A pilot study of radiologic measures of abdominal adiposity: weighty contributors to early pancreatic carcinogenesis worth evaluating?

OBJECTIVE: Intra-abdominal fat is a risk factor for pancreatic cancer (PC), but little is known about its contribution to PC precursors known as intraductal papillary mucinous neoplasms (IPMNs). Our goal was to evaluate quantitative radiologic measures of abdominal/visceral obesity as possible diagnostic markers of IPMN severity/pathology. METHODS: In a cohort of 34 surgically-resected, pathologically-confirmed IPMNs (17 benign; 17 malignant) with preoperative abdominal computed tomography (CT) images, we calculated body mass index (BMI) and four radiologic measures of obesity: total abdominal fat (TAF) area, visceral fat area (VFA), subcutaneous fat area (SFA), and visceral to subcutaneous fat ratio (V/S). Measures were compared between groups using Wilcoxon two-sample exact tests and other metrics. RESULTS: Mean BMI for individuals with malignant IPMNs (28.9 kg/m2) was higher than mean BMI for those with benign IPMNs (25.8 kg/m2) (P=0.045). Mean VFA was higher for patients with malignant IPMNs (199.3 cm2) compared to benign IPMNs (120.4 cm2),P=0.092. V/S was significantly higher (P=0.013) for patients with malignant versus benign IPMNs (1.25vs. 0.69 cm2), especially among females. The accuracy, sensitivity, specificity, and positive and negative predictive value of V/S in predicting malignant IPMN pathology were 74%, 71%, 76%, 75%, and 72%, respectively. CONCLUSIONS: Preliminary findings suggest measures of visceral fat from routine medical images may help predict IPMN pathology, acting as potential noninvasive diagnostic adjuncts for management and targets for intervention that may be more biologically-relevant than BMI. Further investigation of gender-specific associations in larger, prospective IPMN cohorts is warranted to validate and expand upon these observations.

3-(2'-Bromopropionylamino)-benzamides as novel S-phase arrest agents.

We report the synthesis, antiproliferative activity, and SAR of novel 3-(2'-bromopropionylamino)-benzamides. Many of the benzamide compounds showed potent cytotoxicities against Molt-3 leukemia cells. Several compounds exihibited cytotoxicities (under 6.5 microM) against five solid tumor cell lines. The mechanism of action of the most potent benzamide 10l does not involve targeting on tubulin but it causes cell cycle S-phase arrest. This active S-phase arrest agent merits further investigation.

[A study of the bacteriophage-based assay for the detection of pyrazinamide resistance in Mycobacterium tuberculosis].

OBJECTIVE: To set up and evaluate the method of phage amplified biologically assay (PhaB) in rapid detection of pyrazinamide (PZA) resistance. METHODS: The PhaB assay was developed and applied in detecting PZA resistance in 108 clinical isolates of Mycobacterium tuberculosis and the results were compared with those of the absolute concentration method. The minimum inhibitory concentration (MIC) was detected for all discrepancy isolates. RESULTS: The results showed that the optimal detecting condition was pH 5.5, PZA 200 microg/ml and 48 h. Of the 108 strains of Mycobacterium tuberculosis, 28 strains were PZA-susceptible and 80 strains were PZA-resistant detected by PhaB; while 32 strains were PZA-susceptible and 76 strains were PZA-resistant by absolute concentration method. Twenty-eight of the 108 strains were PZA-susceptible and 71 were PZA-resistant by the two methods. The concordant isolates of determination of PZA resistance were 99 by the two methods and the concordance rates was 91.7%. There were 9 strains in discordant isolates, of which 7 were the same with MIC method and gene chip in drug susceptibility. If the results of absolute concentration method was the gold standard, the sensitivity, specificity, positive and negative predictive values, as well as accuracy of PhaB assay was 94.7%, 84.8%, 93.4%, 87.5% and 91.7% respectively. CONCLUSION: The PhaB assay can be used as a rapid screening method for detection of drug susceptibility of PZA in clinical isolates of Mycobacterium tuberculosis.

Antiviral therapeutic efficacy of foscarnet in hepatitis B virus infection.

Foscarnet (PFA), a viral DNA polymerase inhibitor, is a clinical agent for herpes viruses. The goal of the study was to evaluate the therapeutic efficacy of PFA in hepatitis B virus (HBV) infection. Intravenous infusion of PFA (1 g/day) for 4 weeks significantly reduced serum HBeAg (p<0.01) and HBV DNA copies (p<0.05) in 31 patients who were diagnosed with active chronic HBV infection (CHB) and had not received antiviral treatment previously. Alanine aminotransaminase (ALT), aspartate aminotransaminase (AST) and gamma glutamyl transpeptidase (gamma-GT) of the patients declined (p<0.001, 0.001 and 0.01, respectively). Kidney function (blood creatinine and urea nitrogen) remained unchanged. Another 21 lamivudine-resistant CHB patients with mutations at the tyrosine-methionine-aspartate-aspartate motif (YMDD) displayed a response to PFA similar to that mentioned above, with reductions in HBeAg (p<0.05), HBV DNA (p<0.01) and liver enzymes (ALT and AST, p<0.001; gamma-GT, p<0.05). Moreover, PFA reduced serum HBeAg (p<0.01), HBV DNA (P<0.05), AST (p<0.05) and ALT (p<0.02) in a cohort of 13 severe CHB patients with advanced liver damage. PFA was also evaluated in vitro and in vivo. PFA inhibited HBV DNA replication in HBV-transfected human HepG2 cells (2.2.15 cells) with reduced amount of HBV RC-DNA and DS-DNA. In the duck HBV-infected ducklings, PFA reduced viral DNA and duck HBsAg in the serum (p<0.01 for both).

[Effect of Chinese herbal medicine Xin-kang oral liquid on interferon-induction and its antiviral activity in coxsackievirus B3 infected mice].

OBJECTIVE: To investigate the effect of Chinese herbal medicine Xin-kang oral liquid on interferon (IFN)-induction and its antiviral activity in Coxsackievirus B3 virus strain (CVB3) infected mice. METHODS: The Xin-kang oral liquid was given orally to mice two days prior to the challenge of CVB3 virus to induce myocarditis. Two dosages of Xin-kang oral liquid crude herbal medicine 30 g x kg(-1) x d(-1) and 12 g x kg(-1) x d(-1) were given to the mice of different treatment groups respectively, sterilized water was given to the mice of virus control group. IFN-alpha 10(6) U x kg(-1) x d(-1) S.C was given to the infected mice as positive drug control group. The mice were sacrificed on 5th, 10th and 20th day of infection for evaluation, the levels of serum interferon (IFN) were titrated with vesicular stomatitis virus (VSV) and cardiac tissue was fixed and sectioned. The quantitative histological changes at various stages of myocarditis were observed. RESULTS: In the infected mice fed with 30 g x kg(-1) x d(-1) or 12 g x kg(-1) x d(-1) of Xin-kang oral liquid orally for 5, 10 and 20 days, the mean titer of serum IFN of Xin-Kang oral liquid treated group was markedly higher (29.3 U/0.1 ml) than that of virus control group (12.6 U/0.1 ml). The level of serum IFN in IFN treated positive control mice was lower than that of Xin-kang treatment groups. The histological examination showed extensive myocardial necrosis and cellular infiltration in virus control group, but necrosis and cellular infiltration were less severe in Xin-kang treatment goups of mice. It is demonstrated that there were close correlation between the degree of myocardial lesions and the level of IFN-induction in treated mice. CONCLUSION: Xin-kang oral liquid could facilitate the induction of endogenous interferon that exerted its antiviral activity in CVB3 infected mice. This can help us to understand better the mechanism of anti-CVB3 effect of Xin-Kang oral liquid.

[Antitumor mechanism of 3-bromopropionylamino benzoylurea on leukemia and lymphoma].

AIM: To study the antitumor mechanism of 3-bromopropionylamino benzoylurea (JIMB01) on leukemia and lymphoma. METHODS: The antitumor effects of JIMB01 in cell culture was detected by MTT staining. JIMB01-induced apoptosis in leukemia and lymphoma cells was tested by Giemsa staining, fluorescent Hoechst33258 staining, as well as DNA gel electrophoresis. Cell cycle was analyzed by flow cytometry. JIMB01-induced Bcl-2 phosphorylation in CEM cell lines was detected by Western blot methods. The activities of caspase-3 and caspase-8 were determined by colorimetric protease assay and that of caspase-9 was determined by fluorescent intensity. RESULTS: This compound showed antiproliferative activities in a panel of nine human leukemia and lymphoma cell lines with IC50 values in the range of 0.25 micromol x L(-10 to 0. 51 micromol x L(-1). Morphological observation and cell cycle analysis indicated that CEM cells were blocked at mitosis phase by JIMB01. The fluorescent Hoechst33258 staining showed apoptotic nuclear degradation dispersed in the cytoplasm of CEM cells exposed to JIMB01 at 0. 80 micromol x L(-1) for 24 h. DNA degradation in the form of a multiple-unit DNA ladder was clearly demonstrated in CEM leukemia cells treated with JIMB01 at 0.15 micromol x L(-1) or higher for 24 h using agarose gel electrophoresis. Bcl-2 phosphorylation became visible, in Western blot, within 6 h in CEM cells treated with JIMB01 at 0.15 micromol x L(-1) or higher for 24 h. JIMB01 increased the activities of caspase-3, -8 and -9 in CEM cells; DEVD-fmk, a caspase-3 inhibitor, inhibited the cytotoxicity of JIMB01 in CEM leukemia cells. CONCLUSION: The antitumor mechanism of JIMB01 is that JIMB01 may induce tumor cell apoptosis through Bcl-2 phosphorylation and then caspase passway.

Inhibition of human hepatocellular carcinoma by L-proline-m-bis (2-chloroethyl) amino-L-phenylalanyl-L-norvaline ethyl ester hydrochloride (MF13) in vitro and in vivo.

A new anticancer tripeptide, L-proline-m-bis (2-chloroethyl) amino-L-phenylalanyl-L-norvaline ethyl ester hydrochloride (MF13), was investigated for its activity and mechanism in human hepatocellular carcinoma (HCC) cell lines. MF13 showed antiproliferative activities in the panel of 7 human HCC cell lines with IC50 in the range of 0.08-2.32 microM. A significant blockade in the S-phase occurred in tumor cells 12 h after their exposure to MF13. The inactivated Rb (phosphorylated Rb, pRb), which is present in the S-phase, was increased within 6 h of treatment. Bcl-2 expression was without change in hepatocarcinoma cells treated with MF13; however, a significant increase of bax was observed, resulting in a decreased ratio of bcl-2/bax. Increased activity of caspase-9, -8 and -3 was detected in the MF13 treated cells, indicating an activated pathway of apoptosis by MF13. Morphological examination as well as DNA gel electrophoresis demonstrated a nuclear fragmentation and DNA degradation in the form of multiple-unit DNA ladder in MF13 treated tumor cells. MF13 alone at 10 mg/kg (i.p.) inhibited HepG2 tumor in nude mice by more than 94% in volume. Bel-7402 tumor originated from a Chinese patient with HCC exhibited a sensitivity to MF13 similar to HepG2 in vivo. Antitumor effect of MF13 in the nude mice bearing human hepatocarcinoma (Bel-7402 or HepG2) was stronger than mitomycin C as well as its precursor m-sarcolysin (p<0.01), and comparable with cyclophosphamide. We believe MF13 merits consideration for further investigation as an agent against human hepatocellular carcinoma.

[Study on the method for rapid detection of Mycobacterium tuberculosis by phage amplified biologically assay].

OBJECTIVE: To set up a method for rapid detection of Mycobacterium tuberculosis (MTB) by phage amplified biologically (PhaB) assay and to investigate the optimal test condition. METHODS: Various test conditions were compared in order to observe the influence on detective results after MTB was infected by Mycobacteriophage. The test condition established was used for detection of sputum samples, and the results were compared with BIOTEC Lab test. RESULTS: The bacterial concentration of MTB in 200 - 500/ml was detected by PhaB assay at 1 x 10(9) plaque forming unite (PFU)/ml of Mycobacteriophage, 37 degrees C for 60 min. The optimal concentration of virucidal for inactivation of Mycobacteriophage was 100 mmol/ml for 5 min at room temperature. The bacteriolytic plaque was clear at the concentration of 1 x 10(8)/ml indicator cells. Bacterium inactivated by heat can not be infected by Mycobacteriophage. Positive result was observed for control strains of H(37)Rv, H(37)Ra and M. bovis while negative result was obtained for 7 strains of non-Mycobacterium and 16 control strains of non-Tuberculosis Mycobacteria (NTM). The 4 strains of NTM (M. fortuitum, M. intrcellulare, M. aurum, M. phlei) showed positive reaction at higher concentrations (> 1 x 10(5)/ml). The repetition test showed that the differentiation coefficient in batch and inner was all under 15%. There was a significantly difference (P < 0.01) in positive rate between two digestion-decontamination procedure with N-acetyl-cysteine-NaOH liquefacient (94%) and NaOH liquefacient (62%). The positive rate of the samples cultured one day (65%) was significantly higher than that of the samples without preculture (40%). The results for detection of clinical samples by two reagents, ours and BIOTEC Lab, were nearly the same. CONCLUSION: Because its rapidity, simplicity, and sensitivity, PhaB assay can be used for rapid detection of MTB, but the condition of test is very important.

Inhibition of microtubule polymerization by 3-bromopropionylamino benzoylurea (JIMB01), a new cancericidal tubulin ligand.

3-Bromopropionylamino benzoylurea (JIMB01) is a small molecular weight compound (MW 313) that has been synthesized in our laboratory. This compound showed antiproliferative activities in a panel of thirteen human tumor cell lines with IC(50) values in the range of 0.25 to 0.51 micro M for leukemia and lymphoma cell lines and 0.33 to 9.26 micro M for solid tumor cell lines. The primary action of JIMB01 is to inhibit microtubule polymerization but not depolymerization. A 4 micro M concentration of the compound caused a complete inhibition of microtubule assembly in a cell-free reaction. An increase in the number of human hepatocarcinoma cells blocked in the M-phase was detected 12hr after exposure to JIMB01. The kinase activity of cyclin B1, which is responsible for the G(2)/M transition, was increased accordingly. Bcl-2 phosphorylation became visible, in a western blot, within 6hr in hepatocarcinoma cells treated with JIMB01 at 0.8 micro M or higher. JIMB01-induced apoptosis in liver cancer cells was confirmed by morphological methods, flow cytometry, as well as DNA gel electrophoresis, which clearly demonstrated DNA degradation in the form of a multiple-unit DNA ladder. Furthermore, in vivo experiments using nude mice showed that intraperitoneal injection of JIMB01 at 15mg/kg (with seven injections at 4-day intervals) significantly inhibited the growth of a human hepatocarcinoma (BEL-7402) by 66% in tumor volume (P=0.01), at least compatible to the inhibition by vincristine (43% inhibition), indicating good bioavailability of the compound in the circulation. Side-effects of the compound were not observed, and the body weight of the treated mice remained stable during the 4-week treatment. Since JIMB01 is a small compound, targets a specific molecule in tumor cells, and has promising activity against human hepatocarcinoma in vivo, we believe JIMB01 merits consideration for further investigation.

Double blockade of cell cycle at g(1)-s transition and m phase by 3-iodoacetamido benzoyl ethyl ester, a new type of tubulin ligand.

3-Iodoacetamido benzoyl ethyl ester (3-IAABE) is a new compound synthesized in our laboratory. The primary action of 3-IAABE is to inhibit microtubule assembly by interacting with -SH groups on tubulin. In contrast to other known microtubule disrupters, 3-IAABE caused a double blockade in the cell cycle at G(1)-S transition and in M phase. The blockade was determined by cell cycle analysis and chromosome distribution. Kinase activities of cyclin E and cyclin-dependent kinase 2 responsible for the G(1)-S transition were increased, as were the activities of mitotic cyclin B and cdc2. 3-IAABE treatment also increased p53 expression and dephosphorylated (or activated) retinoblastoma protein. Investigation of the signal transduction pathway showed that 3-IAABE induced bcl-2 phosphorylation, followed by activation of caspase-9, -3, and -6, but not caspase-8. DNA fragmentation factor and poly(ADP-ribose) polymerase, the downstream substrates of caspase-3 and -6, were cleaved after 3 h of exposure to 3-IAABE, followed by DNA fragmentation. Pretreatment of the cells with inhibitors of caspase-9, -3, or -6, respectively, inhibited the cleavage of DNA fragmentation factor and poly(ADP-ribose) polymerase and thus inhibited the onset of apoptosis. 3-IAABE showed antitumor activities in the panel of 60 National Cancer Institute human tumor cell lines with total growth inhibition in the range of 0.22-4.3 micro M for solid tumor lines and 0.025-0.22 micro M for leukemia/lymphoma cell lines. The 3-IAABU total growth inhibition of phytohemagglutinin-stimulated healthy human lymphocytes was 450-fold greater than that of leukemic cells. 3-IAABE significantly inhibited the growth of human hepatocarcinoma (BEL-7402) in nude mice by 72% in tumor volume, more strongly than did vincristine (43 percent inhibition). Besides being a novel lead for the design of new anticancer tubulin ligands, the activity of 3-IAABE in the cell cycle may also help us to understand the molecular pharmacology of microtubule-active drugs.