Anti-myeloma effect of homoharringtonine with concomitant targeting of the myeloma-promoting molecules, Mcl-1, XIAP, and β-catenin

Since a variety of cell intrinsic and extrinsic molecular abnormalities cooperatively promote tumor formation in multiple myeloma (MM), therapeutic approaches that concomitantly target more than one molecule are increasingly attractive. We herein demonstrate the anti-myeloma effect of a cephalotaxus alkaloid, homoharringtonine (HHT), an inhibitor of protein synthesis, through the induction of apoptosis. HHT significantly reduced Mcl-1, a crucial protein involved in myeloma cell survival, in all three myeloma cell lines examined, whereas certain BH3-only proteins, such as Bim, Bik, and Puma, remained unchanged following HHT treatment, and their expression levels depended on the cell type. HHT also reduced the levels of c-FLIP(L/S), activated caspase-8, and induced active truncated-Bid. Thus, HHT-induced apoptosis appears to be mediated via both intrinsic and extrinsic apoptosis pathways, and the resultant imbalance between BH3-only proteins and Mcl-1 may be pivotal for apoptosis by HHT. In addition, HHT treatment resulted in reduced levels of beta-catenin and XIAP proteins, which also contribute to disease progression and resistance to chemotherapy in MM. In combination, HHT enhanced the effects of melphalan, bortezomib, and ABT-737. These results suggest that HHT could constitute an attractive option for MM treatment though its ability to simultaneously target multiple tumor-promoting molecules.     

Targeted deletion of the murine corneodesmosin gene delineates its essential role in skin and hair physiology

Controlled proteolytic degradation of specialized junctional structures, corneodesmosomes, by epidermal proteases is an essential process for physiological desquamation of the skin. Corneodesmosin (CDSN) is an extracellular component of corneodesmosomes and, although considerable debate still exists, genetic studies have suggested that the CDSN gene in the major psoriasis-susceptibility locus (PSORS1) may be responsible for susceptibility to psoriasis, a human skin disorder characterized by excessive growth and aberrant differentiation of keratinocytes. CDSN is also expressed in the inner root sheath of hair follicles, and a heterozygous nonsense mutation of the CDSN gene in humans is associated with scalp-specific hair loss of poorly defined etiology. Here, we have investigated the pathogenetic roles of CDSN loss of function in the development of skin diseases by generating a mouse strain with targeted deletion of the Cdsn gene. Cdsn-deficient mouse skin showed detachment of the stratum corneum from the underlying granular layer and/or detachment within the upper granular layers due to the disrupted integrity of the corneodesmosomes. When grafted onto immunodeficient mice, Cdsn-deficient skin showed rapid hair loss together with epidermal abnormalities resembling psoriasis. These results underscore the essential roles of CDSN in hair physiology and suggest functional relevance of CDSN gene polymorphisms to psoriasis susceptibility.     

HTLV-I associated interstitial pneumonia--a case report and bronchoalveolar lavage study

A 70-year-old male was admitted to our hospital because of fever and dyspnea. The patient was seropositive for HTLV-I and ATL cells were seen in the peripheral blood in the percentage of 2-5. The proviral DNA was positive and the diagnosis of smoldering ATL was made. His chest X-ray film showed diffuse reticulo-nodular infiltrates in both lung fields. The lung tissue obtained by transbronchial lung biopsy showed the lymphocytic infiltrations in the alveolar septa and the submucosa of the bronchioles. Bronchoalveolar lavage (BAL) fluid showed an increased proportion of lymphocytes that consisted mainly of CD3+ DR+ cells and the CD4+/CD8+ ratio was 2.1 during exacerbation and 0.8 after steroid therapy. Anti-HTLV-I IgG and IgA antibodies were positive in both serum and BAL fluid by Western blotting method. It is suggested that T-lymphocyte alveolitis may occur in patients who are seropositive for HTLV-I and the immunological mechanism seems to be responsible.     

Antisense oligonucleotides of hepatoma-derived growth factor (HDGF) suppress the proliferation of hepatoma cells

BACKGROUND/AIMS: Human hepatoma-derived growth factor, purified from the conditioned medium of hepatoma-derived cell line, HuH-7, stimulates the growth of Swiss 3T3 fibroblasts and HuH-7 cells. To evaluate the role of hepatoma-derived growth factor on the growth of hepatoma cells, we investigated the effects of recombinant hepatoma-derived growth factor protein and hepatoma-derived growth factor antisense oligonucleotides on the proliferation of several hepatoma cell lines. METHODOLOGY: We examined the effects of hepatoma-derived growth factor antisense oligonucleotides on the growth of hepatoma cells by cell growth assay. RESULTS: Hepatoma-derived growth factor stimulated the proliferation of some hepatoma cells (HuH-7, HLF, HepG2, AH66tc cells) about 15-70% over than the control. Hepatoma-derived growth factor antisense oligonucleotides, phosphorothioate-linked or encapsulated in liposome, can inhibit the growth of hepatoma cells. The ID50 of hepatoma-derived growth factor antisense phosphorothioate oligonucleotides for HuH-7 cells, in which hepatoma-derived growth factor expression was abundant, was 3 microM by the assay of cell proliferation and [3H]-thymidine incorporation. Their ID50 for AH66tc cells, on which the effects of exogenous hepatoma-derived growth factor were weak, was higher than 10 microM. To omit the toxic effects due to phosphorothioate modification of oligonucleotides and keep the cellular uptake more without their destruction in the culture medium, we used oligonucleotides encapsulated in cationic liposome. Hepatoma-derived growth factor antisense oligonucleotides encapsulated in liposome suppressed the growth of hepatoma cells effectively (ID50:2.0 microM). CONCLUSIONS: These findings suggest that hepatoma-derived growth factor is one of important autocrine, and/or intracrine factors for hepatoma cells, and that hepatoma-derived growth factor anti-sense oligonucleotides may be useful for human hepatocellular carcinoma as an anti-cancer agent.     

Urokinase type plasminogen activator receptor expression in colorectal neoplasms

Background—The urokinase type plasminogenactivator receptor (uPAR) may play a critical role in cancer invasionand metastasis.
Aims—To study the involvement of uPAR incolorectal carcinogenesis.
Methods—The cellular expression and localisationof uPAR were investigated in colorectal adenomas and invasivecarcinomas by in situ hybridisation, immunohistochemistry, and northernand western blot analyses.
Results—uPAR mRNA expression was found mainly inthe cytoplasm of dysplastic epithelial cells of 30% of adenomas withmild (19%), moderate (21%), and severe (47%) dysplasia, and in that of carcinomatous cells of 85% of invasive carcinomas: Dukes' stages A(72%), B (93%), and C (91%). Some stromal cells in the adjacent neoplastic epithelium were faintly positive. Immunoreactivity for uPARwas detected in dysplastic epithelial cells of 14% of adenomas and incarcinomatous cells of 49% of invasive carcinomas. uPAR mRNA andprotein concentrations were significantly higher in severe than in mildor moderate dysplasia (p<0.05); they were notably higher in Dukes'stage A than in severe dysplasia (p<0.05), and significantly higher inDukes' stage B than in stage A (p<0.05), but those in stage B werenot different from those in stage C or in metastatic colorectalcarcinomas of the liver.
Conclusions—Colorectal adenoma uPAR, expressedessentially in dysplastic epithelial cells, was upregulated withincreasing severity of atypia, and increased notably during thecritical transition from severe dysplasic adenoma to invasivecarcinoma. These findings implicate uPAR expression in the invasive andmetastatic processes of colorectal cancer.

Keywords:urokinase type plasminogen activator receptor; colorectal adenoma; colorectal cancer; adenoma-carcinoma sequence

     

Inhibition by protein kinase-C inhibitor and cycloheximide of phorbol ester- and epidermal growth factor-induced arachidonic acid metabolism in cultured porcine thyroid cells

In an attempt to elucidate possible mechanism(s) for stimulated arachidonic acid metabolism by phorbol 12-myristate 13-acetate (PMA) and epidermal growth factor (EGF) in porcine thyroid cells, we examined the effects of protein kinase inhibitors, isoquinolinesulfonamide derivatives (H-7 and HA-1004), and cycloheximide. The production of PGE2 stimulated by either PMA or EGF was strongly inhibited by H-7, with an ID50 value of approximately 20 to 25 mumol/L in each case, as well as by cycloheximide, with an ID50 value of less than 0.5 micrograms/mL in each case. In contrast, 100 mumol/L of HA-1004 showed less inhibition of PGE2 production provocated by either PMA or EGF. On the other hand, PGE2 production in basal or stimulated condition by exogenously added arachidonic acid, was inhibited to an even lesser extent by both H-7 and cycloheximide. The EGF- and PMA-stimulated release of 3H-arachidonic acid from the cells was also strongly inhibited by H-7 and cycloheximide. These results suggest an induction of synthesis of some proteins responsible for the release of arachidonic acid, which might be attributed to protein kinase-C activation in arachidonic acid metabolism stimulated by PMA or EGF. Moreover, PGE2 production was potently induced by PMA and slightly by EGF in the cyclooxygenase-inactivated cells by acetyl salicylate pretreatment, which also suggests that both agents might induce the synthesis of cyclooxygenase in cultured porcine thyroid cells, although we did not measure its activity.