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Here we demonstrate the ability of mannosylated liposomes (ML) targeted to mannose receptors (MR) to perform the targeted delivery of model plasmid DNA encoding EGFP and total tumour RNA into murine bone-marrow-derived dendritic cells (DCs) and enhance the efficiency of anti-tumour response triggered by these DCs in murine melanoma model. ML consist of cationic lipid 2X3 (1,26-Bis(cholest-5-en-3β-yloxycarbonylamino)-7,11,16,20-tetraazahexacosan tetrahydrochloride), helper lipid DOPE (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine), and 2.5, 5 or 10% mol. of novel mannosylated lipoconjugates. In the structure of lipoconjugates D-mannose was attached to ditetradecylglycerol residue via succinyl (lipoconjugate 1) or diethylsquarate (lipoconjugate 2) linker groups. ML spontaneously form complexes with plasmid DNA and RNA due to electrostatic interaction between positively charged lipid amino group and negatively charged phosphate of nucleic acids. ML demonstrated the benefit in transfection efficiency (TE) of pDNA into DC progenitors and immature DCs in comparison with the control liposomes at low N/P (nitrogen to phosphate) ratios (1/1 and 2/1) but not at high N/P ratios where the TE was comparable with control liposomes. Moreover, ML at low N/P were more effective in RNA delivery into immature DCs in comparison with DC progenitors. At high N/P ratios liposomal formulations containing 5 and 10% mol. of mannosylated lipoconjugate 2 with diethylsquarate linker were the most effective (up to 50% of transfected cells). DCs transfected ex vivo with ML/melanoma B16 RNA complexes after i.v. injection into mice caused five- to six-fold inhibition of melanoma lung metastasis number. Moreover, the i.v. injection of ML/melanoma B16 RNA complexes into mice induced generation of the melanoma B16-specific cytotoxic T-lymphocytes, which were two-fold more efficient in B16 cell killing than those from control liposome group., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Recent data on the application of dendritic cells (DCs) as anti-tumor vaccines has shown their great potential in therapy and prophylaxis of cancer. Here we report on a comparison of two treatment schemes with DCs that display the models of prophylactic and therapeutic vaccination using three different experimental tumor models: namely, Krebs-2 adenocarcinoma (primary tumor), melanoma (B16, metastatic tumor without a primary node) and Lewis lung carcinoma (LLC, metastatic tumor with a primary node). Dendritic cells generated from bone marrow-derived DC precursors and loaded with lysate of tumor cells or transfected with the complexes of total tumor RNA with cationic liposomes were used for vaccination. Lipofectamine 2000 and liposomes consisting of helper lipid DOPE (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine) and cationic lipid 2D3 (1,26-Bis(1,2-de-O-tetradecyl-rac-glycerol)-7,11,16,20-tetraazahexacosan tetrahydrocloride) were used for RNA transfection. It was shown that DCs loaded with tumor lysate were ineffective in contrast to tumor-derived RNA. Therapeutic vaccination with DCs loaded by lipoplexes RNA/Lipofectamine 2000 was the most efficient for treatment of non-metastatic Krebs-2, where a 1.9-fold tumor growth retardation was observed. Single prophylactic vaccination with DCs loaded by lipoplexes RNA/2D3 was the most efficient to treat highly aggressive metastatic tumors LLC and B16, where 4.7- and 10-fold suppression of the number of lung metastases was observed, respectively. Antimetastatic effect of single prophylactic DC vaccination in metastatic melanoma model was accompanied by the reductions in the levels of Th2-specific cytokines however the change of the levels of Th1/Th2/Th17 master regulators was not found. Failure of double prophylactic vaccination is explained by Th17-response polarization associated with autoimmune and pro-inflammatory reactions. In the case of therapeutic DC vaccine the polarization of Th1-response was found nevertheless the antimetastatic effect was less effective in comparison with prophylactic DC vaccine.

Exogenous ribonucleases are known to inhibit tumor growth via apoptosis induction in tumor cells, allowing to consider them as promising anticancer drugs for clinical application. In this work the antitumor potential of binase was evaluated in vivo and the mechanism of cytotoxic effect of binase on tumor cells was comprehensively studied in vitro. We investigated tumoricidal activity of binase using three murine tumor models of Lewis lung carcinoma (LLC), lymphosarcoma RLS 40 and melanoma B-16. We show for the first time that intraperitoneal injection of binase at a dose range 0.1-5 mg/kg results in retardation of primary tumor growth up to 45% in LLC and RLS 40 and inhibits metastasis up to 50% in LLC and RLS 40 and up to 70% in B-16 melanoma. Binase does not exhibit overall toxic effect and displays a general systemic and immunomodulatory effects. Treatment of RLS 40-bearing animals with binase together with polychemotherapy revealed that binase decreases the hepatotoxicity of polychemotherapy while maintaining its antitumor effect. It was demonstrated that the cytotoxic effect of binase is realized via the induction of the intrinsic and extrinsic apoptotic pathways. Activation of intrinsic apoptotic pathway is manifested by a drop of mitochondrial potential, increase in calcium concentration and inhibition of respiratory activity. Subsequent synthesis of TNF-α in the cells under the action of binase triggers extrinsic apoptotic pathway through the binding of TNF with cell-death receptors and activation of caspase 8. Thus binase is a potential anticancer therapeutics inducing apoptosis in cancer cells.