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Renal αKlotho along with fibroblast growth factor 23 regulates phosphate and vitamin D metabolism. Its cleavage yields soluble Klotho controlling intracellular processes. αKlotho has anti‐inflammatory and antioxidant effects and is nephro‐ and cardioprotective. AMP‐dependent kinase (AMPK) is a nephro‐ and cardioprotective energy sensor. Given that both αKlotho and AMPK have beneficial effects in similar organs, we studied whether AMPK regulates αKlotho gene expression in Madin–Darby canine kidney, normal rat kidney 52E, and human kidney 2 cells. Using quantitative real‐time PCR and western blotting, we measured αKlotho expression upon pharmacological manipulation or siRNA‐mediated knockdown of AMPKα. AMPK activator 5‐aminoimidazole‐4‐carboxamide ribonucleoside (AICAR) enhanced αKlotho expression, an effect reduced in the presence of AMPK inhibitor compound C or siRNA targeting AMPK catalytic α‐subunits (α1 and α2). Similarly, AMPK activators metformin and phenformin upregulated αKlotho transcripts. Taken together, our results suggest that AMPK is a powerful inducer of αKlotho and could thereby contribute to the development of future therapeutic interventions. [ABSTRACT FROM AUTHOR]

N-methyl-D-aspartate (NMDA) receptors are inhibited by many medicinal drugs. The recent successful repurposing of NMDA receptor antagonists ketamine and dextromethorphan for the treatment of major depressive disorder further enhanced the interest in this field. In this work, we performed a screening for the activity against native NMDA receptors of rat CA1 hippocampal pyramidal neurons among biguanide compounds using the whole-cell patch-clamp method. Antimalarial biguanides proguanil and cycloguanil, as well as hypoglycemic biguanide phenformin, inhibited them in micromolar concentrations, while another hypoglycemic biguanide metformin and antiviral biguanide moroxydine were practically ineffective. IC50 values at −80 mV holding voltage were 3.4 ± 0.6 µM for cycloguanil, 9.0 ± 2.2 µM for proguanil and 13 ± 1 µM for phenformin. The inhibition by all three compounds was not competitive. Cycloguanil acted as an NMDA receptor voltage-dependent trapping channel blocker, while proguanil and phenformin acted as allosteric inhibitors. Our results support the potential clinical repurposing of biguanide compounds for the treatment of neurodegenerative disorders linked to glutamatergic excitotoxicity while also providing a better understanding of structural determinants of NMDA receptor antagonism by biguanides. [ABSTRACT FROM AUTHOR]

Renal αKlotho along with fibroblast growth factor 23 regulates phosphate and vitamin D metabolism. Its cleavage yields soluble Klotho controlling intracellular processes. αKlotho has anti‐inflammatory and antioxidant effects and is nephro‐ and cardioprotective. AMP‐dependent kinase (AMPK) is a nephro‐ and cardioprotective energy sensor. Given that both αKlotho and AMPK have beneficial effects in similar organs, we studied whether AMPK regulates αKlotho gene expression in Madin–Darby canine kidney, normal rat kidney 52E, and human kidney 2 cells. Using quantitative real‐time PCR and western blotting, we measured αKlotho expression upon pharmacological manipulation or siRNA‐mediated knockdown of AMPKα. AMPK activator 5‐aminoimidazole‐4‐carboxamide ribonucleoside (AICAR) enhanced αKlotho expression, an effect reduced in the presence of AMPK inhibitor compound C or siRNA targeting AMPK catalytic α‐subunits (α1 and α2). Similarly, AMPK activators metformin and phenformin upregulated αKlotho transcripts. Taken together, our results suggest that AMPK is a powerful inducer of αKlotho and could thereby contribute to the development of future therapeutic interventions.

Altered glycolytic metabolism has been associated with chemoresistance in acute myeloid leukemia (AML). However, there are still aspects that need clarification, as well as how to explore these metabolic alterations in therapy. In the present study, we aimed to elucidate the role of glucose metabolism in the acquired resistance of AML cells to cytarabine (Ara-C) and to explore it as a therapeutic target. Resistance was induced by stepwise exposure of AML cells to increasing concentrations of Ara-C. Ara-C-resistant cells were characterized for their growth capacity, genetic alterations, metabolic profile, and sensitivity to different metabolic inhibitors. Ara-C-resistant AML cell lines, KG-1 Ara-R, and MOLM13 Ara-R presented different metabolic profiles. KG-1 Ara-R cells exhibited a more pronounced glycolytic phenotype than parental cells, with a weaker acute response to 3-bromopyruvate (3-BP) but higher sensitivity after 48 h. KG-1 Ara-R cells also display increased respiration rates and are more sensitive to phenformin than parental cells. On the other hand, MOLM13 Ara-R cells display a glucose metabolism profile similar to parental cells, as well as sensitivity to glycolytic inhibitors. These results indicate that acquired resistance to Ara-C in AML may involve metabolic adaptations, which can be explored therapeutically in the AML patient setting who developed resistance to therapy. [ABSTRACT FROM AUTHOR]

Experimental and clinical studies have revealed the influence of AMP-activated protein kinase (AMPK) signaling on the development of non-alcoholic liver fibrosis. Currently, the results of experimental studies demonstrate that inhibition of AMPK promotes fibrogenesis, while its activation prevents the development of liver fibrosis. The purpose of this work is to establish the effect of activation of AMP-activated protein kinase by the administration of phenformin on the content of glycosaminoglycans, oxyproline and sialic acids in the liver of rats under the conditions of long-term administration of ethanol. The study was conducted on 24 male Wistar rats. The animals were randomly divided into 4 groups of 6 animals each, on which we modeled ethanol-induced liver damage and administered phenformin hydrochloride at a dose of 10 mg/kg. The experiment lasted 63 days. In the liver of rats, the content of total glycosaminoglycans, the concentration of heparin-heparan, keratan-dermatan and chondroitin fractions of glycosaminoglycans, the content of free oxyproline and sialic acids were studied. Long-term alcoholization leads to a violation of the extracellular matrix of the liver of rats, which is evidenced by a decrease in the concentration of proteoglycans and a redistribution of their fractions in the direction of a decrease in anti-inflammatory and regenerative fractions. Chronic intake of alcohol increases the processes of desialylation of glycoconjugates and the intensity of collagenolysis. Activation of AMP-activated protein kinase by administration of phenformin under the conditions of simulating ethanol-induced liver damage leads to an increase in the concentration of glycosaminoglycans due to the growth of heparin-heparan and chondroitin fractions and reduces the intensity of desialylation of glycoconjugates and collagenolysis in the liver of rats. [ABSTRACT FROM AUTHOR]

Lung cancer is one of the most fatal cancers worldwide. Resistance to conventional therapies remains a hindrance to patient treatment. Therefore, the development of more effective anti‐cancer therapeutic strategies is imperative. Solid tumors exhibit a hyperglycolytic phenotype, leading to enhanced lactate production; and, consequently, its extrusion to the tumor microenvironment. Previous data reveals that inhibition of CD147, the chaperone of lactate transporters (MCTs), decreases lactate export in lung cancer cells and sensitizes them to phenformin, leading to a drastic decrease in cell growth. In this study, the development of anti‐CD147 targeted liposomes (LUVs) carrying phenformin is envisioned, and their efficacy is evaluated to eliminate lung cancer cells. Herein, the therapeutic effect of free phenformin and anti‐CD147 antibody, as well as the efficacy of anti‐CD147 LUVs carrying phenformin on A549, H292, and PC‐9 cell growth, metabolism, and invasion, are evaluated. Data reveals that phenformin decreases 2D and 3D‐cancer cell growth and that the anti‐CD147 antibody reduces cell invasion. Importantly, anti‐CD147 LUVs carrying phenformin are internalized by cancer cells and impaired lung cancer cell growth in vitro and in vivo. Overall, these results provide evidence for the effectiveness of anti‐CD147 LUVs carrying phenformin in compromising lung cancer cell aggressiveness. [ABSTRACT FROM AUTHOR]

Introduction and aim. Hydrogen sulfide (H2S) has attracted the attention of researchers as a novel signaling molecule that affects vascular metabolism, immune function, stress and inflammation. It plays an important role in pathophysiological disorders under the conditions of the development of obesity, diabetes, non-alcoholic fatty liver disease and cardiovascular diseases. The purpose of this work is to establish correlation ratios of H2S concentration with markers of oxidative-nitrosative stress and extracellular matrix metabolism of the liver during chronic alcoholic hepatitis modeling and AMPK modulation by phenformin and doxorubicin. Material and methods. The experiments were performed on 36 white, sexually mature male Wistar rats, weighing 180-220 g. Alcoholic hepatitis was modelled by alcohol administration, on the background of alcoholic hepatitis animals received phenformin orally at a dose of 10 mg/kg or doxorubicin at a dose of 1.25 mg/kg intraperitoneally. Statistical processing of the results of biochemical studies was carried out using the non-parametric method of Spearman to determine correlations. Results. H2S during alcoholic hepatitis inversely proportionally strongly correlates with the concentration of nitrites, oxyproline and arginase activity. Phenformin administration during alcoholic hepatitis leads to formation of inversely proportionally strongly correlation of H2S with the production of superoxide anion radical, the concentration of malondialdehyde, activities of constitutive NO-synthases, nitrite reductases, nitrate reductases, and arginase. Doxorubicin administration during alcoholic hepatitis leads to formation of directly proportional strongly correlation of H2S with the activity of constitutive NO-synthases, nitrite reductases, nitrate reductases. Conclusion. Administration of phenformin or doxorubicin expands correlations between H2S and indicators of oxidative-nitrosative stress. [ABSTRACT FROM AUTHOR]

Ampk is an energy gatekeeper that responds to decreases in ATP by inhibiting energy-consuming anabolic processes and promoting energy-generating catabolic processes. Recently, we showed that Lkb1, an understudied kinase in B lymphocytes and a major upstream kinase for Ampk, had critical and unexpected roles in activating naïve B cells and in germinal center formation. Therefore, we examined whether Lkb1 activities during B cell activation depend on Ampk and report surprising Ampk activation with in vitro B cell stimulation in the absence of energy stress, coupled to rapid biomass accumulation. Despite Ampk activation and a controlling role for Lkb1 in B cell activation, Ampk knockout did not significantly affect B cell activation, differentiation, nutrient dynamics, gene expression, or humoral immune responses. Instead, Ampk loss specifically repressed the transcriptional expression of IgD and its regulator, Zfp318. Results also reveal that early activation of Ampk by phenformin treatment impairs germinal center formation but does not significantly alter antibody responses. Combined, the data show an unexpectedly specific role for Ampk in the regulation of IgD expression during B cell activation.

Aim: Breast cancer is the most common malignancy in women. This disease is a critical public health problem and further research at the molecular level would define its prognosis and specific treatment. Phenformin is an antidiabetic agent. Recent studies showed that it should be considered as a potential agent for the prevention and treatment of cancer cell lines. This study aimed to investigate the antiproliferative effect of phenformin on MDA-MB-231 and MCF-7 breast cancer cell lines. Materials and Methods: MDA-MB-231 and MCF-7 breast cancer cell lines were used in the study. Experimental groups; control group, 1 μM, 2.5 μM, and 5 μM phenformin administered groups were planned. WST-1 analysis was performed to evaluate the viability of cells 24 h after the treatments. Statistical analysis was carried out using the SPSS 17.0 statistical program. p<0.05 was considered to indicate a statistically significant difference. Results: In the MDA-MB-231 breast cancer cell line, there were statistically significant differences among all groups (p<0.05, for all), except between the 1 μM phenformin-treated and control groups (p>0.05). In the MCF-7 breast cancer cell line; there were statistically significant differences between the control group and 5 μM phenformin-treated, and between 1 μM phenformin-treated and 5 μM phenformin-treated (p<0.05, for all) groups. Conclusion: Phenformin seems to exert antiproliferative effects on MDA-MB-231 and MCF-7 breast cancer cell lines. It was observed that the antiproliferative effect was stronger in the MDA-MB-231 cell line. It showed a stronger antiproliferative effect in the MDAMB-231 cell line of antiproliferative effect. [ABSTRACT FROM AUTHOR]

Phenformin, a biguanide compound, has attracted increased attention due to its prominent antitumor activity. As a multi‑target agent, the antitumor effects of phenformin involve a wide range of factors, including inhibition of mitochondrial complex I, activation of AMP‑activated protein kinase, impact on the tumor microenvironment, suppression of cancer stem cells and others. In addition, phenformin has been shown to markedly augment the effectiveness of various clinical treatment methods, including radiotherapy, chemotherapy, targeted therapy and immunotherapy. It is noteworthy that breakthrough progress has been made in the treatment of cancer with phenformin with application in clinical trials for the treatment of melanoma. Phenformin not only reduces the lesion area of patients, but also enhances the efficacy of dalafinib/trimetinib. In the present review, the novel breakthroughs in the antitumor effects and mechanisms of phenformin were discussed. In addition, the current review focuses on the clinical development value of phenformin, striving to provide new insights into the future research direction of phenformin in the field of tumor treatment.

The treatment of many skin inflammation diseases, such as psoriasis and atopic dermatitis, is still a challenge and inflammation plays important roles in multiple stages of skin tumor development, including initiation, promotion and metastasis. Phenformin, a biguanide drug, has been shown to play a more efficient anti-tumor function than another well-known biguanide drug, metformin, which has been reported to control the expression of pro-inflammatory cytokines; however, little is known about the effects of phenformin on skin inflammation. This study used a mouse acute inflammation model, ex vivo skin organ cultures and in vitro human primary keratinocyte cultures to demonstrate that phenformin can suppress acute skin inflammatory responses induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in vivo and significantly suppresses the pro-inflammatory cytokines IL-1β, IL-6 and IL-8 in human primary keratinocytes in vitro. The suppression of pro-inflammatory cytokine expression by phenformin was not directly through regulation of the MAPK or NF-κB pathways, but by controlling the expression of c-Myc in human keratinocytes. We demonstrated that the overexpression of c-Myc can induce pro-inflammatory cytokine expression and counteract the suppressive effect of phenformin on cytokine expression in keratinocytes. In contrast, the down-regulation of c-Myc produces effects similar to phenformin, both in cytokine expression by keratinocytes in vitro and in skin inflammation in vivo. Finally, we showed that phenformin, as an AMPK activator, down-regulates the expression of c-Myc through regulation of the AMPK/mTOR pathways. In summary, phenformin inhibits the expression of pro-inflammatory cytokines in keratinocytes through the down-regulation of c-Myc expression to play an anti-inflammation function in the skin. [ABSTRACT FROM AUTHOR]

Summary: Background. Myeloproliferative neoplasms (MPN) are disorders characterized by an alteration at the hematopoietic stem cell (HSC) level, where the JAK2 mutation is the most common genetic alteration found in classic MPN (polycythemia vera, essential thrombocythemia, and primary myelofibrosis). We and others previously demonstrated that metformin reduced splenomegaly and platelets counts in peripheral blood in JAK2V617F pre-clinical MPN models, which highlighted the antineoplastic potential of biguanides for MPN treatment. Phenformin is a biguanide that has been used to treat diabetes, but was withdrawn due to its potential to cause lactic acidosis in patients. Aims. We herein aimed to investigate the effects of phenformin in MPN disease burden and stem cell function in Jak2V617F-knockin MPN mice. Results. In vitro phenformin treatment reduced cell viability and increased apoptosis in SET2 JAK2V67F cells. Long-term treatment with 40 mg/kg phenformin in Jak2V617F knockin mice increased the frequency of LSK, myeloid progenitors (MP), and multipotent progenitors (MPP) in the bone marrow. Phenformin treatment did not affect peripheral blood counts, spleen weight, megakaryocyte count, erythroid precursors frequency, or ex vivo clonogenic capacity. Ex vivo treatment of bone marrow cells from Jak2V617F knockin mice with phenformin did not affect hematologic parameters or engraftment in recipient mice. Conclusions. Phenformin increased the percentages of LSK, MP, and MPP populations, but did not reduce disease burden in Jak2V617F-knockin mice. Additional studies are necessary to further understand the effects of phenformin on early hematopoietic progenitors. [ABSTRACT FROM AUTHOR]

Psoriasis is a chronic inflammatory skin disease characterized by a complex pathogenesis involving various types of cells and cytokines. Among those, the pro-inflammatory cytokine IL-23/IL-17A axis plays a crucial role in the development and rapid progression of psoriasis. Phenformin, a derivative of metformin and a member of the biguanide class of drugs, exhibits superior anti-inflammatory and anti-tumor efficacy compared to metformin. However, the potential role of phenformin in anti-psoriatic skin inflammation has not been explored. In this study, we utilized a mouse model of psoriasis and an in vitro model using human keratinocytes to investigate whether phenformin can suppress psoriasis-like inflammatory responses. Our results demonstrate that the topical application of phenformin significantly inhibited acute skin inflammatory responses in the psoriasis mouse model induced by imiquimod (IMQ). Additionally, phenformin suppressed the expression of psoriasis-related cytokines IL-17, IL-23, IL-8, and S100A8/S100A9 in an in vitro psoriatic keratinocyte model induced by IMQ. Furthermore, we found that IMQ-induced psoriatic skin and IMQ-treated keratinocytes exhibited high expression of the c-Myc gene, which was downregulated by phenformin. The c-Myc inhibitor JQ1 similarly inhibited the psoriatic inflammatory response and the expression of psoriasis-related cytokines in both in vitro and in vivo models. phenformin ameliorates the psoriasis-like inflammatory response by inhibiting c-Myc expression in keratinocytes, suggesting its potential as a topical drug for the treatment of psoriasis. • Topical application of phenformin significantly suppresses skin inflammatory responses in the in vivo psoriasis mouse model. • Phenformin inhibited the expression of psoriasis-related cytokines IL-17, IL-23, IL-8, and S100A8/S100A9 in an in vitro psoriatic cell model. • The c-Myc inhibitor JQ1 similarly inhibited the psoriatic inflammatory response and the expression of psoriasis-related cytokines. • Phenformin, inhibiting c-Myc to suppress the psoriasis-like inflammatory response, can be used as a topical drug for psoriasis treatment. [ABSTRACT FROM AUTHOR]

The Liver Kinase B1 (LKB1) tumor suppressor acts as a metabolic energy sensor to regulate AMP-activated protein kinase (AMPK) signaling and is commonly mutated in various cancers, including non-small cell lung cancer (NSCLC). Tumor cells deficient in LKB1 may be uniquely sensitized to metabolic stresses, which may offer a therapeutic window in oncology. To address this question we have explored how functional LKB1 impacts the metabolism of NSCLC cells using 13C metabolic flux analysis. Isogenic NSCLC cells expressing functional LKB1 exhibited higher flux through oxidative mitochondrial pathways compared to those deficient in LKB1. Re-expression of LKB1 also increased the capacity of cells to oxidize major mitochondrial substrates, including pyruvate, fatty acids, and glutamine. Furthermore, LKB1 expression promoted an adaptive response to energy stress induced by anchorage-independent growth. Finally, this diminished adaptability sensitized LKB1-deficient cells to combinatorial inhibition of mitochondrial complex I and glutaminase. Together, our data implicate LKB1 as a major regulator of adaptive metabolic reprogramming and suggest synergistic pharmacological strategies for mitigating LKB1-deficient NSCLC tumor growth.

Inhibitors of ataxia–telangiectasia mutated (ATM), such as KU‐55933 (Ku), represent a promising class of novel anticancer drugs. In addition, the biguanide derivative phenformin exhibits antitumor activity superior to that of the AMPK activator metformin. Herein, we assessed the potential combinatorial therapeutic efficacy of phenformin and Ku when used to inhibit the growth of liver cancer cells, and we assessed the mechanisms underlying such efficacy. The Hep‐G2 and SMMC‐7721 liver cancer cell lines were treated with phenformin and Ku either alone or in combination, after which the impact of these drugs on cellular proliferation was assessed via 3‐(4,5‐dimethylthiazol) 2, 5‐diphenyltetrazolium and colony formation assays, whereas Transwell assays were used to gauge cell migratory activity. The potential synergy between these two drugs was assessed using the CompuSyn software, while flow cytometry was employed to evaluate cellular apoptosis. In addition, western blotting was utilized to measure p‐ATM, p‐AMPK, p‐mTOR, and p‐p70s6k expression, while mitochondrial functionality was monitored via morphological analyses, JC‐1 staining, and measurements of ATP levels. Phenformin and Ku synergistically impacted the proliferation, migration, and apoptotic death of liver cancer cells. Together, these compounds were able to enhance AMPK phosphorylation while inhibiting the phosphorylation of mTOR and p70s6k. These data also revealed that phenformin and Ku induced mitochondrial dysfunction as evidenced by impaired ATP synthesis, mitochondrial membrane potential, and abnormal mitochondrial morphology. These findings suggest that combination treatment with phenformin and Ku may be an effective approach to treating liver cancer via damaging mitochondria within these tumor cells.

Objective To investigate the effect of phenformin combined with hexokinase inhibitor 2-deoxyglucose (2-DG) on the treatment of triple-negative breast cancer cell lines 4T1 and MDA-MB-231. Methods Following treatment with phenformin, 2-DG or phenformin combined with 2-DG on 4T1 and MDA-MB-231 cells for 48 h, the cell proliferation in each group was detected by SRB and the apoptosis of cells was detected by flow cytometry. The concentration of glucose and lactic acid in cell culture supernatant was detected by ELISA. The activity of mitochondrial respiratory chain complex Ⅰ was detected by FlexStation3 and the mitochondrial oxygen consumption (OCR) was assayed with the Seahorse X Fe Analyzer. Results The hexokinase expression (4.6±0.17,3.73±0.21), glucose consumption (356±31, 397±42) μg/105 cells, Lactic acid concentration (5.59±0.52, 7.83±0.78) μmol/L in the supernatant of AT1 and MDA-MB-231 cells in Phenformin group were higher than that in control group ( 1±0.15, 1±0.12 ), ( 289±25, 301±32) μg/105cells, ( 2.37±0.18, 4.01±0.45) μmol/L (P < 0.01). Even if the dose was reduced by 90%, the cell viability of phenformin combined with 2-DG group (64.63±2.28, 51.97±2.29) % was still higher than that of phenformin group (86.70±1.83, 85.53±1.46) % (P<0.001). The combination of the two drugs significantly promoted the apoptosis of AT1 and MDA-MB-231. In addition, compared with the phenformin group (5.59±0.52, 7.83±0.78) μmol/L, the phenformin combined with 2-DG group (3.46±0.37, 5.18±0.62) μmol/L cell lactic acid production also greatly reduced (P<0.01). Compared with the phenformin or 2-DG single-drug group, the phenformin combined with 2-DG group can significantly inhibit the growth rate of tumors in tumor-bearing mice (P<0.01). The median survival time of tumor-bearing mice in the phenformin combined with 2-DG group was 72.5 d, which was higher than that in the phenformin group 57 d and 2-DG group 55.5 d (P<0.01). Conclusion Hexokinase inhibitor 2-DG significantly enhances the therapeutic effects of phenformin on triple-negative breast cancer cells. [ABSTRACT FROM AUTHOR]

Metformin is a widely prescribed medication for the treatment and management of type 2 diabetes. It belongs to a class of biguanides, which are characterized by a wide range of diverse biological properties, including anticancer, antimicrobial, antimalarial, cardioprotective and other activities. It is known that biguanides serve as excellent N-donor bidentate ligands and readily form complexes with virtually all transition metals. Recent evidence suggests that the mechanism of action of metformin and its analogues is linked to their metal-binding properties. These findings prompted us to summarize the existing data on the synthetic strategies and biological properties of various metal complexes with metformin and its analogues. We demonstrated that coordination of biologically active biguanides to various metal centers often resulted in an improved pharmacological profile, including reduced drug resistance as well as a wider spectrum of activity. In addition, coordination to the redox-active metal centers, such as Au(III), allowed for various activatable strategies, leading to the selective activation of the prodrugs and reduced off-target toxicity. [ABSTRACT FROM AUTHOR]

Cancer is one of the main causes of human mortality and brain tumors, including invasive pituitary adenomas, medulloblastomas and glioblastomas are common brain malignancies with poor prognosis. Therefore, the development of innovative management strategies for refractory cancers and brain tumors is important. In states of mitochondrial dysfunction – commonly encountered in malignant cells – cells mostly shift to anaerobic glycolysis by increasing the expression of LDHA (Lactate Dehydrogenase-A) gene. Oxamate, an isosteric form of pyruvate, blocks LDHA activity by competing with pyruvate. By blocking LDHA, it inhibits protumorigenic cascades and also induces ROS (reactive oxygen species)-induced mitochondrial apoptosis of cancer cells. In preclinical studies, oxamate blocked the growth of invasive pituitary adenomas, medulloblastomas and glioblastomas. Oxamate also increases temozolomide and radiotherapy sensitivity of glioblastomas. Oxamate is highly polar, which may preclude its clinical utilization due to low penetrance through cell membranes. However, this obstacle could be overcome with nanoliposomes. Moreover, different oxamate analogs were developed which inhibit LDHC4, an enzyme also involved in cancer progression and germ cell physiology. Lastly, phenformin, an antidiabetic agent, exerts anticancer effects via complex I inhibition in the mitochondria and leading the overproduction of ROS. Oxamate combination with phenformin reduces the lactic acidosis-causing side effect of phenformin while inducing synergistic anticancer efficacy. In sum, oxamate as a single agent and more efficiently with phenformin has high potential to slow the progression of aggressive cancers with special emphasis to brain tumors.

Biguanide drugs (metformin and phenformin) have drawn interest for potential cancer treatments, and laboratory studies show that some cancer cells are selectively sensitive to growth-inhibitory effects of biguanides. Examining metabolic pathways affected by biguanide treatments in cancer cells that are highly sensitive to biguanides, we found that biguanide treatment depletes cellular levels of both aspartate and NAD+. Experiments to replenish these metabolites or block steps of the aspartate-malate shuttle suggest that depletion of both metabolites, rather than either aspartate of NAD+ individually, is critical for growth-inhibitory effects of biguanide exposure. Even in sensitive cancer cells, though, biguanide treatment alone over a broad range of doses only inhibits cell replication without significantly affecting cell viability. Noting that clinical observations of biguanide efficacy have used combinations of agents that typically include cisplatin, we found that biguanide treatment at a cytostatic level substantially decreases survival of lung cancer and breast cancer cells when co-treated with cisplatin at doses that alone are also non-cytotoxic. This striking enhancement of cisplatin toxicity by biguanides depends on reductions of levels of NAD+ and aspartate, since addition of either of these metabolites prevented this potentiation of cisplatin cytotoxicity. Thus, biguanide drugs can have cytotoxic effects when used in combination with other cancer drugs, such as cisplatin, and depleting cellular levels of NAD+ and aspartate is critical for enhancing the cytotoxicity of cisplatin by biguanide drugs in sensitive cancer cells. [ABSTRACT FROM AUTHOR]

Radiotherapy (RT) is a common treatment for head and neck cancers, but central nervous system function can be impaired by clinical radiation doses. This experimental study evaluated the protective efficacy of the anti-hyperglycaemic/anti-neoplastic agent phenformin against radiation-induced developmental toxicity in zebrafish embryos. Zebrafish embryos pre-treated with 25 μM phenformin 1 h before x-ray irradiation were compared to irradiation-only embryos for mortality, hatching rate, morphology, spontaneous movement, heart beat, larval swimming, activities of the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT), malondialdehyde content (MDA, a by-product of membrane lipid oxidation), and acetylcholinesterase (AChE) activity. In addition, expression levels of multiple genes related to neural development and apoptosis (sod2, bdnf, ache, p53, bax, and bcl-2) were compared by RT-PCR and associated protein expression levels by western blotting. Pre-treatment with phenformin increased hatching rate, spontaneous movement, heart beat, and larval motor activity, decreased mortality and malformation rate, increased SOD, CAT, and AChE activities, and reduced MDA compared to irradiation-only embryos. The mRNA expression levels of anti-apoptotic sod2, bdnf, ache, and bcl-2 were enhanced while mRNA expression of p53 and pro-apoptotic bax were reduced in the phenformin pre-treatment group. Further, p53, Bax, and γ-H2AX (a biomarker of DNA damage) were downregulated while Bcl-2 and BDNF were upregulated by phenformin pre-treatment. Taken together, this study supports the protective efficacy of phenformin against radiation toxicity in zebrafish embryos by suppressing oxidative stress and ensuing apoptosis.

Inactivation of the LKB1 tumor suppressor is a frequent event in non-small cell lung carcinoma (NSCLC) leading to the activation of mTOR complex 1 (mTORC1) and sensitivity to the metabolic stress inducer phenformin. In this study, we explored the combinatorial use of phenformin with the mTOR catalytic kinase inhibitor MLN0128 as a treatment strategy for NSCLC bearing comutations in the LKB1 and KRAS genes. NSCLC is a genetically and pathologically heterogeneous disease, giving rise to lung tumors of varying histologies that include adenocarcinomas and squamous cell carcinomas (SCC). We demonstrate that phenformin in combination with MLN0128 induced a significant therapeutic response in KRAS/LKB1-mutant human cell lines and genetically engineered mouse models of NSCLC that develop both adenocarcinomas and SCCs. Specifically, we found that KRAS/LKB1-mutant lung adenocarcinomas responded strongly to phenformin + MLN0128 treatment, but the response of SCCs to single or combined treatment with MLN0128 was more attenuated due to acquired resistance to mTOR inhibition through modulation of the AKT-GSK signaling axis. Combinatorial use of the mTOR inhibitor and AKT inhibitor MK2206 robustly inhibited the growth and viability of squamous lung tumors, thus providing an effective strategy to overcome resistance. Taken together, our findings define new personalized therapeutic strategies that may be rapidly translated into clinical use for the treatment of KRAS/LKB1-mutant adenocarcinomas and squamous cell tumors.

Chun-Lin Kuo,1 Shu-Man Hsieh Li,*,2 Shu-Yi Liang,*,2 Shu-Ting Liu,2 Li-Chun Huang,2 Wei-Ming Wang,3 Li-Chen Yen,2,4 Shih-Ming Huang21Department of Orthopaedic Surgery, Tri-Service General Hospital, National Defense Medical Center, Taiwan, Republic of China; 2Department of Biochemistry, National Defense Medical Center, Taiwan, Republic of China; 3Department of Dermatology, Tri-Service General Hospital, National Defense Medical Center, Taiwan, Republic of China; 4Department of Microbiology and Immunology, National Defense Medical Center, Taiwan, Republic of China*These authors contributed equally to this workBackground: Differentiated embryo chondrocyte 1 (DEC1) is a helix-loop-helix transcription factor that directly binds to the class B E-box in target genes. DEC1 exerts both pro-survival and pro-apoptotic effects in a cell- and tissue-dependent manner. Its actions play role the progression of cancer remains unclear.Methods: We first examined the functional roles of DEC1 using the transient promoter reporter assay. Then, the knockdown of DEC1 expression was performed with the short hairpin RNA strategy in HeLa and A2058 cancer cell lines to check the cell cycle and mitochondrial function profile using the flow cytometry and Seahorse assays. We later clarified the role of DEC1 in the tumorigenesis using the colony formation, anchorage-independent growth assay, and cellular proliferation analysis.Results: In the present study, we tested two guanide-containing drugs, metformin and phenformin, and found that both exhibit cytotoxicity against HeLa cervical carcinoma and A2058 melanoma cells. This effect was mediated, at least in part, through activation of the AMPK pathway; degradation of important cellular proteins, such as DEC1 and p53; and suppression of mitochondrial function, colony formation, and anchorage-independent cell proliferation. Our results further suggest that the cytotoxicity of metformin and phenformin reflect the impact of the repressive actions of DEC1 on gene expression, including DEC1 itself. This in turn suppresses both anchorage-independent growth and cell proliferation.Conclusion: These findings provide several lines of evidence suggesting that DEC1 activity contributes to tumorigenicity and that the antitumor properties of biguanides reflect their ability to inhibit DEC1 functions.Keywords: differentiated embryo chondrocyte 1, p53, metformin, phenformin, tumorigenesis

The treatment of many skin inflammation diseases, such as psoriasis and atopic dermatitis, is still a challenge and inflammation plays important roles in multiple stages of skin tumor development, including initiation, promotion and metastasis. Phenformin, a biguanide drug, has been shown to play a more efficient anti-tumor function than another well-known biguanide drug, metformin, which has been reported to control the expression of pro-inflammatory cytokines; however, little is known about the effects of phenformin on skin inflammation. This study used a mouse acute inflammation model, ex vivo skin organ cultures and in vitro human primary keratinocyte cultures to demonstrate that phenformin can suppress acute skin inflammatory responses induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in vivo and significantly suppresses the pro-inflammatory cytokines IL-1β, IL-6 and IL-8 in human primary keratinocytes in vitro. The suppression of pro-inflammatory cytokine expression by phenformin was not directly through regulation of the MAPK or NF-κB pathways, but by controlling the expression of c-Myc in human keratinocytes. We demonstrated that the overexpression of c-Myc can induce pro-inflammatory cytokine expression and counteract the suppressive effect of phenformin on cytokine expression in keratinocytes. In contrast, the down-regulation of c-Myc produces effects similar to phenformin, both in cytokine expression by keratinocytes in vitro and in skin inflammation in vivo. Finally, we showed that phenformin, as an AMPK activator, down-regulates the expression of c-Myc through regulation of the AMPK/mTOR pathways. In summary, phenformin inhibits the expression of pro-inflammatory cytokines in keratinocytes through the down-regulation of c-Myc expression to play an anti-inflammation function in the skin.

Inhibitors of ataxia–telangiectasia mutated (ATM), such as KU‐55933 (Ku), represent a promising class of novel anticancer drugs. In addition, the biguanide derivative phenformin exhibits antitumor activity superior to that of the AMPK activator metformin. Herein, we assessed the potential combinatorial therapeutic efficacy of phenformin and Ku when used to inhibit the growth of liver cancer cells, and we assessed the mechanisms underlying such efficacy. The Hep‐G2 and SMMC‐7721 liver cancer cell lines were treated with phenformin and Ku either alone or in combination, after which the impact of these drugs on cellular proliferation was assessed via 3‐(4,5‐dimethylthiazol) 2, 5‐diphenyltetrazolium and colony formation assays, whereas Transwell assays were used to gauge cell migratory activity. The potential synergy between these two drugs was assessed using the CompuSyn software, while flow cytometry was employed to evaluate cellular apoptosis. In addition, western blotting was utilized to measure p‐ATM, p‐AMPK, p‐mTOR, and p‐p70s6k expression, while mitochondrial functionality was monitored via morphological analyses, JC‐1 staining, and measurements of ATP levels. Phenformin and Ku synergistically impacted the proliferation, migration, and apoptotic death of liver cancer cells. Together, these compounds were able to enhance AMPK phosphorylation while inhibiting the phosphorylation of mTOR and p70s6k. These data also revealed that phenformin and Ku induced mitochondrial dysfunction as evidenced by impaired ATP synthesis, mitochondrial membrane potential, and abnormal mitochondrial morphology. These findings suggest that combination treatment with phenformin and Ku may be an effective approach to treating liver cancer via damaging mitochondria within these tumor cells. [ABSTRACT FROM AUTHOR]

Metformin is a widely prescribed medication for the treatment and management of type 2 diabetes. It belongs to a class of biguanides, which are characterized by a wide range of diverse biological properties, including anticancer, antimicrobial, antimalarial, cardioprotective and other activities. It is known that biguanides serve as excellent N-donor bidentate ligands and readily form complexes with virtually all transition metals. Recent evidence suggests that the mechanism of action of metformin and its analogues is linked to their metal-binding properties. These findings prompted us to summarize the existing data on the synthetic strategies and biological properties of various metal complexes with metformin and its analogues. We demonstrated that coordination of biologically active biguanides to various metal centers often resulted in an improved pharmacological profile, including reduced drug resistance as well as a wider spectrum of activity. In addition, coordination to the redox-active metal centers, such as Au(III), allowed for various activatable strategies, leading to the selective activation of the prodrugs and reduced off-target toxicity.

Metabolic flexibility represents a potential point of attack for novel cancer treatments. We recently described the signaling mechanism inducing a metabolic shift in response to metformin and phenformin in leukemia and lymphoma cells. Enhanced glucose utilization was critically dependent on mitochondrial stress signaling/hypoxia-inducible factor-1α representing a therapeutic vulnerability.

Objective(s): Breast cancer remains a global challenge, and further chemopreventive therapies are still immediately required. Emerging evidence has revealed the potent anti-cancer effects of biguanides, Metformin (MET) and phenformin (PHE). Thus, to explore an efficient chemopreventive strategy for breast cancer, the antiproliferative effects of the combination of MET and PHE against breast cancer cells were assessed. Materials and Methods: Cytotoxicity of the drugs individually and in combination against T47D and MDA-MB-231 breast cancer cells were assessed using MTT assay and the median-effect method was used to analyze the precise nature of the interaction between MET and PHE. Besides, the expression levels of hTERT after 48 hr drug exposure were determined using qRT-PCR. Results: Based on the cytotoxicity assay, both MET and PHE further inhibited the growth of MDA-MB-231 cells compared with T47D cells. It was found that MET+PHE reduced the IC50s of MET and PHE in both cells drastically more than the single treatments in a synergistic manner. Importantly, MET+PHE showed higher antiproliferative effect with smaller IC50 values against MDA-MB-231 cells than against T47D cells. Real-time PCR results revealed that hTERT expression was significantly reduced in both breast cancer cell lines treated with MET+PHE than the single treatments. In comparison between two types of breast cancer cells, it was detected that MET+PHE could further decline hTERT expression in MDA-MB-231cells than in T47D cells (P

Background/Aims: Biguanides are anti-hyperglycaemic agents used to treat diabetes by acting primarily on the liver, inhibiting hepatic gluconeogenesis. However, biguanides may target other key metabolic tissues to exert beneficial actions. As the “master endocrine gland”, the pituitary is a true homeostatic sensor that controls whole body homeostasis and metabolism by integrating central and peripheral signals. However, whether the pituitary is a primary site of biguanides action in normal adult humans/primates remains unknown. Therefore, we aimed to elucidate the direct effects of two biguanides (metformin/phenformin) on the expression and secretion of all anterior pituitary hormones in two non-human primate species (Papio anubis and Macaca fascicularis), and the molecular/signalling-mechanisms behind these actions. Methods: Primary pituitary cell cultures from baboons and macaques were used to determine the direct impact of metformin/phenformin (alone and combined with primary regulators) on the functioning of all pituitary cell-types (i.e. expression/secretion/signaling-pathways, etc). Results: Metformin/phenformin inhibited basal, but not GHRH/ghrelin-stimulated GH/ACTH/ FSH-secretion and GH/POMC-expression, without altering secretion or expression of other pituitary hormones (PRL/LH/TSH), FSH-expression or cell viability in both primate models. These biguanide actions are likely mediated through modulation of: 1) common (mTOR/PI3K/intracellular-Ca2+mobilization) and distinct (MAPK) signaling pathways; and 2) gene expression of key receptors regulating somatotrope/corticotrope/gonadotrope function (i.e. upregulation of SSTR2/SSTR5/INSR/IGF1R/LEPR). Conclusion: The pituitary gland is a primary target of biguanide actions wherein they modulate somatotrope/corticotrope/gonadotrope-function through multiple molecular/signaling pathways in non-human primate-models. This suggests that the well-known metabolic effects of biguanides might be, at least in part, influenced by their actions at the pituitary level.

Abstract Background In previous studies, we have shown that the combination of metformin and gefitinib inhibits the growth of bladder cancer cells. Here we examined whether the metformin analogue phenformin, either used alone or in combination with gefitinib, could inhibit growth of bladder cancer cells. Methods The growth-inhibitory effects of phenformin and gefitinib were tested in one murine and two human bladder cancer cell lines using MTT and clonogenic assays. Effects on cell migration were assessed in a wound healing assay. Synergistic action between the two drugs was assessed using CompuSyn software. The potential involvement of AMPK and EGFR pathways in the effects of phenformin and gefitinib was explored using Western blotting. Results In MTT and clonogenic assays, phenformin was > 10-fold more potent than metformin in inhibiting bladder cancer cell growth. Phenformin also potently inhibited cell migration in wound healing assays, and promoted apoptosis. AMPK signaling was activated; EGFR signaling was inhibited. Phenformin was synergistic with gefitinib, with the combination of drugs showing much stronger anticancer activity and apoptotic activation than phenformin alone. Conclusions Phenformin shows potential as an effective drug against bladder cancer, either alone or in combination with gefitinib.

Phenformin (Phen), a potent activator of AMPK, is effective against some resistant cancers. This study evaluated the inhibition of proliferation, migration, invasion, and angiogenesis by Phen in aggressive cancer cells and investigated the underlying mechanism of the inhibition. Cholangiocarcinoma (CCA) KKU-156 and KKU-452 cells were used in this study. The results showed that Phen suppressed cell proliferation and induced apoptosis in both cells. Phen suppressed migration and invasion of cancer cells in wound healing and transwell chamber assays, respectively. The effects were associated with depletions of glutathione (GSH) and decreased glutathione redox ratio which represents cellular redox state. The redox stress was linked with the loss of mitochondrial transmembrane potential, as evaluated by JC-1 assay. The effect of Phen on angiogenesis was performed using HUVEC cultured cells. Phen alone did not affect tube formation of HUVEC cells. However, conditioned media from CCA cell cultures treated with Phen suppressed the tube-like structure formation. The antitumor effect of Phen was associated with AMPK activation and suppression of mTOR phosphorylation, HIF-1A, and VEGF protein expression. In conclusion, Phen inhibits cell proliferation, migration, invasion, and angiogenesis probably through AMPK-mTOR and HIF-1A-VEGF pathways. Phen may be repurposed as chemoprevention of cancer. [ABSTRACT FROM AUTHOR]

Medicines are usually prescribed for repeated use over shorter or longer times. Unfortunately, repeated-dose animal toxicity studies do not correlate well with observations in man. As emphasized by the '3Rs' and the desire to phase-out animal research, in vitro models are needed. One potential approach uses clinostat-cultured 3D HepG2--C3A liver-mimetic spheroids. They take 18 days to recover in vivo physiological functionality and reach a metabolic equilibrium, which is thereafter stable for a year. Acute and chronic repeated-dose studies of six drugs (amiodarone, diclofenac,metformin, phenformin, paracetamol and valproic acid) suggest that spheroids are more predictive of human in vivo toxicity than either 2D-cultured HepG2 cells or primary human hepatocytes. Repeated non-lethal treatment results in a clear response and return to equilibrium. Mitochondrial toxic compounds can be identified using a galactose-based medium. Some drugs induced a protective (or stress) response that intensifies after the second treatment. This 3D spheroid model is inexpensive, highly reproducible and well-suited for the determination of repeated-dose toxicity of compounds (naturally or chemically synthesized). [ABSTRACT FROM AUTHOR]

Diabetes mellitus is a serious metabolic disorder affecting millions of people worldwide. Phenformin and metformin are biguanide antidiabetic agents that are conveniently synthesized in a single‐step chemical reaction. Phenformin was once used to lower blood glucose levels, but later withdrawn from the market in several countries because it was frequently associated with lactic acidosis. Metformin is still a widely prescribed medication for the treatment of type 2 diabetes despite the introduction of several newer antidiabetic agents. Metformin is administered orally and has desirable pharmacokinetics. Incidence of metformin‐induced lactic acidosis is serious but very rare. Imeglimin, a novel molecule being investigated by Poxel and Sumitomo Dainippon Pharma in Japan, is currently in clinical trials for the treatment of type 2 diabetes. Unlike metformin, imeglimin is a cyclic molecule containing a triazine ring. However, like metformin, imeglimin is also a basic small molecule. Imeglimin is synthesized from metformin as a precursor via a single step chemical reaction. Recent mechanism of action studies suggests that imeglimin improves mitochondria function, when given in combination with metformin it helps achieve better glycemic control in patients with type 2 diabetes. We herein describe and compare the current status, synthesis, physicochemical properties, pharmacokinetic parameters, mechanism of action, and preclinical/clinical studies of metformin and imeglimin. [ABSTRACT FROM AUTHOR]

Radiotherapy (RT) is a common treatment for head and neck cancers, but central nervous system function can be impaired by clinical radiation doses. This experimental study evaluated the protective efficacy of the anti-hyperglycaemic/anti-neoplastic agent phenformin against radiation-induced developmental toxicity in zebrafish embryos. Zebrafish embryos pre-treated with 25 μM phenformin 1 h before x-ray irradiation were compared to irradiation-only embryos for mortality, hatching rate, morphology, spontaneous movement, heart beat, larval swimming, activities of the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT), malondialdehyde content (MDA, a by-product of membrane lipid oxidation), and acetylcholinesterase (AChE) activity. In addition, expression levels of multiple genes related to neural development and apoptosis (sod2, bdnf, ache, p53, bax, and bcl-2) were compared by RT-PCR and associated protein expression levels by western blotting. Pre-treatment with phenformin increased hatching rate, spontaneous movement, heart beat, and larval motor activity, decreased mortality and malformation rate, increased SOD, CAT, and AChE activities, and reduced MDA compared to irradiation-only embryos. The mRNA expression levels of anti-apoptotic sod2, bdnf, ache, and bcl-2 were enhanced while mRNA expression of p53 and pro-apoptotic bax were reduced in the phenformin pre-treatment group. Further, p53, Bax, and γ-H2AX (a biomarker of DNA damage) were downregulated while Bcl-2 and BDNF were upregulated by phenformin pre-treatment. Taken together, this study supports the protective efficacy of phenformin against radiation toxicity in zebrafish embryos by suppressing oxidative stress and ensuing apoptosis. [ABSTRACT FROM AUTHOR]

Dengue is the disease caused by dengue virus (DENV), and is associated with significant morbidity, mortality and economic cost worldwide. Despite its relevance, there is currently no specific, safe and effective pharmacological treatment against this disease. Our group was able to demonstrate the anti-DENV effect of metformin in both in vitro and in vivo models. On the other hand, the Pharmaceutical Chemistry Laboratory of the Faculty of Pharmacy of the Autonomous University of the State of Morelos developed new structural and functional analogues of metformin and phenformin. Considering the similarity of the analogues with metformin, it is possible that their therapeutic properties are similar and with it, the antiviral activity on DENV infection. Therefore, in the present work we analyzed the effect of various metformin and phenformin analogues on DENV-2 infection in Huh-7 cells. The results demonstrate that metformin and phenformin analogues are effective antiviral agents for inhibiting DENV infection in vitro and could be good candidates for the treatment of dengue. Among them, the analog EGL-1 and EGL-2 were found to have the most promising characteristics. Thus, the latter two compounds could be an alternative to metformin, the only biguanide currently available. [ABSTRACT FROM AUTHOR]

The results presented in this paper confirm the beneficial role of an easy-to-use and low-cost thin-layer chromatography (TLC) technique for describing the retention behavior and the experimental lipophilicity parameter of two biguanide derivatives, metformin and phenformin, in both normal-phase (NP) and reversed-phase (RP) TLC systems. The retention parameters (RF, RM) obtained under different chromatographic conditions, i.e., various stationary and mobile phases in the NP-TLC and RP-TLC systems, were used to determine the lipophilicity parameter (RMW) of metformin and phenformin. This study confirms the poor lipophilicity of both metformin and phenformin. It can be stated that the optimization of chromatographic conditions, i.e., the kind of stationary phase and the composition of mobile phase, was needed to obtain the reliable value of the chromatographic lipophilicity parameter (RMW) in this study. The fewer differences in the RMW values of both biguanide derivatives were ensured by the RP-TLC system composed of RP2, RP18, and RP18W plates and the mixture composed of methanol, propan-1-ol, and acetonitrile as an organic modifier compared to the NP-TLC analysis. The new calculation procedures for logP of drugs based on topological indices 0χν, 0χ, 1χν, M, and Mν may be a certain alternative to other algorithms as well as the TLC procedure performed under optimized chromatographic conditions. The knowledge of different lipophilicity parameters of the studied biguanides can be useful in the future design of novel and more therapeutically effective metformin and phenformin formulations for antidiabetic and possible anticancer treatment. Moreover, the topological indices presented in this work may be further used in the QSAR study of the examined biguanides.

Microglial functioning depends on Ca2+ signaling. By using Ca2+ sensitive fluorescence dye, we studied how inhibition of mitochondrial respiration changed spontaneous Ca2+ signals in soma of microglial cells from 5–7-day-old rats grown under normoxic and mild-hypoxic conditions. In microglia under normoxic conditions, metformin or rotenone elevated the rate and the amplitude of Ca2+ signals 10–15 min after drug application. Addition of cyclosporin A, a blocker of mitochondrial permeability transition pore (mPTP), antioxidant trolox, or inositol 1,4,5-trisphosphate receptor (IP3R) blocker caffeine in the presence of rotenone reduced the elevated rate and the amplitude of the signals implying sensitivity to reactive oxygen species (ROS), and involvement of mitochondrial mPTP together with IP3R. Microglial cells exposed to mild hypoxic conditions for 24 h showed elevated rate and increased amplitude of Ca2+ signals. Application of metformin or rotenone but not phenformin before mild hypoxia reduced this elevated rate. Thus, metformin and rotenone had the opposing fast action in normoxia after 10–15 min and the slow action during 24 h mild-hypoxia implying activation of different signaling pathways. The slow action of metformin through inhibition of complex I could stabilize Ca2+ homeostasis after mild hypoxia and could be important for reduction of ischemia-induced microglial activation.

Abstract Background The Connectivity Map (CMAP) database, an important public data source for drug repositioning, archives gene expression profiles from cancer cell lines treated with and without bioactive small molecules. However, there are only one or two technical replicates for each cell line under one treatment condition. For such small-scale data, current fold-changes-based methods lack statistical control in identifying differentially expressed genes (DEGs) in treated cells. Especially, one-to-one comparison may result in too many drug-irrelevant DEGs due to random experimental factors. To tackle this problem, CMAP adopts a pattern-matching strategy to build “connection” between disease signatures and gene expression changes associated with drug treatments. However, many drug-irrelevant genes may blur the “connection” if all the genes are used instead of pre-selected DEGs induced by drug treatments. Methods We applied OneComp, a customized version of RankComp, to identify DEGs in such small-scale cell line datasets. For a cell line, a list of gene pairs with stable relative expression orderings (REOs) were identified in a large collection of control cell samples measured in different experiments and they formed the background stable REOs. When applying OneComp to a small-scale cell line dataset, the background stable REOs were customized by filtering out the gene pairs with reversal REOs in the control samples of the analyzed dataset. Results In simulated data, the consistency scores of overlapping genes between DEGs identified by OneComp and SAM were all higher than 99%, while the consistency score of the DEGs solely identified by OneComp was 96.85% according to the observed expression difference method. The usefulness of OneComp was exemplified in drug repositioning by identifying phenformin and metformin related genes using small-scale cell line datasets which helped to support them as a potential anti-tumor drug for non-small-cell lung carcinoma, while the pattern-matching strategy adopted by CMAP missed the two connections. The implementation of OneComp is available at https://github.com/pathint/reoa . Conclusions OneComp performed well in both the simulated and real data. It is useful in drug repositioning studies by helping to find hidden “connections” between drugs and diseases.

Activations of Akt or ERK pathway induced by clinical drugs promote therapeutic failure due to decrease of drug response, and no available strategies have been developed to solve these problems. In this study, we found that pirarubicin (THP), one important chemotherapeutic drug for treating bladder cancer intravesically, dramatically elevated phosphorylations of both Akt and Erk1/2 in addition to inducing DNA damage. MK2206 or AZD6244, representative Akt and Erk1/2 inhibitors, respectively, profoundly sensitized bladder cancer cells to THP treatment. Interestingly, we found that inhibition of a single arm of either Akt or Erk1/2 pathway would induce the increase of another arm, indicating the existence of the crosstalk between these two pathways. Thus, simultaneous suppression of both signals may be needed for increasing the sensitivity of THP. On the other hand, we revealed that phenformin efficiently inhibited both Akt and Erk1/2 phosphorylation in a dose-dependent manner. Furthermore, we demonstrated that phenformin, mimicking dual inhibitors, plays dramatically synergistic action with THP both in vitro and in vivo. Our findings suggest that combination therapy of THP with dual inhibitors may constitute a successful strategy for improving chemotherapy response.

Adipose tissue macrophages (ATMs) are crucial in maintaining a low-grade inflammatory microenvironment in adipose tissues (ATs). Modulating ATM polarization to attenuate inflammation represents a potential strategy for treating obesity with insulin resistance. This study develops a combination therapy of celastrol (CLT) and phenformin (PHE) using chondroitin sulfate-derived micelles. Specifically, CLT-loaded 4-aminophenylboronic acid pinacol ester-modified chondroitin sulfate micelle (CS-PBE/CLT) and chondroitin sulfate-phenformin conjugate micelles (CS-PHE) were synthesized, which were shown to actively target ATs through CD44-mediated pathways. Furthermore, the dual micellar systems significantly reduced inflammation and lipid accumulation via protein quantification and Oil Red O staining. In preliminary in vivo studies, we performed H&E staining, immunohistochemical staining, insulin tolerance test, and glucose tolerance test, and the results showed that the combination therapy using CS-PBE/CLT and CS-PHE micelles significantly reduced the average body weight, white adipose tissue mass, and liver mass of high-fat diet-fed mice while improving their systemic glucose homeostasis. Overall, this combination therapy presents a promising alternative to current treatment options for diet-induced obesity.

In the 1920s, guanidine, the active component of Galega officinalis, was shown to lower glucose levels and used to synthesize several antidiabetic compounds. Metformin (1,1 dimethylbiguanide) is the most well-known and currently the only marketed biguanide in the United States, United Kingdom, Canada, and Australia for the treatment of non-insulin-dependent diabetes mellitus. Although phenformin was removed from the US market in the 1970s, it is still available around the world and can be found in unregulated herbal supplements. Adverse events associated with therapeutic use of biguanides include gastrointestinal upset, vitamin Bi 2 deficiency, and hemolytic anemia. Although the incidence is low, metformin toxicity can lead to hyperlactatemia and metabolic acidosis. Since metformin is predominantly eliminated from the body by the kidneys, toxicity can occur when metformin accumulates due to poor clearance from renal insufficiency or in the overdose setting. The dominant source of metabolic acidosis associated with hyperlactatemia in metformin toxicity is the rapid cytosolic adenosine triphosphate (ATP) turnover when complex I is inhibited and oxidative phosphorylation cannot adequately recycle the vast quantity of H + from ATP hydrolysis. Although metabolic acidosis and hyperlactatemia are markers of metformin toxicity, the degree of hyperlactatemia and severity of acidemia have not been shown to be of prognostic value. Regardless of the etiology of toxicity, treatment should include supportive care and consideration for adjunct therapies such as gastrointestinal decontamination, glucose and insulin, alkalinization, extracorporeal techniques to reduce metformin body burden, and metabolic rescue. [ABSTRACT FROM AUTHOR]

Activations of Akt or ERK pathway induced by clinical drugs promote therapeutic failure due to decrease of drug response, and no available strategies have been developed to solve these problems. In this study, we found that pirarubicin (THP), one important chemotherapeutic drug for treating bladder cancer intravesically, dramatically elevated phosphorylations of both Akt and Erk1/2 in addition to inducing DNA damage. MK2206 or AZD6244, representative Akt and Erk1/2 inhibitors, respectively, profoundly sensitized bladder cancer cells to THP treatment. Interestingly, we found that inhibition of a single arm of either Akt or Erk1/2 pathway would induce the increase of another arm, indicating the existence of the crosstalk between these two pathways. Thus, simultaneous suppression of both signals may be needed for increasing the sensitivity of THP. On the other hand, we revealed that phenformin efficiently inhibited both Akt and Erk1/2 phosphorylation in a dose-dependent manner. Furthermore, we demonstrated that phenformin, mimicking dual inhibitors, plays dramatically synergistic action with THP both in vitro and in vivo. Our findings suggest that combination therapy of THP with dual inhibitors may constitute a successful strategy for improving chemotherapy response. [ABSTRACT FROM AUTHOR]