Shinji Endo

Potent in vitro and in vivo antitumor effects of MDM2 inhibitor nutlin‐3 in gastric cancer cells

The tumor suppressor gene p53 is the most frequently mutated gene in human cancers. However, its mutation rate is relatively low in gastric cancer compared with other cancers. In this study, we investigated the mechanisms underlying the antitumor effects of nutlin‐3, an inhibitor of human homolog of murine double minute 2 (MDM2). MDM2 is a negative regulator of p53. Four gastric cancer cell lines with wild‐type p53 (wt p53) and three with mutant‐type p53 (mt p53) were analyzed for MDM2 and MDM4 expression by immunoblotting, and for their gene amplification by quantitative real‐time PCR. Moreover, the viability of cells exposed to nutlin‐3 was examined by WST‐8 assay, and the expression of p53 and its downstream genes was analyzed by immunoblotting. Nutlin‐3 stabilized p53 and increased the expression of p21WAF1 and Noxa, and cleaved poly (ADP)‐ribose polymerase regardless of the pre‐expression levels of MDM2 and MDM4 in gastric cancer cells with wt p53. Flow cytometry revealed that nutlin‐3 arrested the cell cycle in G1 phase and induced apoptosis in the cell lines. These nutlin‐3 effects were not observed in the cell lines with mt p53. Nutlin‐3 exerted additive or synergistic cytotoxicity in combination with 5‐fluorouracil or cisplatin in most cell lines with wt p53. An in vivo antitumor effect of nutlin‐3 alone and its additive augmentation by 5‐fluorouracil were confirmed in an MDM2 overexpressed xenograft tumor model. Nutlin‐3 showed potent antitumor activity against human gastric cancer cells with wt p53 and shows promise as a single agent and in combination with conventional anticancer drugs. (Cancer Sci 2011; 102: 605–613)

Granular cell tumor occurring in the sigmoid colon treated by endoscopic mucosal resection using a transparent cap (EMR-C)

A case of granular cell tumor occurring in the sigmoid colon is reported. The patient, a 56-year-old man, visited our hospital for further evaluation of occult blood in his stool. Endoscopic examination revealed a yellowish, hemispheric submucosal tumor (SMT) with redness, about 6 mm in diameter, in the sigmoid colon. Endoscopic mucosal resection using a transparent cap (EMR-C) was performed, and histological examination revealed that the tumor consisted of a nested growth of large tumor cells with ample granular cytoplasm and small round nuclei. The tumor cells expressed S-100 protein and were stained with neuron-specific enolase (NSE) and periodic acid-Schiff, but were negative for desmin, vimentin, and cytokeratin. The resected tumor was diagnosed as a granular cell tumor. This may be the first report of a colorectal granular cell tumor successfully treated with EMR-C.

Therapeutic efficacy of the Qing Dai in patients with intractable ulcerative colitis

Ulcerative colitis (UC) is a chronic inflammatory bowel disease that may become intractable when treated with conventional medications such as aminosalicylates, corticosteroids, and azathioprine. The herbal medicine Qing Dai has traditionally been used in Chinese medicine to treat UC patients, but there is a lack of published data on the efficacy of Qing Dai in UC treatment. We report several cases of patients with intractable UC who take Qing Dai in a retrospective observational study. Furthermore, we explore the mechanisms of action of Qing Dai. Nine patients with active UC who received conventional medications but wished to receive Qing Dai as an alternative medication were included in our analysis. The UC severity level was determined based on the clinical activity index (CAI). Additionally, 5 of the 9 patients were endoscopically evaluated according to the Matts grading system. Each patient received 2 g/d of Qing Dai orally and continued taking other medications for UC as prescribed. Electron spin resonance was applied to explore the mechanisms of action of Qing Dai. After 4 mo of treatment with Qing Dai, the CAI score decreased from 8.3 ± 2.4 to 2.4 ± 3.4 (mean ± SD; P < 0.001). Similarly, the endoscopic Matts grade decreased from 3.4 ± 0.5 to 2.2 ± 0.8 (P = 0.02). Six of 7 patients who were on prednisolone upon enrollment in the study were able to discontinue this corticosteroid. Electron spin resonance revealed that Qing Dai possesses strong hydroxyl radical scavenging activity. Qing Dai showed significant clinical and endoscopic efficacy in patients who failed to respond to conventional medications. Scavenging of hydroxyl radicals appears to be a potential mechanism through which Qing Dai acts, but the significance of the scavenging ability of Qing Dai with respect to the anti-inflammatory effect in UC patients warrants further investigation.

Antitumor Effects of a Sirtuin Inhibitor, Tenovin-6, against Gastric Cancer Cells via Death Receptor 5 Up-Regulation

Up-regulated sirtuin 1 (SIRT1), an NAD+-dependent class III histone deacetylase, deacetylates p53 and inhibits its transcriptional activity, leading to cell survival. SIRT1 overexpression has been reported to predict poor survival in some malignancies, including gastric cancer. However, the antitumor effect of SIRT1 inhibition remains elusive in gastric cancer. Here, we investigated the antitumor mechanisms of a sirtuin inhibitor, tenovin-6, in seven human gastric cancer cell lines (four cell lines with wild-type TP53, two with mutant-type TP53, and one with null TP53). Interestingly, tenovin-6 induced apoptosis in all cell lines, not only those with wild-type TP53, but also mutant-type and null versions, accompanied by up-regulation of death receptor 5 (DR5). In the KatoIII cell line (TP53-null), DR5 silencing markedly attenuated tenovin-6-induced apoptosis, suggesting that the pivotal mechanism behind its antitumor effects is based on activation of the death receptor signal pathway. Although endoplasmic reticulum stress caused by sirtuin inhibitors was reported to induce DR5 up-regulation in other cancer cell lines, we could not find marked activation of its related molecules, such as ATF6, PERK, and CHOP, in gastric cancer cells treated with tenovin-6. Tenovin-6 in combination with docetaxel or SN-38 exerted a slight to moderate synergistic cytotoxicity against gastric cancer cells. In conclusion, tenovin-6 has potent antitumor activity against human gastric cancer cells via DR5 up-regulation. Our results should be helpful for the future clinical development of sirtuin inhibitors.

The sirtuin inhibitor tenovin-6 upregulates death receptor 5 and enhances cytotoxic effects of 5-fluorouracil and oxaliplatin in colon cancer cells.

It has been reported that upregulated SIRT1 (NAD(+)-dependent class III histone deacetylase) deacetylates the p53 protein, represses its function, and allows for tumor cell growth in various cancers. Here we investigated antitumor effects of tenovin-6, a small-molecule inhibitor of SIRT1 and SIRT2, in various colon cancer cell lines. Tenovin-6 induced apoptosis in all five colon cancer cell lines investigated (two cell lines with wild-type p53 and three with mutant p53) regardless of the p53 mutation status. This effect was accompanied by accumulation of death receptor 5 (DR5) in most cell lines. DR5 silencing in HCT116 cells strongly attenuated tenovin-6-induced apoptosis. We investigated the effect of combining tenovin-6 with conventional anticancer agents 5-fluorouracil (5-FU), SN-38 (an active metabolite of irinotecan), and oxaliplatin. Synergistic antitumor effects of tenovin-6 were observed in combination with either 5-FU or oxaliplatin in vitro. The combination of tenovin-6 and oxaliplatin exhibited potent growth inhibition of HCT116 xenograft tumors in vivo. In conclusion, tenovin-6 induced apoptosis in human colon cancer cells through the activation of the DR5 signaling pathway and enhanced the antitumor properties of 5-FU and oxaliplatin. These results may help develop a novel treatment option for colorectal cancer using a SIRT inhibitor.

Enhanced specificity of HPV16 E6E7 siRNA by RNA-DNA chimera modification.

Although efforts have been made to develop new drugs for infectious and neoplastic diseases utilizing synthetic small interfering RNA(siRNAs), those intrinsically have undesirable effects, including silencing of unintended genes (off-target effect) and nonspecific cytotoxicity. Off-target effects can be avoided by DNA substitution in the guide strand (GS) seed region of nucleotide positions 1–8 and its complementary part of the passenger strand plus the 3′ overhang, which is designated as a double-strand RNA–DNA chimera (dsRDC). In this study, we found that the specificity of potent siRNAs targeting human papillomavirus 16 (HPV16) E6 and E7 oncogenes, which we previously reported, could be enhanced by short dsRDC modification (first six nucleotides from the 5′ end of the GS and its complementary nucleotides of the passenger strand). Such dsRDC modification reduced nonspecific cytotoxicity in two of three siRNAs (497 and 752), although not in the other (573), which correlated with their off-target effects. In addition, silencing activity was marginally impaired in two dsRDCs (497 and 573) and moderately in one (752). Finally, dsRDC-497 induced E6E7-specific growth suppression of cervical cancer cells as well as E6E7-immortalized human keratinocytes. Our results show that dsRDC modification enhances the specificity of E6E7 siRNA, which is required for use in in vivo settings.

Panitumumab-Induced Interstitial Lung Disease in a Case of Metastatic Colorectal Cancer

Background: A Japanese postmarketing survey of panitumumab revealed that panitumumab-associated interstitial lung disease (ILD) occurred in approximately 1% (19/1767) of patients, causing death in 36.8% of these cases. Case Report: We report the case of a 60-year-old Japanese man who developed ILD associated with panitumumab therapy (third-line therapy) for metastatic sigmoid colon cancer involving the liver, lymph nodes, and lungs. 2 months after the initiation of panitumumab therapy, he developed a progressive nonproductive cough, dyspnea, and a fever, and was diagnosed with ILD. Intravenous pulse methylprednisolone treatment led to quick recovery. The patient had some risk factors for ILD associated with epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors. Conclusion: Further studies are required to elucidate the association between anti-EGFR antibodies and ILD.

Correlations of survival with progression-free survival, response rate, and disease control rate in advanced biliary tract cancer: a meta-analysis of randomised trials of first-line chemotherapy.

Background:The need to promote novel drug development for advanced biliary tract cancer (ABTC) has emphasised the importance of determining whether various efficacy end points can act as surrogates for overall survival (OS).Methods:We conducted a literature search of randomised trials of first-line chemotherapy for ABTC and investigated correlations between efficacy end points and OS using weighted linear regression analysis. The ratios of the median OS, median progression-free survival (PFS), response rate, and disease control rate in each trial were used to summarise treatment effects. The surrogate threshold effect (STE), which was the minimum treatment effect on PFS required to predict a non-zero treatment effect on OS, was calculated.Results:Seventeen randomised trials with 36 treatment arms were identified, and a sample size of 2148 patients with 19 paired arms was analysed. The strongest correlation between all evaluated efficacy end points was observed between median OS and median PFS ratios (r2=0.66). In trials with gemcitabine-containing therapies and targeted agents, the r2-values were 0.78. The STE was estimated at 0.83 for all trials and 0.81 for trials with gemcitabine-containing therapies, and was not calculated for trials with targeted agents.Conclusions:The median PFS ratio correlated well with the median OS ratio, and may be useful for planning a clinical trial for novel drug development.

C-Arm Cone Beam Computed Tomography Guidance for Radiofrequency Ablation in Hepatocellular Carcinoma.

Objective: To assess the usefulness of C-arm cone beam computed tomography (CBCT) combined with ultrasound for the treatment of hepatocellular carcinoma (HCC) by radiofrequency ablation (RFA). Methods: Patients underwent RFA following transcatheter arterial chemoembolization (TACE) or RFA alone under ultrasound or CBCT guidance combined with ultrasound-based techniques. They were divided into 2 groups based on the use (C group) and nonuse (NC group) of CBCT guidance. The technical success of RFA and local tumor progression after the first RFA session were evaluated by dynamic contrast-enhanced imaging methods. Between-group differences were assessed retrospectively. Results: We enrolled 198 patients with 260 HCC nodules. The complete ablation rates were 63.0 and 89.4% in the NC and C groups, respectively. In log-rank testing, local tumor progression occurred significantly more often in the NC group when RFA was used without TACE, in males when des-gamma-carboxy prothrombin was ≥29 mAU/mL, and when the diameter of a nodule was ≥18 mm. On Cox proportional-hazards regression analysis, the NC group, RFA alone without TACE, and male gender were significant independent variables. Conclusion: TACE followed by RFA under CBCT and ultrasound guidance improves the reliability of ablation of target HCC nodules, reduces the need for additional treatment sessions, and prevents local tumor progression.

Successful Treatment with Modified FOLFOX6 and Panitumumab in a Cecal Cancer Patient Undergoing Hemodialysis

Combination chemotherapy of mFOLFOX6 (5-fluorouracil, leucovorin, and oxaliplatin) plus panitumumab, a fully human monoclonal antibody against epidermal growth factor receptor (EGFR), is one of the standard treatments for metastatic colorectal cancer (mCRC) without KRAS mutation. A few reports suggested no need of dose adjustment of cetuximab, a similar chimeric anti-EGFR antibody, in patients with renal impairment. However, panitumumab combined with cytotoxic drugs for hemodialysis patients has not been reported. We herein report a case of a hemodialysis mCRC patient successfully treated with mFOLFOX6 and panitumumab combination therapy.