α-Phellandrene exhibits antinociceptive and tumor-reducing effects in a mouse model of oncologic pain.

Medical reports indicate a prevalence of pain in 50% of patients with cancer. In this context, this article investigated the antinociceptive activity of α-PHE using in vivo Sarcoma-180-induced hypernociception in mice to detail its mechanism(s) of antinociception under different conditions of treatment and tumor progression. Firsty, in vitro cytotoxic action was assessed using melanoma B-16/F-10 and S-180 murine cells and colorimetric MTT assays. For in vivo studies, acute treatment with α-PHE (6.25, 12.5, 25 and 50 mg/kg orally by gavage) was performed on the 1st day after S-180 inoculation. Subacute treatments were performed for 8 days starting on the next day (early protocol) or on day 8 after S-180 inoculation (late protocol). For all procedures, mechanical nociceptive evaluations were carried out by von Frey's technique in the subaxillary region peritumoral tissue (direct nociception) and in right legs of S-180-bearing mice (indirect nociception). α-PHE showed in vitro cytotoxic action on B-16/F-10 and S-180 (CI50 values of 436.0 and 217.9 µg/mL), inhibition of in vivo tumor growth (ranging from 47.3 to 82.7%) and decreased direct (peritumoral tissue in subaxillary region) and indirect (right leg) mechanical nociception in Sarcoma 180-bearing mice with early and advanced tumors under acute or subacute conditions of treatment especially at doses of 25 and 50 mg/kg. It improved serum levels of GSH as well as diminished systemic lipid peroxidation, blood cytokines (interleukin-1ß, -4, -6, and tumor necrosis factor-α). Such outcomes highlight α-PHE as a promising lead compound that combines antinociceptive and antineoplasic properties. Its structural simplicity make it a cost-effective alternative, justifying further mechanistic investigations and the development of pharmaceutical formulations. Moreover, the protocols developed and standardized here make it possible to use Sarcoma-180 hypernociception model to evaluate the capacity of new antinociceptive molecules under conditions of cancer-related allodynia.

Anti-tumor activity and immunogenicity of a succinoglycan riclin.

Natural polysaccharides have attracted considerable interests due to diverse biological activities. Succinoglycan is an extracellular polysaccharide produced by most Agrobacterium strains. Here, we confirmed riclin was a typical succinoglycan by NMR and methylation analysis, and investigated the antitumor effects of riclin in sarcoma 180 tumor-bearing mice. The results showed that riclin inhibited the tumor growth significantly as well as cyclophosphamide (CTX). While CTX caused serious damage to spleen structure, riclin increased the spleen index and promoted lymphocytes proliferation in peripheral blood, spleen and lymph nodes. Riclin decreased splenocytes apoptosis as evidenced by alterations of B-cell lymphoma-2 family proteins and Cleaved Caspase-3 protein. Moreover, 1H nuclear magnetic resonance (NMR)-based metabolomics analysis revealed that riclin partially altered the metabolic profiles of splenocytes. In conclusion, riclin is a succinoglycan that performed strong immunogenicity and suppressed sarcoma growth in mice. Succinoglycan riclin could be a potential antitumor agent for functional food and pharmaceutical purpose.

The Effects of the Alkaloid Tambjamine J on Mice Implanted with Sarcoma 180 Tumor Cells.

The tambjamines are a small group of bipyrrolic alkaloids that, collectively, display a significant range of biological activities including antitumor, antimicrobial and immunosuppressive properties. The key objective of the present study was to undertake preclinical assessments of tambjamine J (T-J) so as to determine its in vivo antitumor effects. To that end, sarcoma 180 cells were transplanted in mice and the impacts of the title compound then evaluated using a range of protocols including hematological, biochemical, histopathological, genotoxic and clastogenic assays. As a result it was established that this alkaloid has a significant therapeutic window and effectively reduces tumor growth (by 40 % and 79 % at doses of 10 and 20 mg/kg/day, respectively). In this regard it displays similar antitumor activity to the anticancer agent cyclophosphamide and alters animal weight in an analogous manner.

In vitro and in vivo antitumoral activity of a ternary copper (II) complex.

Recently, a high number of copper derivatives has been evaluated as DNA-targeting metallodrugs, due to the lower toxicity and its potential to cleave DNA. Several strategies have been testing to develop metal compounds effective against tumour cells. In this work, the ternary copper (doxycycline)-(1,10-phenanthroline) complex [Cu(dox)(phen)]2+ was especially designed as an antitumoral drug, previously showing high cytotoxicity and DNA cleavage activity. We aimed to further investigate the in vitro cytotoxic activity in both tumoral and non-tumoral cells, in vitro genotoxic potential, and in vivo antitumor activity using BALB/C mouse injected with sarcoma S180 and Ehrlich cell lines. Our results indicated that this compound exhibits a moderate genotoxic potential, with selective growth inhibition of tumor cells, especially the murine melanoma B16F10. Its main mechanism of action seems to be through ROS generation. We have further shown a significant reduction of the implanted tumor size in the animal model, suggesting that this compound has great antitumoral potential against many tumor types. [Cu(dox)(phen)]2+ is selectively cytotoxic for melanoma B16F10 and showed high chemotherapeutic potential in vivo against implanted sarcoma S180 and Ehrlich ascites tumours.

Antinociceptive activity of Schinus terebinthifolia leaf lectin (SteLL) in sarcoma 180-bearing mice.

ETHNOPHARMACOLOGY RELEVANCE: Schinus terebinthifolia Raddi leaves have been used in folk medicine due to several properties, including antitumor and analgesic effects. The variable efficacy and adverse effects of analgesic drugs have motivated the search for novel antinociceptive agents. It has been reported that the S. terebinthifolia leaf lectin (SteLL) has antitumor activity against sarcoma 180 in mice. AIM OF THE STUDY: This work aimed to evaluate whether SteLL would reduce cancer pain using an orthotopic tumor model. MATERIALS AND METHODS: A sarcoma 180 cell suspension was inoculated into the right hind paws of mice, and the treatments (150 mM NaCl, negative control; 10 mg/kg morphine, positive control; or SteLL at 1 and 2 mg/kg) were administered intraperitoneally 24 h after cell inoculation up to 14 days. Spontaneous nociception, mechanical hyperalgesia, and hot-plate tests were performed. Further, the volume and weight of the tumor-bearing paws were measured. RESULTS: SteLL (2 mg/kg) improved limb use during ambulation. The lectin (1 and 2 mg/kg) also inhibited mechanical hyperalgesia and increased the latency time during the hot-plate test. Naloxone was found to reverse this effect, indicating the involvement of opioid receptors. The tumor-bearing paws of mice treated with SteLL exhibited lower volume and weight. CONCLUSION: SteLL reduced hyperalgesia due to sarcoma 180 in the paws of mice, and this effect can be related to its antitumor action.

PGE2-HIF1α reciprocal induction regulates migration, phenotypic alteration and immunosuppressive capacity of macrophages in tumor microenvironment.

BACKGROUND: Very little is known about the role inflammation and mechanism(s) that enables the tumor to evade host's anti-tumor immune function during very initial days of tumor establishment. Our study focuses on the immune response and local inflammation specially the pro-inflammatory and immune modifier components that are responsible for tumor-induced immune-suppression, tumor-associated macrophages (TAM) at tumor microenvironment in mouse model from very early to late phase of tumor progression. METHODS: 1 × 105 Ascites tumor, EAC in Swiss albino or Sarcoma-180 (S-180) in Balb c mice strain were inoculated intra-peritonially and grouped into Control (0 day or no tumor), initial phase (3 day tumor), early (7 Day), Late (14 day) and terminal (21 day tumor) sets. T cell activity, tumor niche macrophage, inflammatory signatures were studied using Confocal microscopy, flowcytometry, ELISA, q-RT PCR and Western blot. RESULTS: We observed increased T cell infiltration at a very early stage of tumorigenesis in the tumor site with elevated percentage of activated/memory T cells. But increased cellular death and functional suppression of tumor site T cells during final stages. We observed increased infiltration of TAMs with skewed M2 phenotype. Increased chemokine receptor expression could be noted on these TAMs. Using HIF-1α inhibitor and prostaglandin receptor antagonists we demonstrated crucial role of these factor in functional alteration in TAMs. HIF-1α inhibition and also by prostaglandin receptor inhibition reduced signature pro-inflammatory gene expression, migration of macrophages and T cell suppression capacity of TAMs. We also demonstrated that PGE2 can induce HIF-1α activation in relatively less hypoxic microenvironment during early stages of tumor. CONCLUSION: Altogether, these findings strongly suggest link between prostaglandin mediated early HIF-1α activation and subsequent hypoxia induced HIF-1α activation that further enhances prostaglandin synthesis driving the recruitment and functional alteration of tumor site macrophages.

Synthesis and Evaluation of Ga-68-Labeled Rhein for Early Assessment of Treatment-Induced Tumor Necrosis.

PURPOSE: This study aimed to synthesize a necrosis-avid agent using rhein as a precursor and labeled with gallium-68 (Ga-68) for positron emission tomography/computed tomography (PET/CT) imaging, to evaluate response to anticancer treatment in a mouse model. PROCEDURES: 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-conjugated rhein was radiolabeled with Ga-68 to formulate [68Ga]DOTA-rhein. The in vitro stability of [68Ga]DOTA-rhein was assessed by radio-HPLC. Necrosis avidity was evaluated in a mouse model of muscle necrosis by microPET/CT imaging, biodistribution study, histochemical staining, and autoradiography studies. Murine tumor models with the subcutaneous implantation of S180 cell lines were generated for the evaluation of therapeutic effect. Tumor necrosis was induced by the treatment of combretastatin A4 disodium phosphate (CA4P), and microPET/CT imaging was performed at 1 h post tracer injection. DNA binding studies were conducted to explore the necrosis avidity mechanism of the tracer. RESULTS: [68Ga]DOTA-rhein exhibited a satisfactory yield, a radiochemical purity over 97 %, and a good serum stability. The uptakes of [68Ga]DOTA-rhein in necrotic muscles and tumors were significantly higher than those in normal muscles and tumors (P < 0.05). The results of autoradiography and histochemical staining were consistent with the selective uptake of the radiotracer in necrotic regions. MicroPET/CT images showed a high uptake of the tracer in necrotic muscles and necrotic tumors. DNA binding studies suggested that necrosis avidity correlated with DNA binding to a certain extent. CONCLUSIONS: Our results demonstrated that [68Ga]DOTA-rhein showed a prominent necrosis avidity and could be a useful probe for early assessment of response to anticancer therapy by PET/CT imaging.

The effect of earthworm extract on mice S180 tumor growth and apoptosis.

Great efforts have been made to explore the potential treatment for cancers, and the most common therapies include surgery, chemotherapy and radiotherapy. As an alternative medication, earthworms have drawn increased attention considering its abundance in resource, easy access and minor side effects compared to traditional therapies. However, few studies had focused on the antitumor effect of earthworm-derived components. The purpose of this study is to investigate whether earthworm extract has an effect on tumor cell apoptosis and growth. Earthworm extract (EE) was purified through multiple steps of centrifugation and chromatography. Mice were inoculated with ascitic fluid derived from mice inoculated with S180 sarcoma tumor cells and fed orally with different amounts of EE for 25 days. Tumor samples were analyzed for size and cell apoptosis. And we found that the weight and sizes of tumor decreased gradually as the amount of EE administered increased. More apoptotic cells and lowered level of lactate dehydrogenase (LDH), a biomarker of tumor invasiveness, was detected in EE-treated group than the untreated group. Our results suggested that EE could dramatically promote tumor apoptosis and reduce tumor size in vivo, suggesting a novel alternative therapy for cancer patients.

Synthesis and Bioevaluation of Novel [18F]FDG-Conjugated 2-Nitroimidazole Derivatives for Tumor Hypoxia Imaging.

Hypoxia imaging can guide tumor treatment and monitor changes in hypoxia during treatment. However, there is still no ideal hypoxia imaging agent for clinical applications. In this study, two novel 2-nitromidazole derivatives were synthesized and directly radiolabeled by [18F]FDG in high radiochemical yield and excellent radiochemical purity. Cell experiments, biodistribution, and positron emission tomography (PET) imaging studies were also conducted in mice-bearing S180 or OS732 tumors. [18F]FDG-2NNC2ON [(2 R,3 S,4 R, E)-2-18F-fluoro-3,4,5,6-tetrahydroxyhexanal O-3-(2-(2-nitro-1 H-imidazole-1-yl)ethylamino)-2-oxopropyl oxime] and [18F]FDG-2NNC5ON [(2 R,3 S,4 R, E)-2-18F-fluoro-3,4,5,6-tetrahydroxyhexanal-O-3-(5-(2-nitro-1 H-imidazole-1-yl)pentylamino)-2-oxopropyl oxime] can be cleared from the blood quickly and specifically target hypoxic tumor cells. The uptake of the probes by hypoxic cells gradually increases with time. After 4 h, the uptake value of [18F]FDG-2NNC2ON in hypoxic cells is 3.2 times higher than that in normoxia cells. In contrast, there is no difference in the uptake of [18F]FDG between hypoxic cells and normoxia cells. Biodistribution resulting from two tumor models indicate that the uptake values of the two radiotracers in the tumor are higher at 1 h than those at 2 and 4 h. At 1 and 2 h, the tumors are clearly observed on the PET images and the imaging features of [18F]FDG-2NNC5ON and [18F]FDG-2NNC2ON are distinct from those of [18F]FDG. Compared with [18F]FDG-2NNC5ON, [18F]FDG-2NNC2ON has a higher proportion of renal excretion, lower digestive tract uptake, and better imaging contrast because of its higher hydrophilicity. At 2 h, [18F]FDG-2NNC2ON shows a good tumor-to-blood (T/B) ratio, tumor-to-muscle ratio based on biodistribution (Bio-T/M ratio), and tumor-to-muscle ratio based on regions of interest on the PET images [region of interest (ROI)-T/M ratio] in the two tumor models (T/B, Bio-T/M, and ROI-T/M ratios are 3.2, 2.6, and 3.9 in the S180 tumor model and are 3.4, 4.2, and 4.6 in the OS732 tumor model, respectively). The imaging features visualized with autoradiography mostly coincided with the positive areas of HIF1α staining by immunofluorescence. Meanwhile, the biodistribution study and PET imaging revealed that the uptake of the radiotracers in the tumor cannot be competed by 5% glucose, confirming that [18F]FDG-2NNC2ON targets the hypoxic regions of the tumors instead of targeting tumors through the glucose metabolism pathway. These results suggest that the new 2-nitroimidazole derivative conjugated with [18F]FDG, [18F]FDG-2NNC2ON, has potential as an imaging agent for hypoxia.

Targeted Delivery of Zoledronate to Tumor-Associated Macrophages for Cancer Immunotherapy.

Tumor-associated macrophages (TAMs) are recruited from circulatory monocytes by tumor-derived factors, which differentiate into macrophages residing in the tumor microenvironment. TAMs play critical roles in promoting angiogenesis, invasion, metastasis and immune escape, and the direct depletion of TAMs is a promising strategy for tumor immunotherapy. In this study, we developed lipid-coated calcium zoledronate nanoparticles (CaZol@pMNPs) containing conjugated mannose, which were sterically shielded with an extracellular pH-sensitive material. The NPs specifically targeted TAMs and induced their apoptosis in vitro and in vivo. In a S180 tumor-bearing mouse model, CaZol@pMNPs effectively depleted TAMs, markedly decreased angiogenesis, reduced immune suppression, and eventually restrained tumor growth without eliciting systemic effects. The collective data indicate the potential of the direct depletion of TAMs using CaZol@pMNPs for cancer immunotherapy.

Action of copper(II) complex with ß-diketone and 1,10-phenanthroline (CBP-01) on sarcoma cells and biological effects under cell death.

This work reports the biological evaluation of a copper complex of the type [Cu(O-O)(N-N)ClO4], in which O-O = 4,4,4-trifluoro-1-phenyl-1,3-butanedione (Hbta) and N-N = 1,10-phenanthroline (phen), whose generic name is CBP-01. The cytotoxic effect of CBP-01 was evaluated by resazurin assay and cell proliferation was determined by MTT assay. DNA fragmentation was analyzed by gel electrophoresis. Cell cycle progression was detected through propidium iodide (PI) staining. Apoptosis and autophagy were determined by, respectively, Annexin V and 7-AAD staining and monodansylcadaverine (MDC) staining. The changes in intracellular reactive oxygen species levels were detected by DCFDA analysis. The copper complex CBP-01 showed in vitro antitumor activity with IC50s values of 7.4 µM against Sarcoma 180 and 26.4 against murine myoblast cells, displaying selectivity toward the tumor cell tested in vitro (SI > 3). An increase in reactive oxygen species (ROS) generation was observed, which may be related to the action mechanism of the complex. The complex CBP-01 may induce DNA damage leading cells to accumulate at G0/G1 checkpoint where, apparently, cells that are not able to recover from the damage are driven to cell death. Evidence has shown that cell death is initiated by autophagy dysfunction, culminating in apoptosis induction. The search for new metal-based drugs is focused on overcoming the drawbacks of already used agents such as acquired resistance and non-specificity; thus, the results obtained with CBP-01 show promising effects on cancer cells.

Antitumor activity of Se-containing tea polysaccharides against sarcoma 180 and comparison with regular tea polysaccharides and Se-yeast.

Selenium (Se)-containing polysaccharide, a Se-conjugate macromolecule, generally exhibited higher antitumor activity than its regular polysaccharide. Previously, we extracted Se-containing tea polysaccharides (Se-TPS) from Se-enriched tea, and explored its structure and antioxidant activity. In this study, we investigated antitumor activity of Se-TPS on sarcoma 180 (S-180), and compared with its regular polysaccharides TPS and dietary supplement Se-yeast. In vitro antitumor activity of Se-TPS was evaluated by MTT and LDH assays, and the results indicated that Se-TPS can significantly inhibit the proliferation of S-180 in dose-dependent manner (R2=0.97, p<0.0001). In S-180 cancer xenograft model in Kunming mice, Se-TPS oral administration at three doses of 50, 100 and 200mg/kg body weight daily for 13days resulted in significant tumor regression. At the same dose, Se-TPS exhibited significantly higher antitumor activity than TPS and Se-yeast. Importantly, Se-TPS can significantly increase the spleen and thymus indices of tumor-bearing mice, suggesting the safety and immunomodulatory activity of Se-TPS. Therefore, Se-TPS may be a desirable antitumor agent for therapeutic and immunomodulatory applications.

Growth-inhibition of S180 residual-tumor by combination of cyclophosphamide and chitosan oligosaccharides in vivo.

Chitosan oligosaccharides (COS), hydrolyzed products of chitosan, have recently been reported to have various biological activities. Herein, the present study was undertaken to assess the ability of COS to potentiate the antitumor effect of cyclophosphamide (CTX) as well as alleviating the CTX-associated toxicities in vivo, in a residual-tumor; a model which is closer to clinical surgery. Sarcoma 180 (S180) residual-tumor mice were divided into 6 groups (n = 6); including control, CTX, COS 40 mg/kg, COS 80 mg/kg, and combination groups (CTX + COS 40, CTX + COS 80). Animals were killed 18 days post-intraperitoneal administration and the tumors were weighed. The spleens were harvested to determine lymphocytes proliferation and NK cell activities; blood cells were evaluated by flow cytometry, and the expression levels of TNF-α were measured using ELISA. Notably, the combined therapy (CTX + COS80) showed the most effective reduction of the tumor weight, the highest inhibition of tumor growth, and proliferation, when compared with control as well single CTX group. Additionally, COS was able to recover the CTX-induced decreases in the lymphocyte proliferation, splenocyte NK cell activity, TNF-α concentration, and abnormal CD4+/CD8+ T lymphocyte subset. The increase in infiltrating T cells and macrophages best explain the immunostimulatory effect of COS. Results herein highlighted the therapeutic potential of COS as adjuvant treatment during tumor chemotherapy.

Effect of lutein and doxorubicin combinatorial therapy on S180 cell proliferation and tumor growth.

OBJECTIVE: Multidrug resistance and toxicity significantly compromise the therapeutic efficacy for sarcomas. We aimed at evaluating the effect of lutein-doxorubicin (DOX) combinatorial therapy on inhibiting S180 (Sarcoma 180) cell proliferation and tumor growth. MATERIALS AND METHODS: S180 cells in logarithmic growth phase were treated with lutein, DOX, or lutein-DOX combinatorial therapy for 48 h. The cell survival rate was determined by MTT assay. Apoptosis was detected by flow cytometry. The expression of PCNA, P53, and NF-κB was assessed by Western blot. Further, mice bearing S180 tumors received lutein, DOX, or lutein-DOX combinatorial therapy by oral gavage. RESULTS: Lutein-DOX combinatorial therapy significantly decreased the proliferation of S180 cells (p<0.01) in vitro. Also, the expression of proliferating cell nuclear antigen (PCNA) (p<0.05) and the apoptosis-relevant gene p53 were decreased, which resulted in increased cell apoptosis (p<0.05). The level of nuclear factor kappa B (NF-κB) was also decreased by the combinatorial therapy. Lutein-DOX combinatorial therapy reduced the cytotoxicity of DOX and reduced the inflammatory response. The inhibitory effect of lutein-DOX combinatorial therapy on cell proliferation was confirmed in vivo. The growth rate and size of the tumor at 30 d after treatment were significantly lower than those of the control group and DOX single therapy. CONCLUSIONS: Lutein and DOX synergistically inhibit sarcoma cell proliferation and tumor growth. This novel therapeutic regimen could potentially improve clinical outcome of sarcoma patients.

Combined use of insoluble ß-glucan from the cell wall of Candida albicans and cyclophosphamide: Validation in S180 tumor-bearing mice.

BACKGROUND: Cyclophosphamide (CTX) is a widely used antitumor drug that can suppress the immune system. The effects of regulating immune response and antitumor of ß-glucan from the cell wall of Candida albicans (CAIBG) have been confirmed. However, the effects of the combined use of CAIBG and CTX remain unclear and warrant further investigation. METHODS: S180 tumor-bearing models were developed for CAIBG (100 mg/10 mL/kg) and CTX (30 mg/10 mL/kg) intervention. The weights of the body, tumor spleen, and Thymus were recorded to calculate the index of the spleen and Thymus. The spleen and Thymus were observed by hematoxylin and eosin staining, whereas the expression of tumor necrosis factor (TNF)-α and interleukin (IL)-1ß was determined by Western blot. The survival times of mice were followed and recorded for analysis. RESULTS: CAIBG, CTX, and combined use of CAIBG and CTX could down-regulate the tumor growth and prolong the survival time. The spleen and Thymus index significantly increased in the CAIBG + CTX group than in the CTX group, but it was lower than that in the CAIBG group. Moreover, the Thymus index was significantly lower in the CAIBG + CTX group than in the CAIBG group. The lymphocytes of the spleen and Thymus decreased significantly in the CTX group but improved significantly in the CAIBG and CAIBG + CTX groups. The expression level of TNF-α and IL-1ß in the CTX+CAIBG group increased significantly compared with that in the CTX group. The survival time of the CAIBG group and CAIBG + CTX group was significantly higher than that of the CTX group. CONCLUSIONS: CAIBG has strong treatment potential in combating tumor growth and prolonging survival time of S180 tumor-bearing mice. Combined use of CAIBG and CTX can compensate the CTX-induced immunosuppression and provide antitumor effects. Future studies are necessary to elucidate the underlying mechanism.

Himatanthus drasticus (Apocynaceae) latex reduces oxidative stress and modulates CD4+, CD8+, FoxP3+ and HSP-60+ expressions in Sarcoma 180-bearing mice.

ETHNOPHARMACOLOGICAL RELEVANCE: In Brazil, latex of Himatanthus drasticus is used to treat inflammation, wound healing and cancer. The present study evaluated the antitumoral potential of H. drasticus latex (HdCL) in Sarcoma 180-bearing mice (S180). MATERIALS AND METHODS: HdCL was obtained in Crato-CE, Brazil. Qualitative phytochemicals assays, nuclear magnetic resonance (NMR) and microbiological analyzes were performed. Swiss mice were divided into six groups, according to tumor forms: 1) ascitic model, GI (Control; 0.9% saline), GII (S180asc) and GIII (S180asc/HdCL/14 days); 2) solid model, GIV (Control; 0.9% saline), GV (S180sol) and GVI (S180sol/HdCL/10 days). HdCL and 0.9% saline were administered at 0.2 mL, SID, by gavage, for 10 or 14 days. For ascitic model, 0.5 mL of S180 suspension (4×106 cells/mL) was inoculated intraperitoneally and for solid model, cells were inoculated subcutaneously (25 µL) on the right hind paw of mice. Blood samples were collected for hematological and oxidative stress evaluation. Thickness, volume and weight of paws were measured in solid model. After euthanasia, spleen, liver and kidney were collected in order to assess the relative organ weight. Tissue fragments of paws and popliteal lymph nodes (PLN) were analyzed by H&E and CD4+, CD8+, HSP-60+ and Foxp3+ immunohistochemistry. RESULTS: HdCL presented milky aspect and pinkish supernatant. Phenols, flavonols, flavanones, free steroids and cinnamoyl derivatives of lupeol, α-amyrin and ß-amyrin were detected at the phytochemistry analysis. HdCL did not alter the relative weight of organs, hematological parameters and volume of ascitic fluid recovered. In solid model, HdCL reduced (P < 0.05) paw volume, but did not altered thickness, paw weight and histological parameters. S180sol induced necrosis, metastasis and destruction of bone, cartilage and muscles. Bleeding, vessel congestion and oncocytes were observed in PLN. In paw, HdCL did not alter FoxP3+ and HSP-60+ expressions but reduced the CD4+ and CD8+ expressions, while at PLN, HdCL reduced the expressions of all markers. HdCL decreased (P < 0.05) serum levels of malondialdehyde in ascitic model. CONCLUSIONS: Treatment with HdCL reduced oxidative damage and modulated the expressions of CD4+, CD8+, FoxP3+and HSP-60+ in S180 solid tumor model, which can be associated to the presence of triterpenes, such as α-amyrin, ß-amyrin and lupeol cinnamate. Present data emphasizes the importance of immune system in cancer and highlights the evaluation of the pharmacological properties of plants used by population as phytoterapics.

Enhanced antitumor efficacy of poly(D,L-lactide-co-glycolide)-based methotrexate-loaded implants on sarcoma 180 tumor-bearing mice.

PURPOSE: Methotrexate is widely used in chemotherapy for a variety of malignancies. However, severe toxicity, poor pharmacokinetics, and narrow safety margin of methotrexate limit its clinical application. The aim of this study was to develop sustained-release methotrexate-loaded implants and evaluate antitumor activity of the implants after intratumoral implantation. MATERIALS AND METHODS: We prepared the implants containing methotrexate, poly(D,L-lactide-co-glycolide), and polyethylene glycol 4000 with the melt-molding technique. The implants were characterized with regards to drug content, morphology, in vitro, and in vivo release profiles. Differential scanning calorimetry (DSC) and Fourier transform infrared spectroscopy (FTIR) were carried out to investigate the physicochemical properties of the implants. Furthermore, the antitumor activity of the implants was tested in a sarcoma 180 mouse model. RESULTS: The implants were prepared as solid rods. Scanning electron microscopy images showed a smooth surface of the implant, suggesting that methotrexate was homogeneously dispersed in the polymeric matrix. The results of DSC and FTIR indicated that no significant interaction between methotrexate and the polymer was observed in the implants. Both in vitro and in vivo release profiles of the implants were characterized by burst release followed by sustained release of methotrexate. Intratumoral implantation of methotrexate-loaded implants could efficiently delay tumor growth. Moreover, an increase in the dose of implants led to a higher tumor suppression rate without additional systemic toxicity. CONCLUSION: These results demonstrate that methotrexate-loaded implants had significant antitumor efficacy in a sarcoma 180 mouse model without dose-limiting side effects, and suggest that the implants could be potentially applied as an intratumoral delivery system to treat cancer.

Surface density of polyarginine influence the size, zeta potential, cellular uptake and tissue distribution of the nanostructured lipid carrier.

Poly-arginines are strong tools to elevate the cellular uptake of nanopreparations. To learn the influence of poly-arginine (RRRRRRRR, R8) density on a series of properties of nanostructured lipid carrier (NLC), we build six R8 modified NLCs with different R8 densities (nR-NLC, where n represents the R8 ratio) by fusion-emulsion method with the aid of stearyl-R8. The pharmaceutical characteristics like size, zeta potential and in vitro drug release, cellular uptake, cytotoxicity to A549 cells and tissue distribution in S180 tumor-bearing mice of the six nR-NLCs are all investigated. It turns out that with as little as 2% weight ratio of stearyl-R8 modified on NLC, its pharmaceutical properties, especially zeta potential changes astonishingly; however, the stearyl-R8 ratio should be higher than 4% to upgrade the cellular uptake and cytotoxicity evidently; in the ex vivo tissue distribution assessment, the nR-NLC with less than 8% R8 showed similar tissue accumulation, while NLC with 10% R8 shows obvious acute toxicity to mice. Our study pays attention to the effect of the R8 ratio on the changes of cargo properties, and the results indicate that this topic is essential and worth to be further developed.

Study on antitumor, antioxidant and immunoregulatory activities of the purified polyphenols from pinecone of Pinus koraiensis on tumor-bearing S180 mice in vivo.

Pinecone polyphenols are bioactive dietary constituents that enhance health and help prevent and treat cancer through improving antioxidant and immunoregulatory activities. This study was designed to investigate the antitumor, antioxidant and immunoregulatory activities of the 40% ethanol eluent of polyphenols from pinecone of pinus koraiensis (PPP-40) in Sarcoma 180 (S180)-bearing mice models in vivo. The results of antitumor activity indicated that PPP-40 significantly inhibited S180 tumor growth and the dose of 150mg/kg exhibited the highest antitumor activity. Moreover, TdT-mediated dUTP nick end labeling (TUNEL) assay results further confirmed the apoptosis of S180 tumor cells. In addition, PPP-40 could obviously promote the expressions of Bax protein and inhibit the Bcl-2 protein, accordingly improve the expressions of activated Caspase-3 as well, which resulted in the activation of mitochondrial apoptotic pathway of tumor cells in S180 mice eventually. The results of antioxidant activity showed that the S180 mice treated with PPP-40 had the higher superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities, the more glutathione (GSH) content, and the lower malondialdehyde (MDA) level in plasma comparing with non-treated control group. Moreover, the administration with PPP-40 (150mg/kg) significantly accelerated the proliferation of splenocytes (p<0.01) and increased the monocyte phagocytosis activity in vivo simultaneously. These results revealed that PPP-40 exerts an effective antitumor activity by activating the mitochondrial apoptotic pathway and improving the antioxidant and immunoregulatory activities.

Loading cisplatin onto 6-mercaptopurine covalently modified MSNS: a nanomedicine strategy to improve the outcome of cisplatin therapy.

In the treatment of cancer patients, cisplatin (CDDP) exhibits serious cardiac and renal toxicities, while classical combinations related to CDDP are unable to solve these problems and may result in worse prognosis. Alternately, this study covalently conjugated 6-mercaptopurine (6MP) onto the surface of mercapto-modified mesoporous silica nanoparticles (MSNS) to form MSNS-6MP and loaded CDDP into the holes on the surface of MSNS-6MP to form MSNS-6MP/CDDP, a tumor-targeting nano-releasing regime for CDDP and 6MP specifically. In the S180 mouse model, the anti-tumor activity and overall survival of MSNS-6MP/CDDP (50 mg·kg-1·day-1, corresponding to 1 mg·kg-1·day-1 of 6MP and 5 mg·kg-1·day-1 of CDDP) were significantly higher than those of CDDP alone (5 mg·kg-1·day-1) or CDDP (5 mg·kg-1·day-1) plus 6MP (1 mg·kg-1·day-1). The assays of serum alanine aminotransferase, aspartate aminotransferase and creatinine, as well as the images of myocardium and kidney histology, support that MSNS-6MP/CDDP is able to completely eliminate liver, kidney and heart toxicities induced by CDDP alone or CDDP plus 6MP.