Kunihiro Takanashi

Aberrant Crypt Foci: Detection, Gene Abnormalities, and Clinical Usefulness

Human aberrant crypt foci (ACF) were first identified as lesions consisting of large thick crypts in colonic mucosa of surgical specimens after staining with methylene blue. Previously we succeeded in identifying ACF by using magnifying endoscopy and analyzed the number, size, and dysplastic features of ACF in normal controls and patients with adenoma or cancer patients. On the basis of these analyses, we strongly suggested that ACF, particularly dysplastic ACF, are precursor lesions of the adenoma-carcinoma sequence in humans. In most sporadic ACF, K-ras mutations were positive, but APC mutations were negative irrespective of nondysplastic or dysplastic features. Conversely, in most ACF from familial adenomatous polyposis patients, APC mutations were positive but K-ras mutations were negative. These results may suggest that the molecular mechanism of sporadic colon carcinogenesis is not necessarily the same as that of familial adenomatous polyposis. It was shown that ACF acquired resistance to apoptosis induced by bile salts, whereas normal colonic epithelial cells are turning over consistently by apoptosis. This apoptosis resistance was closely associated with glutathione S-transferase P1-1 expression. One of the most important clinical applications of ACF observation with magnifying endoscopy is its use as a target lesion for chemoprevention. Because ACF are tiny lesions, they should be eradicated during a short time by administration of chemopreventive agents. In fact, we performed an open chemopreventive trial of sulindac and found that the number of ACF was reduced markedly in 2 months. We currently are proceeding with a randomized double-blind trial targeting ACF.

Chemoprevention of colorectal cancer

Colorectal cancer is a disease with a high mortality rate and it has been increasing in prevalence worldwide. Chemoprevention, as well as primary and secondary prevention, for colorectal cancer have attracted much attention. Many chemopreventive trials have been performed, and several agents, including nonsteroidal antiinflammatory drugs, such as aspirin and sulindac, cyclooxygenase-2 selective inhibitors, such as celecoxib, vitamin D, folate, and calcium, have been shown to have some effect. In these chemopreventive trials, the targeted lesions used for evaluation were mainly polyps. However, the chemopreventive effects of some agents on polyps may require several years to evaluate. Further, larger polyps may not be susceptible to chemopreventive agents. Aberrant crypt foci (ACF) are tiny lesions at the earliest stage of colorectal carcinogenesis, which consist of large, thick crypts identified by dense, methylene blue staining. We succeeded in identifying human ACF in situ using magnifying endoscopy and found that the number of ACF, particularly dysplastic ACF, increased significantly from normal subjects to adenoma patients and then to cancer patients. We also found that the number, size, and dysplastic features of ACF are significantly correlated with the number of adenomas in adenoma patients. Thus, it was surmised that ACF are precursor lesions of the adenoma-carcinoma sequence in humans and that ACF may be the most appropriate lesions as targets for chemoprevention. We have shown that the number of ACF was significantly reduced in patients treated with sulindac. We are currently proceeding with a randomized, double-blind, chemopreventive trial targeting ACF.

Exacerbation of bloody diarrhea as a side effect of mesalamine treatment of active ulcerative colitis.

Mesalamine has been used as the first-line therapy for the treatment of ulcerative colitis (UC) because of its efficacy and fewer side effects. However, earlier study showed that mesalamine occasionally causes diarrhea. We are presenting a patient with active UC in whom bloody diarrhea accompanied by abdominal pain and fever occurred and the symptoms were aggravated after administration of mesalamine. In order to clarify the reason of symptoms aggravation, drug lymphocyte stimulation test and rechallenge trial with mesalamine were performed. The results indicated the possibility that aggravation was related to allergic reaction and was dose-dependent. Furthermore, we examined colonoscopic views but there was no remarkable change in before and after rechallenge trial. Based on the above result, the patient was diagnosed with mesalamine intolerance. In order to differentiate whether the exacerbation of bloody diarrhea is due to the side effects of the mesalamine or a true relapse of UC, taking careful history before and after increasing mesalamine dosage as well as being aware of side effects of mesalamine are required. Clinicians should be aware of diarrhea as a side effect of mesalamine particularly after onset of mesalamine formulation, change in mesalamine formulation, or change in mesalamine dose.

A Phase I Trial of Arterial Infusion Chemotherapy with Gemcitabine and 5-Fluorouracil for Unresectable Advanced Pancreatic Cancer after Vascular Supply Distribution via Superselective Embolization

BACKGROUND We previously reported that arterial infusion chemotherapy improved the response rate and survival of the patients with pancreatic cancer at advanced stages in an open trial. We conducted a Phase I trial of arterial infusion chemotherapy with gemcitabine and 5-fluorouracil for advanced pancreatic cancer after vascular supply distribution via superselective embolization. METHODS Patients were treated after arterial embolization for hemodynamic change to restrict the blood flow into the pancreas (mainly to the great pancreatic artery and the caudal pancreatic artery). Arterial infusion chemotherapy consisted of gemcitabine in doses that were increased from 600 to 1000 mg/m(2) in subsequent cohorts on Day 1 plus continuous infusion of 5-fluorouracil 300 mg/m(2)/day on Days 1-5 every 2 weeks. Result Twelve patients were enrolled. The maximum tolerated dose of gemcitabine was determined to be Level 3 (1000 mg/m(2)). Only very mild hematological and non-hematological toxicities were noted. The overall response rate was 33.3%. The median survival time was 22.7 (95% CI; 9.5-24.5) months and the 1- and 2-year overall survival rates were 83.3 and 25.0%, respectively. CONCLUSION Arterial infusion chemotherapy using 1000 mg/m(2) gemcitabine on Day 1 and 300 mg/m(2)/day 5-fluorouracil on Days 1-5 every 2 weeks warrants a Phase II study.

Advanced pancreatic carcinoma showing a complete response to arterial infusion chemotherapy

We report a patient with advanced carcinoma of the pancreatic body and tail with multiple liver metastases who showed a complete response to hepatic and splenic arterial infusion chemotherapy (HSAIC) with gemcitabine and 5-fluorouracil, following transcatheter peripancreatic arterial embolization (TPPAE) and partial splenic embolization (PSE). Nonresectable advanced pancreatic carcinoma tends to have a low response to medical treatment, with the median survival time being 6 months or less for stage IV cases. We disclose herein that the median survival time of patients receiving HSAIC after TPPAE is more than three times longer than the survival time attained with conventional treatments. However, in patients with advanced carcinoma of the pancreatic tail, for which TTPAE is not applicable, survival times remain low. Thus, in the patient described here, we also performed embolization of the left gastric and short gastric arteries as well as PSE to increase the flow within the great pancreatic and caudal pancreatic arteries via the splenic artery, and gemcitabine and 5-fluorouracil were administered via the splenic artery. As a result of these procedures, marked reduction in the advanced carcinoma of the pancreatic body and tail and of liver metastases was attained.

Laparoscopic-endoscopic cooperative surgery is a safe and effective treatment for superficial nonampullary duodenal tumors

The use of endoscopic submucosal dissection (ESD) for duodenal neoplasms has increased in recent years, but delayed perforation and bleeding are also known to frequently occur. We present two cases in which duodenal adenoma was successfully treated with laparoscopic‐endoscopic cooperative surgery. ESD was combined with laparoscopic seromuscular sutures. The lesions in both cases were located in the second portion of the duodenum. The patients requested resection of the lesion, and we performed laparoscopic‐endoscopic cooperative surgery. After the laparoscopic surgeon mobilized the duodenum, the endoscopic surgeon performed ESD for the duodenal tumor without perforation. The laparoscopic surgeon sutured the duodenal wall in the seromuscular layer to strengthen the ulcer bed after ESD. Histopathological studies confirmed that the surgical margins were tumor‐free in both cases. The patients were discharged with no complications. This unique laparoscopic‐endoscopic cooperative procedure is a safe and effective method for resecting superficial nonampullary duodenal tumors.

Human infection by acanthocephalan parasites belonging to the genus Corynosoma found from small bowel endoscopy.

A 73-year-old man with a suspected ileus in January 2013 and subsequently suffered melena in February 2014 was endoscopically examined. As a result of the examinations, unidentified species of Corynosoma sp. and Corynosoma villosum were recovered from the small intestine, further endoscopic diagnosis suggested relevance between abdominal pain and the present infections in the small intestine. The recovered worms were composed of gravid females with developed eggs, suggesting that these parasites can survive for a long time in the intestine after infection. In this case, the short interval between infections appears to be due to the individuals eating habits which consist of regularly consuming uncooked seafood.

Abstract 1065: Regulation of autophagy and MAPK signaling by glutathione S-transferase-π in KRAS mutated cancer cells

Background: Mutated KRAS (mKRAS) is the most commonly found oncogene which may play a role in carcinogenesis at relatively early stage. Although Raf-1/MEK/ERK pathway has been well characterized as a proliferative signal at downstream of mKRAS, other signals related to senescence or apoptosis were also found at downstream of mKRAS and thus mKRAS function is still needed to be explored at a molecular level. We have hitherto disclosed a close correlation of GST-π expression and mKRAS in cancer tissues, induction of GST-π expression by KRAS transfection (Gastroenterology121:865-874, 2001) and significant reduction of mKRAS positive colon polyps in GST-π KO mice (unpublished observation), suggesting some cooperative function of GST-π for mKRAS. Objective: In the present study, we attempted to clarify the molecular mechanisms of the cooperative function of GST-π for mKRAS activity in tumor cells. Results: We first intended to confirm the correlation between GST-π expression and KRAS mutation in various human cancer cell lines by Western blotting. Those cells with no mKRAS (HepG2, HeLa, MCF7) were negative for GST-π expression while mKRAS-expressing cells (PANC-1, A549, M7609) were positive for GST-π. We then using M7609 cells, examined the effect of GST-π siRNA on their growth and found a significant impairment of cell proliferation. When GST-π KD M7609 cells were examined under electron microscopy, 40% of cells showed intracytoplasmic vesicles. The vesicles were positive for LC3 punctate signal by immunofluorescent staining and showed a characteristic feature of autophagosome, an organelle enclosed by a double lipid bilayer, by electron microscopy. In these cells, phosphorylation of Akt and EGFR was substantially reduced. Since interaction of GST-π with EGFR has been reported, it is plausible that this interaction positively regulates EGFR/class I PI3K/Akt/mTOR signal which in turn inhibits autophagy formation. Another mKRAS related signal: Raf-1/MEK/ERK was also explored in its relation to GST-π function. In M7609 KD cells, both Raf-1 protein and p-Raf-1 (S338), active form of Raf-1, were significantly reduced and phosphorylation of S621 residue which is known to play a crucial role in prevention of ubiquitination of Raf-1 protein was suppressed. In these cells, phosphorylation of downstream kinases of MEK and ERK were also reduced though their total protein levels were unchanged. When the cells were treated with proteasome inhibitor (MG132), recovery of Raf-1 and p-Raf-1 protein was observed. In parental M7609 cells, GST-π was found to be preferentially bound to p-Raf-1 on co-immunoprecipitation analysis, indicating that GST-π forms complex with Raf-1to prevent the Raf-1 from proteasomal degradation. Conclusion: GST-π, on one hand prevents the cancer cells from autophagy formation and on the other hand, stimulates Raf-1/MEK/ERK signal by stabilizing Raf-1 protein in mKRAS cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1065. doi:10.1158/1538-7445.AM2011-1065