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.     

Immunohistochemical analysis of TrkA neurotrophin receptor expression in human non-neuronal carcinomas

The Trk family of tyrosine protein kinase receptors plays a significant role in the development and maintenance of neural tissues. It has been recently shown that Trk receptors are also expressed by a wide range of normal non-neuronal tissues in humans in a cell type-specific manner. In the present study, the expression patterns of TrkA in 337 non-neuronal invasive carcinomas of 15 different human tissues were investigated immunohistochemically. Overall, 133 (39%), 101 (30%) and 103 (31%) tumors exhibited strong, moderate and no TrkA Immunoreactivity, respectively. Esophageal and thyroid carcinomas expressed high levels of TrkA, whereas the levels in gastric and colon cancers were low. TrkA expression was detected not only in carcinomas originating from TrkA-positive normal counterpart tissues, Including the esophagus, breast, lung and uterus, but also in those from TrkA-negative tissues/cells of the thyroid, liver and ovary. Immunostaining for nerve growth factor-β, the specific ligand for TrkA, in esophageal and breast carcinomas demonstrated its immunoreactivity in stromal fibroblasts and some TrkA-expressing tumor cells. These results suggest that paracrine/autocrine regulation via stromal/tumoral NGF-tumoral TrkA interaction may be involved In the growth of certain non-neuronal carcinomas.     

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.     

Active production of anti-human T-lymphotropic virus type I (HTLV-I) IgM antibody in HTLV-I-associated myelopathy

We investigated the presence of anti-human T-lymphotropic virus type I (HTLV-I) IgM in sera and cerebrospinal fluid from patients with HTLV-I-associated myelopathy (HAM) by Western blot analysis. Analyses of 36 serum samples revealed that most patients (31/36; 86.1%) had anti-HTLV-I IgM, whereas only four of 23 (17.4%) HTLV-I carriers had it. In studies of cerebrospinal fluid, anti-HTLV-I IgM was detected in 24 of 36 (66.7%) HAM patients, whereas none was detected in nine HTLV-I carriers. The differences were statistically significant (p less than 0.01). These results suggest that persistent active replication of HTLV-I occurs in the central nervous system as well as in the peripheral blood of HAM patients, and may contribute to the development of HAM.     

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.     

Interruption of pulmonary arterial flow with inadequate ventilation leads to pulmonary infarction

We examined the effect of interruption of pulmonary arterial flow and inadequate ventilation on the development of pulmonary infarction in rats. Pulmonary arterial flow was blocked by the injection of agar into the inferior vena cava and inadequate ventilation was produced by obstructing the left main bronchus with a polypropylene tip. Histological and angiographic examination of the lung demonstrated that: pulmonary artery embolism alone does not induce pulmonary infarction; obstruction of a bronchus does not induce significant changes, but that pulmonary infarction develops when pulmonary artery embolism and obstruction of a bronchus occur simultaneously. It has been thought that pulmonary infarction is caused by acute obstruction of a pulmonary artery, however, the alveolar walls are supplied with oxygen by both the pulmonary circulation and by ventilation. Interruption of pulmonary arterial flow alone is probably not sufficient to induce pulmonary infarction, which is probably caused by deficiency of oxygen supply to the alveolar walls by a synergy between interruption of pulmonary arterial flow and inadequate ventilation.     

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.