Inhibition of p70 S6 kinase (S6K1) activity by A77 1726 and its effect on cell proliferation and cell cycle progress.

Leflunomide is a novel immunomodulatory drug prescribed for treating rheumatoid arthritis. It inhibits the activity of protein tyrosine kinases and dihydroorotate dehydrogenase, a rate-limiting enzyme in the pyrimidine nucleotide synthesis pathway. Here, we report that A77 1726, the active metabolite of leflunomide, inhibited the phosphorylation of ribosomal protein S6 and two other substrates of S6K1, insulin receptor substrate-1 and carbamoyl phosphate synthetase 2, in an A375 melanoma cell line. A77 1726 increased the phosphorylation of AKT, p70 S6 (S6K1), ERK1/2, and MEK through the feedback activation of the IGF-1 receptor-mediated signaling pathway. In vitro kinase assay revealed that leflunomide and A77 1726 inhibited S6K1 activity with IC50 values of approximately 55 and 80 µM, respectively. Exogenous uridine partially blocked A77 1726-induced inhibition of A375 cell proliferation. S6K1 knockdown led to the inhibition of A375 cell proliferation but did not potentiate the antiproliferative effect of A77 1726. A77 1726 stimulated bromodeoxyuridine incorporation in A375 cells but arrested the cell cycle in the S phase, which was reversed by addition of exogenous uridine or by MAP kinase pathway inhibitors but not by rapamycin and LY294002 (a phosphoinositide 3-kinase inhibitor). These observations suggest that A77 1726 accelerates cell cycle entry into the S phase through MAP kinase activation and that pyrimidine nucleotide depletion halts the completion of the cell cycle. Our study identified a novel molecular target of A77 1726 and showed that the inhibition of S6K1 activity was in part responsible for its antiproliferative activity. Our study also provides a novel mechanistic insight into A77 1726-induced cell cycle arrest in the S phase.

Major histocompatibility complex class I-related chain A/B (MICA/B) expression in tumor tissue and serum of pancreatic cancer: role of uric acid accumulation in gemcitabine-induced MICA/B expression.

BACKGROUND: Major histocompatibility complex class I-related chain A and B (MICA/B) are two stress-inducible ligands that bind the immunoreceptor NKG2D and play an important role in mediating the cyotoxicity of NK and T cells. In this study, we sought to study MICA/B expression in pancreatic cancer and to determine whether and how genotoxic drugs such as gemcitabine can affect MICA/B expression and natural killer cytotoxity. METHODS: Seven pancreatic cancer cell lines were analyzed for MICA/B expression by flow cytometry and for their sensitivity to NK-92 cell killing by a 51Cr release assay. MICA/B expression in tumor tissues and sera of pancreatic cancer was analyzed by immunohistochemical staining (IHC) and ELISA, respectively. RESULTS: Two MICA/B-positive cell lines were sensitive to the cytotoxic activity of NK-92 cells. Other two MICA/B-positive cell lines and three MICA/B-negative cell lines were resistant to NK-92 cell killing. MICA/B expression was positive in 17 of 25 (68%) pancreatic ductal adenocarcinomas but not in normal pancreatic ductal epithelial cells. Serum MICA/B levels were significantly elevated in patients with pancreatic adenocarcinomas but did not correlate with the stage of pancreatic cancer and patient survival. Gemcitabine therapy led to increased serum MICA levels in 6 of 10 patients with detectable serum MICA. Allopurinol, an inhibitor of xanthine oxidoreductase that converts xanthine to uric acid, blocked uric acid production, MICA/B expression, and sensitivity to NK-92 cell killing toward a PANC-1 cancer cell line exposed to radiation and two genotoxic drugs, gemcitabine and 5-fluorouracil. CONCLUSIONS: The levels of MICA/B expression in serum and tissue of pancreatic cancer are elevated. DNA damage-induced MICA/B expression is mediated through increased uric acid production.

Effectiveness of multidrug therapy in multibacillary leprosy: a long-term follow-up of 34 multibacillary leprosy patients treated with multidrug regimens till skin smear negativity.

The World Health Organization (WHO) Field Trials of multidrug therapy (MDT) started at Schieffelin Leprosy Research and Training Centre (SLR & IC), Karigiri, India in December 1981. The patients were treated with two MDT regimens. The first (regimen A) consisted of 600mg rifampicin and 300mg of clofazimine given under supervision on 2 consecutive days monthly, 225mg injection of acedapsone bimonthly and dapsone 100mg daily. The second regimen (regimen B) was the conventional MDT (WHO/MDT), rifampicin 600mg and clofazimine 300mg supervised once a month, dapsone 100mg and clofazimine 50mg daily, unsupervised. Both the regimens were administered for a minimum period of 2 years or until skin smear negativity, whichever occurred later. Thirty-four newly detected previously untreated MB patients, 16 of whom received regimen A and 18 regimen B, were reassessed. Both regimens were well accepted and well tolerated by the patients. Clofazimine discolouration was the only adverse effect of MDT seen in these patients. After completion of treatment with MDT, the patients were followed up for a total duration of 466 person-years with a mean of 13.7 +/- 1.4 years per patient. No relapse was seen.

Relapse as histoid leprosy after receiving multidrug therapy (MDT); a report of three cases

The histoid type of leprosy has been described as occurring in lepromatous leprosy patients who relapse after many years of apparently successful dapsone monotherapy. Three patients who had received the World Health Organization-recommended regimens of multidrug therapy (WHO/MDT) relapsed as histoid leprosy 12-15 years after completion of treatment. In one patient, through mouse foot pad studies, the bacilli were found to be sensitive to rifampin and clofazimine and resistant to dapsone. In the other two patients mouse foot pad studies were inconclusive. The patients were re-started on WHO/MDT. Two patients took regular treatment and improved, both clinically and bacteriologically. One patient was irregular in treatment, and 1 year after re-starting WHO/MDT nodules were still present although the bacterial index had fallen slightly.