BackgroundLentinus edodes is the second most widely cultivated edible mushroom globally. This study aimed to enrich L. edodes with selenium, quantify total selenium and its species, and evaluate the in vivo antidiabetic activity of selenium-enriched extracts using Wistar rats as the model organism.MethodsThe experiment involved healthy male Wistar strain laboratory rats, divided into 8 groups. Along with general health status, the animals' body weight, blood glucose levels and lipid concentrations as well as selenoprotein P1 and glutathione peroxidase 1 (GPx-1) expression were monitored during the experiment.ResultsThe highest selenium accumulation was observed in the first flushes, reaching 90.4 ± 9.0 mg Se kg-1. Selenomethionine (Se-Met) was identified as the sole selenium species in both aqueous and enzymatic extracts. The findings reveal subtle variations in the physiological effects of L. edodes harvested at different times, particularly regarding body weight, blood glucose, cholesterol, and triglyceride levels, as well as pancreatic tissue health and the expression of selenoprotein P1 and GPx-1 in rats. Selenium-enriched L. edodes extracts exhibited notable antidiabetic properties, especially in regulating blood glucose and preserving pancreatic β-cell structure in an alloxan-induced diabetes model. A significant positive correlation was found between selenoprotein P1 levels and total cholesterol (ρ = 0.622; p < 0.001).DiscussionThe findings of this study provide a basis for monitoring the quality and reproducibility of the selenium fortification process. Furthermore, these results suggest that further investigation in additional animal models is warranted to confirm the broader applicability and robustness of this approach.

Background/Objectives: Metformin is a prominent candidate for cancer drug repurposing, backed by preclinical and epidemiological evidence showing reduced cancer incidence in diabetic patients. Its pleiotropic effects include AMPK activation, protein synthesis inhibition, and metabolic alterations. This review integrates global preclinical data via a comprehensive tabular overview alongside a cross-analysis of specific investigations of hamster fibrosarcoma. Since head-to-head comparisons of metformin-based combinations on hamster fibrosarcoma remain limited, this work performs an integrated cross-study evaluation to establish a clear comparative hierarchy of various repurposed adjuvants combined with metformin. Methods: A literature review and cross-study re-analysis of peer-reviewed preclinical studies were conducted, focusing on in vivo therapeutic outcomes within the BHK-21/C13-induced hamster fibrosarcoma model. Treatment regimens from distinct primary studies were evaluated side-by-side using tumor endpoint data expressed as a percentage of control mean (± SD). Only statistically significant pairwise differences (p < 0.05) determined the comparative hierarchy. Results: Integrated analysis identified disulfiram and diclofenac as the most promising adjuvants among compared metformin combinations, showing the highest statistical significance across all evaluated endpoints. The statistical ranking of the metformin-based combinations followed a descending order: disulfiram, diclofenac, nitroglycerin, itraconazole, and caffeine. Conclusions: By synthesizing previously fragmented primary data into a unified comparative framework, these findings provide a strong, consolidated preclinical rationale for metformin-based combination strategies. The established hierarchy of adjuvant potency resolves structural ambiguity and supports further translational and clinical investigation to evaluate their therapeutic potential in fibrosarcoma and other malignancies.

Background/Objectives: The guinea pig is a unique experimental model because of the evolutionary loss of the GULO gene, which encodes an enzyme involved in vitamin C synthesis. Since vitamin C plays an essential role in collagen biochemistry, numerous studies have investigated the effects of pre- and postnatal vitamin C deficiency. However, only a few studies, including ours, have indicated a possible link between vitamin C deprivation and potential weakening of the basement membrane, which may lead to significant alterations in brain structure. Methods: The experiment included guinea pig foetuses completely deprived from the 10th (E2 group) and the 20th (E1 group) to the 50th day of intrauterine life. Tissue samples from the cerebrum and cerebellum were taken for biochemical, molecular, and immunohistochemical analyses. Results: In the E2 group alone, we found marked gross changes: cerebral bleeding, porencephaly, and a lissencephalic cerebellar surface. Microscopic examination revealed diffuse bleeding in the cerebrum along with a loss of neurons in the area of the defect, specifically in the E2 group. The complete maturation of ectopic neurons characterised dysplastic changes in the cerebellum. Hydroxyproline analysis of both the cerebrum and cerebellum showed no significant differences among the E1, E2, and control groups. However, decreased expression of COL1, COL4A1, and SLC23A1 was observed solely in the cerebellar tissue of the E1 group. Conclusions: The morphological, biochemical, and molecular results represent preliminary associations with vitamin C deficiency, but require further validation.

The prevention of nutrition-related diseases holds paramount significance for human well-being. Sweet basil (Ocimum basilicum L., Lamiaceae) emerges as a noteworthy candidate due to its robust antioxidant properties attributed to a high concentration of phenolic and flavonoid compounds, promising potential health benefits for humans. Notably, bile acids and their derivatives exert influence on the metabolic effects of both conventional and herbal drugs. The aim of this study was to examine the effects of administering an aqueous basil extract for seven days on glycemia and in vivo antioxidant activity in both healthy and diabetic animals, either alone or in combination with a bile acid derivative. The experimental design involved normoglycemic and diabetic Wistar rats subjected to a seven-day regimen of saline and basil water extract. Hyperglycemia was induced using alloxan. Following the treatment period, animals were euthanized by cardiopuncture, and serum analyses were conducted to assess fasting blood glucose levels and biochemical parameters. Additional assessments included oral glucose tolerance tests and antioxidative stress enzyme assays. The findings showed a significant hypoglycemic effect of the aqueous basil extract in both normoglycemic and diabetic animals. The extract also decreased lipid peroxidation and increased the activity of antioxidant enzymes. Basil extract treatment displayed protective effects on glycemia in both normoglycemic and diabetic animals, indicating considerable antioxidant potential. These effects were evident through increased antioxidant enzyme activity and decreased lipid peroxidation, affirming the beneficial impact of aqueous basil extract on health parameters.

The edible mushroom Coprinus comatus has a long history of use in metabolic diseases, which is increasingly documented by modern research. Due to its favorable nutritional composition, it was assumed that this mushroom could accelerate tissue recovery after acutely induced damage with subsequent disturbance of primarily carbohydrate metabolism. To test this hypothesis, the alloxan diabetes model was used, where experimental animals’ change in body weight and biochemical and histological indicators of recovery were monitored. Before performing the in vivo part, HPLC analysis of the C. comatus extract was carried out with subsequent in silico and in vitro tests. Comparing the animals treated with the mushroom in three different doses, no significant change in body weight was observed. Still, the change was also noticed in the lipid status and glycemia, with a dose-dependent beneficial effect. Morphometric analysis of pancreatic tissue stained by immuno-histochemical methods showed that long-term treatment with C. comatus leads to increased numerical density, nuclear volume, and absolute number of beta cells of the islets of Langerhans, which suffered severe damage after alloxan administration. Overall, C. comatus may contribute to faster tissue recovery after acute diabetic-relevant damage with chronic consequences.

In the original publication [...].

Backgrounds: Burn injuries present significant medical challenges due to their complexity in healing and potential for severe scarring. This study evaluates the efficacy of Manuka honey in accelerating burn wound healing compared to conventional antibiotic ointments. Methods: Using a porcine model resembling human skin, nine Landrace breed female pigs with standardized deep dermal burns were treated with either Manuka honey in alginate or a combination of antibiotic ointments. Wound healing was assessed through macroscopic evaluation, a histopathological analysis, and immunohistochemical staining over a 60-day period. Results: Our findings indicate that the Manuka honey treatment was associated with significantly increased collagen density in the treated wounds compared to the control group (p < 0.05). The immunohistochemical analysis revealed lower macrophage activity (Iba1 staining) and a reduction in Ki67 expression on days 10 and 17 in the Manuka honey group, suggesting a more rapid transition toward tissue remodeling. The quantitative analysis showed a trend toward delayed epithelialization and increased inflammation in the control group, while wounds treated with Manuka honey exhibited faster reepithelialization and improved epidermal regeneration. However, additional studies are required to further assess collagen fiber organization and overall dermal architecture. Conclusions: These findings support the potential of Manuka honey as a beneficial treatment for burn wound healing, with evidence of enhanced reepithelialization and collagen deposition. Further research, including clinical trials, is necessary to fully elucidate its role in clinical practice and optimize treatment protocols.

The pineal gland is a small but essential endocrine gland that secretes the hormone melatonin, which regulates thehuman circadian rhythm. This study aims to analyze regressive changes in human pineal gland tissue by examining the expression ofGFAP, vimentin, and CD34. The study included 30 paraffin-embedded blocks of human pineal glands obtained from cadaver autopsies.Samples were stained immunohistochemically with the antibodies GFAP, Vimentin, and CD34. Morphometric data were obtained usingthe threshold color plugin in Fiji software, and the results were analyzed by age and gender. A statistically significant difference wasfound in the areal fraction occupied by vimentin between Groups I and II, and Groups I and III. The average areal density of the GFAPmarker is highest in the youngest cohort. No statistically significant difference was noted among the analyzed groups in comparing theresults. Statistical analysis revealed a statistically significant difference in the density of blood vessels between Group I and Group II. Thequantity of pinealocytes and astrocytes diminishes over time, as evidenced by the reduction in the positivity of GFAP-positive cells.Vimentin positivity demonstrates the increase in the proportion of stroma throughout the aging process. Furthermore, the rise in the densityof blood vessels, identified by anti-CD34, is relative to their localization within the stroma, which becomes predominant over time.

Numerous studies reported about potential effects of L-carnosine in regulation of tumor growth and metabolism. We evaluated the effects of different concentrations of L-carnosine from Karnozin EXTRA (R) supplement on mitochondrial respiratory chain complexes of human embryo lung fibroblasts (MRC-5) and human breast cancer cells (MCF-7), with different energy pathways. Also, we analyzed the proliferation index and expression of various markers of oxidative stress. Treatment with Karnozin EXTRA (R) (concentration of L-carnosine were 2, 5 and 10 mM) for 24 hours gradually decreased the number of cells and changed their morphological features. In both cell lines, a dose-dependent reduction of cell viability was recorded compared to the control group. Also, experimental groups showed a concentration-dependent decrease in fluorescence intensity of SOD2 expressions in MCF-7, while in MRC-5 we noticed higher fluorescence intensity in Carnosine 2 mM group. Treated cells, in both cell lines, showed different intensity of iNOS cytoplasmic immunopositivity in a concentration-dependent manner. In all experimental groups, we noticed an increased expression of marker of oxidative stress-cytochrome P450 2E1 (CYP2E1). The effects of Karnozin EXTRA (R) capsule on mitochondrial respiration, assessed with the Clark-type electrode, were manifested as a reduction of: basal cell respiration, maximum capacity of electron transport chain and mitochondrial ATP-linked respiration. Also, significant decrease in the activity of complex I (NADH-ubiquinone oxidoreductase), complex II (succinate dehydrogenase) and complex IV (cytochrome c oxidase) was observed in both cell lines. Bearing in mind that Karnozin EXTRA (R) is a potential regulator of energy metabolism of MCF-7 and MRC-5, these results provide a good basis for further preclinical and clinical research.

The aim was to detect and correlate anticancer effects of metformin in combinations with other repurposed drugs, already registered for other indications, which may be immediately applied and clinically investigated in oncology, reducing the time and cost of research for new cancer treatments. Immunohistochemistry was performed for tumors treated by dual drug combinations containing metformin with deoxycholic acid, caffeine, itraconazole, nitroglycerin, disulfiram or diclofenac. The drugs were applied in Syrian golden hamsters (6 animals per group) with the inoculated BHK21/C13 fibrosarcoma in doses equivalent to usual human doses, <50 % LD50. The anticancer effects were assessed by: p53 (mutational status); Ki-67 and PCNA (tumor proliferation); CD34 and CD31 (neoangiogenesis); GLUT1 (glucose metabolism); iNOS (NO metabolism); COX4, Cytochrome C and caspase 3 (apoptosis); immunohistochemical markers. Also, biophysical characteristics of fibrosarcoma, animal blood samples and the toxicity on main organs were analyzed. Treatments significantly (P < 0.05) reduced mutational status, tumor proliferation, neoangiogenesis, glucose metabolism, NO metabolism and modulated apoptosis, in correlation with tumor size, without toxicity and influence on biochemical blood and hematological tests. The administration of metformin in two-drug combination with deoxycholic acid, caffeine, itraconazole, nitroglycerin, disulfiram or diclofenac may be recommended for further clinical investigations in oncology.

Background: Alarming data revealed that 19% to 34% of adults with diabetes mellitus develop chronic wounds, which are characterized by impaired healing and a higher risk of infections. Inspired by the traditional use of immortelle for wound healing and the lack of scientific evidence regarding how it thoroughly influences tissue regeneration, we aimed to formulate a hydrogel loaded with immortelle essential oil and assess its effectiveness on diabetic excision wounds. Methods: The rheological properties of the hydrogel, an in vivo safety test, as well as wound healing capacity, were determined in rats with induced diabetes and excision wounds. Diabetic rats were divided into four groups: untreated, treated with 1% silver sulfadiazine ointment, treated with a gel base, and treated with the immortelle essential oil-based hydrogel. Results: It was revealed that the hydrogel exerts pseudoplastic behavior and has no potential to act as an irritant, thus highlighting its suitability for skin application. Moreover, analysis of macroscopic, biochemical, and histopathological data revealed that the immortelle essential oil-based hydrogel significantly improves wound repair. Superior re-epithelialization, scar maturation, and increased collagen fiber density were achieved after immortelle essential oil-based gel application. Conclusions: These findings suggest that the immortelle essential oil-based hydrogel could be a natural, safe, and effective wound-healing dressing.

Background and Objective: The health benefits of isoflavone consumption are often attributed to its antioxidant properties. Since the gut microbiome has a large impact on isoflavone metabolism, probiotics might enhance isoflavone bioavailability and in vivo antioxidant activity. The aim of this study was to assess the antioxidant effects of isoflavones, probiotics and their combination on Carbon Tetrachloride (CCl4)-induced oxidative stress in rats. Materials and Methods: Wistar rats were allocated to seven groups of six animals, group I: Control group received saline solution (1 mL/kg p.o.), II: Soy isoflavones (50 mg/kg p.o.), III: Probiotics (10(9) CFU/kg p.o.), IV: CCl4 (1 mg/kg i.p.) and V-VII: After CCl4, which was fed isoflavones, probiotics or their combination for 14 days, after which the animals were sacrificed. The antioxidative effects of isoflavones, probiotics and their combination were determined by biochemical parameters in serum, the oxidative stress parameters in liver and kidney homogenates and by histological assessment. Results: In comparison to those in the CCl4 treatment group, the functional markers of hepatotoxicity, particularly Alanine Aminotransferase (ALT) and Aspartate Aminotransferase (AST) activity, were significantly lower in the group treated with the combination of isoflavones and probiotics. This combination successfully reduced the CCl4-induced increase in malondialdehyde levels in liver tissue, while histopathological changes in hepatocytes, were notably attenuated. Conclusion: The results indicate that the combined consumption of isoflavones and probiotics inhibits the level of lipid peroxidation and has a hepatocurative effect on CCl4-induced oxidative stress in rats, significantly improving the functional and morphological parameters of the liver.

Background: The 50% maximum probability of breech presentation (BP) implies that the same number of fetuses/newborns with the same characteristics take either BP or cephalic presentation (CP) with equal probability. Sirenomelia offers the possibility to investigate the incidence of BP in the simultaneous presence of several medical entities, each associated with an increased incidence of BP (Potter anomaly, lower limb fusion, prematurity, additional medical entities). The aim of the study is to investigate the probability of BP at delivery in sirenomelic singleton gestations to confirm the rule that the maximum probability for BP is 50%. Methods: A systematic retrospective literature search using the keywords: sirenomelia, sympus apus, sympus monopus, and sympus dipus, in the SCOPUS database. Inclusion criteria: presentation of singleton fetuses at delivery in the second and third trimester of pregnancy. Comparison with chi-square test (level of significance p < 0.5) of overall incidence of BP and CP as well as depending on the type of sirenomelia, prematurity, presence of additional medical entities associated with increased incidence of BP. Results: In total, 67 articles were identified, and 83 cases were found. The number of cases in BP vs. CP with a statistical difference to expected 50%:50% distribution of presentations: All cases—BP44 CP39 p = 0.583; Premature birth—BP25 CP20 p = 0.456; Term birth—BP17 CP18 p = 0.866; Sympus apus—BP15 CP14 p = 0.853; Sympus monopus—BP6 CP9 p = 0.439; Sympus dipus—BP21 CP14 p = 0.237; With additional entities—BP16 CP10 p = 0.239; Without additional entities—BP23 CP26 p = 0.668. Also, there were no statistically significant differences in the distribution of presentations between premature and term birth (p = 0.589), three types of sirenomelia (p = 0.601), with and without additional entities (p = 0.228). Conclusions: This study supports the rule that the maximum probability for BP in any medical entity, including sirenomelia, is 50%. The outcome of the BP should be evaluated with the case-control matching procedure: subtract the number of fetuses/newborns from the CP group that is identical to the number of fetuses/newborns in the BP group, with identical characteristics, and add this group to the BP group before comparing them to the rest of the CP fetuses/newborns.

Brain malformations represent a heterogeneous group of abnormalities of neural morphogenesis, often associated with aberrations of neuronal connectivity and brain volume. Prenatal detection of brain malformations requires a clear understanding of embryology and developmental morphology through the various stages of gestation. This expert panel review is written with the central aim of providing an easy-to-understand roadmap to improve prenatal detection and characterization of structural malformations based on the current understanding of normal and aberrant brain development. The utility of each available neuroimaging modality including prenatal multiplanar neurosonography, anatomical magnetic resonance imaging (MRI), and advanced MRI techniques, as well as further insights from post-mortem imaging have been highlighted for every developmental stage.

This study aimed to develop novel topical formulations based on a natural component (0.5% of Siberian pine essential oil) and to assess its wound-healing capacity through macroscopic, histopathological, and biochemical examination. The phytochemical profile of Pinus sibirica essential oil (PSEO) and rheological analysis and safety potential of formulations were determined. The wound-healing effect was evaluated on an excision wound model in diabetic Wistar albino rats randomly divided into the following groups topically treated with (1) untreated, (2) 1% silver sulfadiazine, (3) ointment base, (4) gel base, (5) PSEO ointment, and (6) PSEO gel. Formulations containing PSEO were stable and safe for skin application. Three weeks of treatment with both PSEO formulations (ointment and gel) led to a significant reduction in wound size (98.14% and 96.28%, respectively) and a remarkably higher level of total hydroxyproline content (9.69 µg/mg and 7.26 µg/mg dry tissue, respectively) relative to the control group (65.97%; 1.81 µg/mg dry tissue). These findings were in correlation with histopathological results. Topically applied PSEO formulations were associated with a significant reduction in most of the measured pro-oxidants and enhanced activity of the antioxidant defense system enzymes (p < 0.05). Our findings showed that gel and ointment with PSEO demonstrated significant wound-repairing capabilities in the excision wound model.

The framework of this study was a comprehensive investigation of Morus nigra L. extracts, with the aim to establish the correlation between chemical composition and antioxidant/hepatoprotective activity of a series of black mulberry extracts obtained from aerial parts of the plant. Black mulberry leaf (MLEE), bark (MBEE), juice (MJ) and fresh fruit (MFEE) extracts were obtained using the conventional Soxhlet extraction, while the supercritical CO2 extraction procedure was employed for preparation of the seed oil (MSO). Analysis of the chemical composition was performed using spectrophotometric, HPLC and GC methods. For the evaluation of antioxidant activity, in vitro FRAP and DPPH assays were applied. In Haan strain NMRI mice with streptozotocin-induced oxidative stress, in vivo antioxidant activity and liver tissue integrity were examined. The content of polyphenolic compounds was the highest in MBEE (68.3 ± 0.7 mgGAE/g) with the most abundant compounds being polyphenolic acids, followed by MLEE (23.4 ± 0.5 mgGAE/g) with the flavonoids isoquercetin and rutin being present in a significant amount. An analysis of MSO revealed a high content of γ-linoleic acid. The highest antioxidant activity in vitro (FRAP and DPPH) was observed for MLEE, MBEE and MSO. Beneficial effects were confirmed in vivo, with lower values of hepatosomatic index, potentiation of the activity of the enzymes superoxide dismutase and catalase, a lower rate of lipid peroxidation and reduced positivity for the P450 enzyme in animals treated with MLEE, MBEE and MSO. Black mulberry leaf and bark extracts as well as seed oil exhibited significant antioxidant activity. Apart from the confirmed biological properties of the fruit and leaf extracts, the observed activities of black mulberry seed oil and bark extract imply its importance as a sustainable source of phytochemicals.

Hereditary spastic paraplegias (HSP) are rare, inherited neurodegenerative or neurodevelopmental disorders that mainly present with lower limb spasticity and muscle weakness due to motor neuron dysfunction. Whole genome sequencing identified bi-allelic truncating variants in AMFR , encoding a RING-H2 finger E3 ubiquitin ligase anchored at the membrane of the endoplasmic reticulum (ER), in two previously genetically unexplained HSP-affected siblings. Subsequently, international collaboration recognized additional HSP-affected individuals with similar bi-allelic truncating AMFR variants, resulting in a cohort of 20 individuals from 8 unrelated, consanguineous families. Variants segregated with a phenotype of mainly pure but also complex HSP consisting of global developmental delay, mild intellectual disability, motor dysfunction, and progressive spasticity. Patient-derived fibroblasts, neural stem cells (NSCs), and in vivo zebrafish modeling were used to investigate pathomechanisms, including initial preclinical therapy assessment. The absence of AMFR disturbs lipid homeostasis, causing lipid droplet accumulation in NSCs and patient-derived fibroblasts which is rescued upon AMFR re-expression. Electron microscopy indicates ER morphology alterations in the absence of AMFR. Similar findings are seen in amfra-/- zebrafish larvae, in addition to altered touch-evoked escape response and defects in motor neuron branching, phenocopying the HSP observed in patients. Interestingly, administration of FDA-approved statins improves touch-evoked escape response and motor neuron branching defects in amfra-/- zebrafish larvae, suggesting potential therapeutic implications. Our genetic and functional studies identify bi-allelic truncating variants in AMFR as a cause of a novel autosomal recessive HSP by altering lipid metabolism, which may potentially be therapeutically modulated using precision medicine with statins.

We describe the preparation, dynamic, assembly characteristics of vase-shaped basket 13- along with its ability to form an inclusion complex with anticancer drug mitoxantrone in abiotic and biotic systems. This novel cavitand has a deep nonpolar pocket consisting of three naphthalimide sides fused to a bicyclic platform at the bottom while carrying polar glycines at the top. The results of 1H Nuclear Magnetic Resonance (NMR), 1H NMR Chemical Exchange Saturation Transfer (CEST), Calorimetry, Hybrid Replica Exchange Molecular Dynamics (REMD), and Microcrystal Electron Diffraction (MicroED) measurements are in line with 1 forming dimer [12]6-, to be in equilibrium with monomers 1(R)3- (relaxed) and 1(S)3- (squeezed). Through simultaneous line-shape analysis of 1H NMR data, kinetic and thermodynamic parameters characterizing these equilibria were quantified. Basket 1(R)3- includes anticancer drug mitoxantrone (MTO2+) in its pocket to give stable binary complex [MTO subset of 1]- (Kd=2.1 mu M) that can be precipitated in vitro with UV light or pH as stimuli. Both in vitro and in vivo studies showed that the basket is nontoxic, while at a higher proportion with respect to MTO it reduced its cytotoxicity in vitro. With well-characterized internal dynamics and dimerization, the ability to include mitoxantrone, and biocompatibility, the stage is set to develop sequestering agents from deep-cavity baskets. Deep-cavity baskets are novel, accessible, modular, and biocompatible hosts. With intriguing conformational dynamics, assembly characteristics and great capacity to bind anticancer drug mitoxantrone, deep-cavity baskets hold promise for developing antidotes as a countermeasure against a broad range of aromatic toxicants in complex biological systems.image

We examined whether combinig diclofenac and metformin in doses equivalent to human doses would synergize their anticancer activity on fibrosarcoma inoculated to hamsters and in vitro. Rescue experiment was performed to examine whether the prosurvival NF-κB stimulation by mebendazole can reverse anticancer effects of the treatment. BHK-21/C13 cell culture was subcutaneously inoculated to Syrian golden hamsters randomly divided into groups (6 animals per group): 1) untreated control; treated daily with 2) diclofenac; 3) metformin; 4) combinations of diclofenac and metformin at various doses; 5) combination of diclofenac, metformin and mebendazole; 6) mebendazole. Dose response curves were made for diclofenac and metformin combination. Tumor growth kinetics, biophysical, pathological, histological and immunohistochemical characteristics of excised tumors and hamster organs as well as biochemical and hematological blood tests were compared among the groups. Single treatments had no anticancer effects. Diclofenac (60 mg/kg/day) exhibited significant (P < 0.05) synergistic inhibitory effect with metformin (500 mg/kg/day) on all tumor growth parameters, without toxicity and influence on biochemical and hematological blood tests. The same results were obtained with double doses of diclofenac and metformin combination. The addition of mebendazole to the diclofenac and metformin combination rescued tumor expansion. Furthermore, diclofenac with metformin demonstrated antiproliferative effects in hamster fibrosarcoma BHK-21/C13, human lung carcinoma A549 (CCL-185), colon carcinoma HT-29 (HTB-38) and cervical carcinoma HeLa (CCL-2) cell cultures, with markedly lower cytotoxicity in the normal fetal lung MRC-5 cells. In conclusion, diclofenac and metformin combination may be recommended for potential use in oncology, due to synergistic anticancer effect in doses achievable in humans.