers were found that showed an inclination to less frequent 'antineoplastic' or more frequent metabolic response to MF which may be helpful in further studies of this drug in cancer patients.

Features of endometrial cancer in patients with 'metabolically healthy' versus 'standard' obesity: the decreasing frequency of metabolically healthy obesity.

BACKGROUND: As endometrial cancer (EC) prevalence increases with obesity, we aimed to determine whether EC characteristics depend upon obesity type: 'standard' (SO) or 'metabolically healthy obesity' (MHO). PATIENTS & METHODS: 258 EC patients were included. Data on anthropometry, blood hormones, lipids and glucose, and tumor features were collected. RESULTS: EC clinicopathologic characteristics and clinical stage correlate differently with BMI and obesity type. BMI is related inversely with tumor grade while SO patients are characterized by a more advanced clinical stage than those with MHO. Besides typical insulin resistance signs, EC patients with SO often display a higher serum leptin/adiponectin ratio compared with MHO patients. Historical data suggest a gradual increase in EC patient height and weight, and a decrease in MHO prevalence. CONCLUSION: It is currently unknown whether the latter observation reflects the evolution of EC, or obesity alongside the current epidemic. Regardless, the reduced MHO prevalence demonstrates the need for more intensive preventive measures aimed at obesity and obesity-associated conditions, including different EC subtypes.

Autoantibodies to estrogen receptors and their involvement in autoimmune diseases and cancer.

The involvement of estrogens, which influence many physiologic processes, has been shown in the development or progression of several diseases including some cancers, most notably breast cancer, and autoimmune disorders. Estrogenic signal is transferred via estrogen receptors (ER) which have dual localization, predominantly intracellular but also in plasma membrane. The discovery of membrane-associated ER (mER) has greatly expanded our understanding of estrogen action; upon ligand binding, mER rapidly activate different signaling pathways inducing downstream transcription factors. Some target genes of the mER pathway may be activated independently of the intracellular ER. Additionally, intracellular ER action can be modulated by mER-initiated signaling. Most notably, the identification of autoantibodies reacting with ER (ERAB) and their possible pathogenic role in autoimmunity and cancer have opened a new path for the research in the estrogen-related receptor activity. In this review, we briefly recapitulate the localization and function of ER and mostly discuss the possible role of ERAB as novel potential prognostic and/or predictive tools in autoimmunity and cancer.

Genetic polymorphisms potentially associated with response to metformin in postmenopausal diabetics suffering and not suffering with cancer.

Metformin is a well-known antidiabetic medication, which, besides diabetes, may be involved into modulation of other age-related pathologies, including cancer. The study concerns 12 gene polymorphisms divided into 2 groups consisting of 6 genes each. The first group was composed from so-called "standard" (S) polymorphisms, for which the connection with metabolic response to metformin is already established. The second group included polymorphisms of genes encoding proteins possibly connected with diabetes mellitus type 2 (DM2), impaired glucose tolerance or cancer and entitled here as "associated" (A). A total of 156 postmenopausal women (average age 60.7 ± 0.7) were included, 37 of them healthy, 64 with type DM2 and concurrent treatment-naïve cancer (mostly breast, endometrial or colorectal cancer), 32 with DM2 without cancer, and 23 with treatment-naïve cancer and normal glucose tolerance. The leading metformin response S-marker in combined group of DM2 patients was the CC variant of OCT1-R61C polymorphism of organic cation transporter protein 1 gene. In cancer patients without DM2, this position belonged to AC and AA genotypes of OCT1_rs622342 polymorphism. Among the A-polymorphisms, GA variant of sex hormone-binding globulin gene SHBG_D356N was less frequently observed in DM2 patients with or without cancer. Besides, in diabetics, the same polymorphic variant of SHBG as well as GC genotype of oxidized lipoprotein receptor OLR1_G501C and GG genotype of locus rs11065987 near BRAP gene were carried rather often in combination with "metformin-positive" variant of OCT1_R61C. In addition, carriers of OCT1_R61C and OCT1_rs622342 polymorphisms with potentially positive reaction to metformin had higher insulin resistance score (HOMA-IR) values. Received data lead to the conclusion that postmenopausal diabetics, both with and without cancer, differ in genetic stigmata of potential response to metformin less than they differ from cancer patients without DM2. As genetic polymorphisms associated with metabolic and anticancer metformin (and, possibly, phenformin) effects may be different, this subject requires further investigation.