Improved formulations of antisense oligodeoxynucleotides using wrapped liposomes.

Previously, we demonstrated that wrapping dextran fluorescein anionic/cationic lipid complexes with neutral lipids produced a stable formulation that markedly increased the duration of the compound in plasma after intravenous administration to rats. The improved drug-delivery properties of the wrapped liposomes (WL) relative to other formulations suggested that this technology could offer important advantages for the administration of other polyanionic drugs, including antisense oligodeoxynucleotides (ODN). In the present study, we investigated the value of WL for formulating fluorescence-labeled phosphorothioated ODN (F-ODN). WL encapsulating F-ODN/cationic lipid complexes were prepared efficiently using similar methodology to that used in our earlier study. Studies confirmed that these WL were stable in vitro. Following intravenous administration to mice, free F-ODN and naked F-ODN/cationic lipid complexes were rapidly eliminated whereas administration of the WL resulted in high blood concentrations of drug that were maintained for several hours. Additional studies were conducted in mice that were inoculated with tumor cells (Caki-1 xenograft model, human kidney); in these experiments, intravenous administration of WL delivered 13 times more F-ODN to the tumor site than achieved after injection of free F-ODN.     

Severe Graves' ophthalmopathy accompanied by myasthenia gravis]

We report a 53-year-old woman with severe Graves' ophthalmopathy accompanied by uncontrolled myasthenia gravis. She presented remarkable exophthalmos, chemosis, and restriction of eye movement. Despite plasma exchange, steroid pulse therapy, local injection of steroid, and irradiation, ocular symptoms did not ameliorate. Since optic neuropathy was seen, orbital decompression surgery was performed in the left eye. Bilateral chemosis was improved after the surgery. Five years after surgery, there was no ocular palsy in the operated left eye, but in the contralateral eye. For the good prognosis of the eye movement, orbital decompression might be recommended in the severe Graves' ophthalmopathy accompanied by the optic neuropathy and/or ophthalmoplegia with proptosis.     

Antitumor effect of KT6124, a novel derivative of protein kinase inhibitor K-252a and its mechanism of action

Summary Novel derivatives of K-252a, (8R*,9S*,11S*)-(–)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8, 11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo [a,g]-cycloocta[cde]trinden-1-one, an inhibitor of protein kinases and calmodulin-dependent phosphodiesterase, were synthesized and evaluated for their antitumor activity in vitro and in vivo. Of ten derivatives tested, four were active against the P388 murine leukemia i. p.-i. p. system, although K-252a was inactive. Among these derivatives, KT6124 was selected for further biological evaluation studies because its efficacy was the highest. KT6124 was also active against sarcoma 180 and B16 melanoma. It exerted a relatively broad spectrum of antiproliferative activity against 20 human tumor cell lines in vitro. To determine the mechanism(s) of action underlying the antitumor activity of KT6124, we tested the drug for inhibition of protein kinases, including Ca²⁺-and phospholipid-dependent protein kinase (PKC), in intact A431 human epidermoid carcinoma cells in comparison with the PKC-inhibitory activity of K-252a. KT6124 did not antagonize the action of phorbol 12-myristate 13-acetate (PMA) in A431 cells, whereas K-252a did, suggesting that KT6124 may not act on protein kinases in the cells. The interaction of KT6124 with DNA in living cells was examined by the alkaline elution method. KT6124 apparantly exhibited DNA scission both dose-and time-dependently in the target cells. The DNA breakage was dependent on proteinase K treatment, suggesting its possible interaction with DNA-related enzyme(s). These results indicate that KT6124 exerts antitumor activity by acting on DNA or on DNA-related enzyme(s) in tumor cells rather than via the inhibition of protein kinases.     

Immunohistochemical and ultrastructural study on effect of fibroblast growth factor on transformation of fibroblasts to regenerated mesothelial cells

To elucidate the effect of fibroblast growth factor on the phenotypical conversion of fibroblasts to mesothelial cells, both immunohistochemical and ultrastructural examinations were carried out on cultured spheroids that were composed of fibroblasts obtained from the parietal pleura of rats with and without addition of antifibroblast growth factor receptor antibody. In the present study, antifibroblast growth factor receptor antibody was employed to block the effect of the autocrine component of fibroblast growth factor in the culture medium. Phenotypical conversion from fibroblast to mesothelial cells was clearly blocked in the experimental group, to which culture medium had been added with antifibroblast growth factor receptor antibody, whereas the control group, cultured without addition of antifibroblast growth factor receptor antibody, showed phenotypical conversion of fibroblasts that was confirmed by the development of macula adherens, microvilli, and positive expression of cytokeratin. These results indicate the possibility that fibroblast growth factor plays a key role in the process of phenotypic conversion of fibroblasts to regenerated mesothelial cells.     

Group-transfer alternating copolymerization of 2-phenyl-1,3,2-dioxaphosphorinane with trimethylsilyl 2-(acryloyloxy)ethanesulfonate

This paper describes a new class of reaction, group-transfer alternating copolymerization (GTAC), between 2-phenyl-1,3,2-dioxaphosphorinane ( 1 ) and trimethylsilyl 2-(acryloyloxy)ethanesulfonate ( 2 ). The copolymerization took place without any added catalyst and proceeded via quantitative trimethylsilyl group-transfer process to give a 1:1 alternating copolymer 3a having a ketene silyl acetal group in the main chain. Results of spectroscopy as well as alkaline hydrolysis and bromination of the copolymer confirmed the structure.     

Comparison of histopathological characteristics of allograft biopsy between responder and non-responder to antiproteinuric effect of angiotensin-converting enzyme inhibitor (ACEI)

Abstract:  Angiotensin-converting enzyme inhibitor (ACEI) has become recognized as agents that have renoprotective effects in the treatment of progressive renal diseases including post-transplant kidneys. Previously we demonstrated the safety and effectiveness of ACEI treatment on the hypertensive proteinuric post-transplant patients ( N  = 10) who had been followed up for 12 months. However, not all patients show good response in urinary protein reduction. We aimed to analyse the histopathological factor(s) affecting the responsiveness of proteinuria to ACEI treatment. Fourteen post-transplant patients with proteinuria who were treated with ACEI and underwent allograft biopsy were analysed. Eight patients showed 50% or more reduction in proteinuria (responder). The other 6 patients showed less (< 50%) reduction in proteinuria (non-responder). There was no difference in clinical characteristics (BP, renal function, donor age, recipient body mass index), dietary sodium or protein intake, and diuretic use between the two groups. As a histopathological characteristic, glomerular size in responder group was significantly larger than that in non-responder group. This suggests that the large glomerular size at least partly contributes to the responsiveness in urinary protein reduction to ACEI treatment in kidney allograft recipients with proteinuria.     

TECHNIQUES AND PITFALLS OF ENDOSCOPIC SUBMUCOSAL DISSECTION FOR COLORECTAL TUMORS

Endoscopic submucosal dissection (ESD) for colorectal tumors is steadily being developed. Safety and standardization of ESD for colorectal tumors have not been yet established because of the technical difficulties and the unsuitable anatomical characteristics of the colon and rectum. The authors mainly use a Flex knife for mucosal incision and a Hook knife for submucosal dissection to perform ESD safely. Skillful colonoscopic control, selection of scope, distal attachment tip hood, adequate high‐frequency generator and correct approach strategy should all be considered for safe performance of ESD. However, the incidence of indicative lesions is rare because the majority of colorectal tumors are adenomatous large laterally spreading tumors, which can be cured by intentional endoscopic piecemeal resection. At present, ESD for colorectal tumors should be performed only at central facilities that have expert colonoscopists. With the development of new devices and associated techniques, technical standardization of ESD for colorectal tumors is expected in the near future.     

High human IgG levels in severe combined immunodeficient mouse reconstituted with human splenic tissues from patients with gastric cancer.

We implanted normal peripheral blood lymphocytes (PBL) from healthy donors and splenic tissues from patients with gastric cancers into the severe combined immunodeficient (SCID) mouse, demonstrating that SCID mouse with splenic tissue can produce a high level of human immunoglobulin G (IgG). The normal PBLs at 10(7) and 10(8)/mouse were implanted intraperitoneally, and three splenic tissues with a size of 3 x 3 x 3 mm from gastric cancer patients were inoculated subcutaneously into the bilateral backs of the mice. At 2, 4, 6 and 8 weeks after inoculation, mice were killed, and the human IgG was assessed by an ELISA method. SCID mice with splenic tissue revealed high human IgG levels from 2 weeks after inoculation and approximately 2 mg of IgG per ml was observed at 8 weeks post-implantation, while the IgG levels in mice treated with PBLs were limited. Since the half life of the extrinsic human IgG was 10.2 days, the high level of human IgG in the SCID mice was supposed to be produced by human plasma cells in the splenic tissue from gastric cancer patients. This model was thought to be adequate for evaluating human immunological functions in vivo.