Hideaki Ichihara

Chemotherapy with hybrid liposomes for human breast tumors along with apoptosis in vivo

Hybrid liposomes (HL-n) can be prepared by dissolving both vesicular and micellar molecules in buffer solution with ultrasonication. A clear solution of HL-n composed of 95 mol% dimyristoylphosphatidylcholine (DMPC) and 5 mol% polyoxyethylenedodecyl ether (C(12)(EO)(n), n=21 and 25) having hydrodynamic diameter of 100 nm could be kept over 1 month on the basis of dynamic light scattering measurements was obtained. (1) Increases of fusion and accumulation of HL-n including NBDPC as a fluorescence probe into human breast tumor (MDA-MB-453) cell membranes were observed. (2) Reduction of mitochondrial membrane potential and activation of caspase-8, caspase-9, and caspase-3 were observed, indicating that apoptotic signal by HL-n should pass through mitochondria and these caspases. (3) Remarkable reduction of tumor volume in a xenograft mice model intravenously treated with HL-n without drugs after the subcutaneously inoculation of MDA-MB-453 cells was verified in vivo. Induction of apoptosis in tumor of xenograft mice treated with HL-n was observed in micrographs on the basis of TUNEL method. It was noteworthy that the therapeutic effects of HL-n along with apoptosis were obtained for xenograft mice model of human breast tumor in vivo.

Intravenous injection of hybrid liposomes suppresses the liver metastases in xenograft mouse models of colorectal cancer in vivo.

Therapeutic effects of hybrid liposomes (HL) composed of l-α-dimyristoylphosphatidylcholine (DMPC) and polyoxyethylene(25) dodecyl ether (C(12)(EO)(25)) on the metastasis of human colon carcinoma (HCT116) cells were examined in vivo. Remarkably high therapeutic effects were obtained in the xenograft mouse models of colorectal cancer (CRC) liver metastases after treatment with HL-25 on the basis of relative liver weight and histological analysis of the liver tissue sections of mouse models with HE staining, and TUNEL staining for detection of apoptotic cells. The survival effects of HL-25 were obtained using xenograft mouse models of CRC liver metastases. Furthermore, with regard to pharmacokinetics, the accumulation of fluorescent labeled HL-25 was observed in the liver tissue of xenograft mouse models of CRC liver metastases for 24 h after the intravenous injection of fluorescent labeled HL-25. Therapeutic effects of HL without any drugs on the liver metastasis of human CRC were revealed for the first time in vivo.

Chemotherapy with hybrid liposomes for acute lymphatic leukemia leading to apoptosis in vivo

Hybrid liposomes (HL) composed of 95 mol% l-α-dimyristoylphosphatidylcholine (DMPC) and 5 mol% polyoxyethylene (25) dodecyl ether (C(12)(EO)(25)) were prepared by sonication. A clear solutions of HL-25 having hydrodynamic diameter of about 50 nm could be maintained over 3 weeks. Remarkable reduction of tumor volume in model mice of acute lymphatic leukemia (ALL) intravenously treated with HL-25 without drugs after the subcutaneously inoculation of human ALL (MOLT-4) cells was verified in vivo. Induction of apoptosis in tumor of model mice of ALL treated with HL-25 was observed in micrographs on the basis of TUNEL method. Remarkable decrease of the ascites in ALL model mice treated with HL-25 was observed. It is noteworthy that prolonged survival (>400%) was obtained in model mice of ALL after the treatment with HL-25 without drugs.

Membrane targeted chemotherapy with hybrid liposomes for tumor cells leading to apoptosis.

We have produced hybrid liposomes (HL) which can be prepared by sonication of a mixture of vesicular and micellar molecules in a buffer solution. The physical properties of HL such as size, shape, and membrane fluidity can be controlled by changing the constituents and compositional ratio. We have employed HL for chemotherapy and interesting results are as follows; (A) The uniform and stable structure of HL composed of L-α-dimyristoylphosphatidylcholine (DMPC) and polyoxyethylenedodecyl ether (C₁₂(EO)(n)) with a diameter of 80 nm was revealed. (B) The remarkable inhibitory effects of HL on the growth of various tumor cells were attained in vitro

Negatively Charged Cell Membranes-Targeted Highly Selective Chemotherapy with Cationic Hybrid Liposomes against Colorectal Cancer In Vitro and In Vivo

Cationic hybrid liposomes (CHL) composed of 87 mol% L-α-dimyristoylphosphatidylcholine (DMPC), 5 mol% polyoxyethylene (21) dodecyl ether (C12(EO)21) and 8 mol% O,O’-ditetradecanoyl-N-(α–trimethylammonioacetyl) diethanolamine chloride (2C14ECl) were prepared by the method of sonication. A clear solution of CHL having a hydrodynamic diameter of 100 nm could be maintained over 4 weeks. The IC50 value of CHL on the growth of human colorectal cancer (CRC; HCT116) cells was remarkably smaller than that of the DMPC liposomes. Immediate fusion of CHL including NBDPC into HCT116 cell membranes was confirmed using a confocal laser and total internal reflection fluorescence (TIRF) microscope without affecting normal colon (CCD-33Co) cells. Activation of caspases for apoptosis of HCT116 cells induced by CHL was verified on the basis of fluorescence microscopy. Carcinoma HCT116 cells have lower membrane potential compared to normal CCD-33Co cells, since negatively charged PS and GM1 in HCT116 cells increased compared with that of normal CCD-33Co. Therapeutic effects of CHL were obtained in xenograft mouse models of CRC in vivo. Induction of apoptosis in tumor of xenograft mouse model of human CRC treated with CHL was observed in micrographs of tissue section on the basis of TUNEL method. Furthermore, no severe side effects of CHL were observed in toxicity tests using normal mice.

Histological bioanalysis for therapeutic effects of hybrid liposomes on the hepatic metastasis of colon carcinoma in vivo

Therapeutic effects of hybrid liposomes (HL) composed of l-alpha-dimyristoylphosphatidylcholine (DMPC) and polyoxyethylene (23) dodecylether (C(12)(EO)(23)) on the metastasis of colon carcinoma (Colon26) cells were examined in vivo. Fluorescent labeled Colon26 cells were observed in the liver tissue of hepatic metastasis mouse models after the intrasplenic inoculation of the cells. Remarkably high therapeutic effects were obtained in the hepatic metastasis mouse models after the treatment with HL on the basis of relative liver weight and histological analysis of the liver tissue sections of mouse models with hematoxylin-eosin staining, Masson trichrome staining, and CEA immunostaining as a histochemical marker of metastatic colon carcinoma. Furthermore, no toxicity was observed in the hepatic metastasis mouse models after the intravenous injection of HL. Therapeutic effects of HL without any drugs on the hepatic metastasis were revealed on the basis of histological analysis for the first time in vivo.

Hybrid liposomes inhibit tumor growth and lung metastasis of murine osteosarcoma cells.

Antitumor effects of hybrid liposomes (HL) composed of l‐α‐dimyristoylphosphatidylcholine (DMPC) and polyoxyethylene(23) dodecyl ether (C12(EO)23) on the metastatic growth of murine osteosarcoma (LM8) cells were investigated in vitro and in vivo. Remarkable inhibitory effects of HL‐23 on the growth of LM8 cells were obtained through the induction of apoptotic cell death in vitro. It was also indicated that HL‐23 should dramatically suppress the invasion of LM8 cells and the formation of filopodia on the cell surface in vitro. Furthermore, significantly high therapeutic effects were observed in the homograft mouse models of LM8 cells with lung metastasis after the treatment with HL‐23 in vivo. That is, the histological analysis demonstrated that the primary tumor growth of LM8 cells implanted subcutaneously into the mice was inhibited along with the induction of apoptosis. In addition, it was found that HL‐23 significantly decreased the lung metastasis of LM8 cells in the mouse models through the inhibition of primary tumor invasion. These results suggest that HL‐23 could be a novel agent for the chemotherapy of osteosarcoma.

Hybrid Liposomes inhibit the Growth and Angiogenesis in Human Breast Cancer Model

Therapeutic effects of hybrid liposomes (HL-25) composed of L-α-dimyristoylphosphatidylcholine (DMPC) and polyoxyethylene(25)dodecyl ether (C12(EO)25) against breast tumor due to anti-angiogenic activity were examined in vitro and in vivo. Inhibitory effects of HL-25 on the formation of capillary tubes in the human umbilical vascular endothelial cells (HUVEC) were obtained in vitro. Remarkable reduction of tumor volume in mouse models of human breast cancer (HBC) was verified after the intravenous treatment with HL-25 without drugs in vivo. Anti-angiogenic activity in mouse models of HBC treated with HL-25 was observed on the basis of immunostaining method using CD34. Therapeutic effects along with anti-angiogenic activity of HL-25 without any drugs on the mouse models of HBC were revealed for the first time in vivo.

Therapeutic effects of hybrid liposomes with downregulation of inflammatory cytokine for model mice of rheumatoid arthritis in vivo

Therapeutic effects of HL for a collagen-induced arthritis (CIA) mouse models of HL-23 composed of 95mol% l-α-dimyristoylphosphatidylcholine (DMPC) and 5mol% polyoxyethylenedodecylether (C12(EO)23) in vivo were examined. Remarkably high therapeutic effects of HL-23 for CIA mouse models were obtained on the basis of clinical assessment of arthritis. The reduction of hyperplastic synovial membrane (pannus tissue) and destruction of the cartilage and bone by HL-23 was revealed on the basis of hematoxylin and eosin (HE) and safranin O staining. Furthermore, the downregulation of inflammatory cytokines such as interleukin (IL)-1β, tumor necrosis factor (TNF)-α, and IL-6 for CIA mouse models treated with HL-23 were investigated. Remarkably high therapeutic effects without joint swelling were obtained in CIA mouse models treated with HL-23.

Therapeutic effects of hybrid liposomes without drugs for rheumatoid arthritis

Abstract Hybrid liposomes (HLs) can be prepared by simply sonicating a mixture of vesicular and micellar molecules in buffer solutions. This study aims to demonstrate inhibitory effects of HLs on the growth of fibroblast-like synoviocytes along with apoptosis and therapeutic effects of HLs in a mouse model with rheumatoid arthritis (RA). HLs composed of 95 mol% l-α-dimyristoylphosphatidylcholine (DMPC) and 5 mol% polyoxyethylene(23)dodecyl ether (C12(EO)23) were prepared by the sonication method. The inhibitory effects of HLs on the growth of human fibroblast-like synoviocytes-RA (HFLS-RA) cells in vitro and their inhibitory mechanism were examined. High inhibitory effects of HLs on the growth of HFLS-RA cells were observed. The induction of apoptosis by HLs was revealed on the basis of flow cytometric analysis. Furthermore, therapeutic effects of HLs in the mouse model with RA were examined in vivo. Our results demonstrate that HLs showed inhibitory effects on the growth of HFLS-RA cells in vitro along with apoptosis and therapeutic effects in mouse models of RA in vivo.