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Distal tubular adenomas found on screening sigmoidoscopy are not markers for proximal advanced neoplasia
Gastrointestinal Oncology A635
April 1998 (120, 240 uM) and 5-FU (50, 100 uM) and ceils were harvested and prepared for flow cytometry at intervals of 24, 48, 72 hrs. Analysis of the cell cycle was performed on DNA histograms generated by FACS analysis using propidium iodide. The sub G0/1 cell population was regarded as apoptotic and this was confirmed using the TUNEL assay. Results. Incubation with SS increased the fraction of cells in G0/1 and decreased the fraction of cells in S-phase (compared to vehicle), consistent with G1 arrest. In contrast, 5-FU treatment produced a significant accumulation of cells in S-phase consistent with its known inhibition of DNA synthesis. Both drugs induced apoptosis in SW480, as well as HT-29, cells in a dose-dependent manner. In conclusion, these results indicate that the pattern of cell cycle inhibition induced by SS is distinct from that of 5-FU, reflecting a different intracellular target. Both drugs appear to activate a common pathway of cell death which may contribute to their anti-tumor effects. The presence of a p53 mutation in SW480 cells indicates that SS and 5-FU induce apoptosis independently of p53. • G2614 DISTAL TUBULAR ADENOMAS FOUND ON SCREENING SIGMOIDOSCOPY ARE NOT MARKERS FOR PROXIMAL ADVANCED NEOPLASIA. T. Levin, J. Selby, S. Grossman. C. Conell, L. Finkler, M. Lawson, A. Palitz. Kaiser Permanente Medical Care Program, Oakland, CA, and Division of Gastroenterology,University of California, San Francisco. PURPOSE: To establish the characteristics of distal colonic neoplasms that make them markers for proximal advanced neoplasia [defined as: tubular adenoma (TA) >-- 1 cm in size, presence of any villous features, severe dysplasia, or carcinoma] in patients undergoing screening sigmoidoscopy in a large HMO. METHODS: In this prospective study, procedure reports from 72,527 screening sigmoidoscopies and 2,944 follow-up colonoscopies were prospectively collected, linked to pathology results, and verified by chart review, when necessary. All subjects were average-risk, asymptomatic, and aged > 50 years old and were screened as part of the Colorectal Cancer Prevention (CoCaP) screening program of Northern California Kaiser Permanente. None had a prior history of colorectal cancer or polyps, nor a family history of colorectal cancer worse than a single first degree relative older than 55 at diagnosis. RESULTS: Neither the presence of a distal TA, nor its size were predictors of proximal advanced neoplasia. However, in a logistic regression model, the presence of any villous features in the distal polyp doubled the odds of proximal advanced neoplasia [OR: 2.3 (1.75, 3.05)]. Among patients over 70, the prevalence of proximal advanced neoplasia was nearly three times that of younger patients. Low risk family history conferred a small, statistically insignificant, increased risk of advanced proximal neeplasia. Sigmoidoscopy Finding
n
Proximal Advanced Neoplasia, % (CI)
No Adenoma
515
5.2 (3.3, 7.2)
>lTA
764
5.5(3.9,7.1)
> 1 TA -> 1 cm
850
5.7 (4.1, 7.2)
Villous Features
815
12.2 (9.9, 14.4)
CONCLUSIONS: The risk of proximal advanced neoplasia in an average risk subject is not inconsequential. However, the risk is the same in a subject with a negative screening sigmoidoscopy as in one with a distal TA. These findings support a strategy of colonoscopy only if a polyp with villous or more advanced features is found on screening sigmoidoscopy in the sixth decade of life. The increase in proximal findings with age suggest the consideration of screening colonoscopy later, perhaps in the latter part of the seventh decade. Dr. Levin is supported by an ADHF Outcomes Research Training Award. • G2615 REG I PROTEIN LOCALIZATION IN COLONIC MUCOSA AT RISK FOR NEOPLASIA. ~ , J Kilcullen, JJ Steinberg, and ME Zenilman. Departments of Surgery and Pathology, Albert Einstein College of Medicine, Montefiore Medical Center, Bronx, N.Y. Pancreatic regenerating gene (reg I) messenger RNA is overexprassed within the pancreas following injury, and its expression corresponds to cellular dedifferentiation. It is rarely expressed in normal colon tissue. While only 50% of colon cancers express reg, 100% of the transition zones between normal and neoplastic colonic tissue ectopically express reg. In this study we mapped the expression of reg in colorectal mucosa at high risk for malignant transformation. We retrospectively evaluated colonic tissue from 14 patients who underwent total colectomy, 11 for familial adenomatous polyposis (FAP), and from 4 for ulcerative colitis (UC). Paraffin-fixed tissue was immnnohistochemically stained with a monoclonal antibody to human reg, mabD4, and examined using light microscopy. Of the 11 FAP patients, 6 patients were male and 5 were female. Their ages ranged from 16 to 55 years (mean=33, median = 35 years). Of the UC patients, 2 were male and 2 were female. Their ages ranged from 45 to 71 (mean=61). Multiple adenomas were seen in the FAP tissue, but no dysplastic changes were seen on any of
the UC specimens. While all normal, hamartomatous and hyperplastic polyp tissue were negative for reg, all adenomatous polyps and adenomatous tissue stained positive for reg. By contrast, only one of the 4 UC patients' tissue was positive for reg. We conclude that reg protein is ectopically expressed in adenomatous colonic mucosa, a n d not in normal, hyperplastic or inflammatory (UC) colonic mucosa. It is not expressed in all colonic tissue at risk for neoplasia. These data suggest that reg I expression in colon tissue is timed close to the development of adenomatous change. G2616 REGRESSION OF LOCALIZED LOW GRADE MALT GASTRIC LYMPHOMA UNDER ANTI-HELICOBACTER PYLORI TREATMENT. M L6vy (1), C Traull6 (2), C Copie-Bergman (1), JC Delchier (1) and le Groupe d'Etudes des Lymphomes de l'Adulte (GELA). (1) HSpital Henri Mondor, 94000 Cr6teil, (2) H6pital Nord, 80054 Amiens, France.
Helicobacter pylori (Hp) eradication may induce remission of low grade MALT gastric lymphoma (MGL). The aim of this work was to study prospectively the influence of anti-Hp treatment on localized types of MGL and the predictive factors on tumoral response. Patients: 25 patients with localized MGL (stage IE) and infected with Hp were included. Histological study showed score 5 lesions in the Wotherspoon's classification in all cases. The treatment consisted in a 7-day tri-therapy. Tumoral response was evaluated after 7 months (2 to 12) by gastroscopy (G) and endoscopic ultrasonography (EUS). Median follow-up was 11 months (2 to 25). Complete remission (CR) was defined by disappearance of endoscopic and histological lesions and partial remission (PR) by regression of endoscopic lesions and persistence of histological abnormalities but decrease of the Wotherspoon score. Results: n
Hp
CR
PR
Persistent disease 4 2 1 5 l 5
Eradicated 22 11 7 Not eradicated 31 1 0 G Gastritis 102 5 4 Large ulcerated folds 15 7 3 EUS Infiltration < 5 mm 13 6 6 Infiltration > 5 mm 10 5 0 (6 to 17 ram) 1These patients remained infected despite 3 different treatments. 2 EUS was not performed in two of these patients.
Conclusion: 1) Anti-Hp treatment in patients with localized low grade MGL induced a tumoral response in 76% with a CR in 48% at 7 months. 2) Factors of treatment failure were Hp eradication failures, large ulcerated folds and tumoral infiltration > 5 mm. • G2617 PROTEIN KINASE C 8 DOWN-REGULATION MEDIATES APOPTOSIS IN THE ADENOCARCINOMA CELL LINE C O L t 205. AE Lewis. MJS Langman, BCY Wong, D Watters*, MC Eggo; Dept of Medicine, University of Birmingham, B 1 5 2 T H , UK and *Queensland Institute of Medical Research, Brisbane, Australia. Background Apoptosis results in the progressive and complete selfdestruction of targeted cells and is essential for normal homeostasis in the colon. Various signalling pathways are involved in this process, including that of protein kinase C (PKC). The effects induced by PKC activators can either be due to PKC activation or to PKC inactivation/down-regulation which succeeds activation. We have examined the role of activation/down regulation of protein kinase C 8, one of the eleven PKC isozymes, in apoptosis in C O L t 205 cells. We used bistratene A (Bis A), a marine polyether which is reported to selectively activate PKC8 (Watters et al (1996) Exp Cell Res 22_99 327-335) and rottlerin, a compound from Mallotus phillippinensis (Gschwendt et al (1994) BBRC 19993-98) which is reported to specifically inhibit PKC5 at ~M concentrations. Methods COLt 205 cells were incubated in serum-free medium for 24h and treated with Bis A (1-100ng/ml) and/or rottlerin (10-8-104M) for a further 24h. Apoptosis was assessed by i) cell cycle analysis using fluorescence activated cell sorting of propidium iodide-labeled cells and by ii) DNA fragmentation. Results Bis A increased the apoptotic fraction to 22.6 ± 2.5% (n=3 + SD) compared with control values of 10.5 -+2.5%. The maximum effect was seen at 10ng/ml. Rottlerin at concentrations which specifically inhibit PKC8 (3-6~tM) produced an increase in apoptosis of 24.9 + 0.1%. Rottlerin was not able to block the effects of Bis A and their effects on apoptosis were additive. Conclusions PKC8 activation with Bis A and inhibition with rottlerin induce apoptosis in COLt 205 cells. Since PKC inhibition with rottlerin is not able to prevent Bis A effects and is additive with Bis we conclude that PKC8 down-regulation mediates apoptosis in these ceils.
April 1998 (120, 240 uM) and 5-FU (50, 100 uM) and ceils were harvested and prepared for flow cytometry at intervals of 24, 48, 72 hrs. Analysis of the cell cycle was performed on DNA histograms generated by FACS analysis using propidium iodide. The sub G0/1 cell population was regarded as apoptotic and this was confirmed using the TUNEL assay. Results. Incubation with SS increased the fraction of cells in G0/1 and decreased the fraction of cells in S-phase (compared to vehicle), consistent with G1 arrest. In contrast, 5-FU treatment produced a significant accumulation of cells in S-phase consistent with its known inhibition of DNA synthesis. Both drugs induced apoptosis in SW480, as well as HT-29, cells in a dose-dependent manner. In conclusion, these results indicate that the pattern of cell cycle inhibition induced by SS is distinct from that of 5-FU, reflecting a different intracellular target. Both drugs appear to activate a common pathway of cell death which may contribute to their anti-tumor effects. The presence of a p53 mutation in SW480 cells indicates that SS and 5-FU induce apoptosis independently of p53. • G2614 DISTAL TUBULAR ADENOMAS FOUND ON SCREENING SIGMOIDOSCOPY ARE NOT MARKERS FOR PROXIMAL ADVANCED NEOPLASIA. T. Levin, J. Selby, S. Grossman. C. Conell, L. Finkler, M. Lawson, A. Palitz. Kaiser Permanente Medical Care Program, Oakland, CA, and Division of Gastroenterology,University of California, San Francisco. PURPOSE: To establish the characteristics of distal colonic neoplasms that make them markers for proximal advanced neoplasia [defined as: tubular adenoma (TA) >-- 1 cm in size, presence of any villous features, severe dysplasia, or carcinoma] in patients undergoing screening sigmoidoscopy in a large HMO. METHODS: In this prospective study, procedure reports from 72,527 screening sigmoidoscopies and 2,944 follow-up colonoscopies were prospectively collected, linked to pathology results, and verified by chart review, when necessary. All subjects were average-risk, asymptomatic, and aged > 50 years old and were screened as part of the Colorectal Cancer Prevention (CoCaP) screening program of Northern California Kaiser Permanente. None had a prior history of colorectal cancer or polyps, nor a family history of colorectal cancer worse than a single first degree relative older than 55 at diagnosis. RESULTS: Neither the presence of a distal TA, nor its size were predictors of proximal advanced neoplasia. However, in a logistic regression model, the presence of any villous features in the distal polyp doubled the odds of proximal advanced neoplasia [OR: 2.3 (1.75, 3.05)]. Among patients over 70, the prevalence of proximal advanced neoplasia was nearly three times that of younger patients. Low risk family history conferred a small, statistically insignificant, increased risk of advanced proximal neeplasia. Sigmoidoscopy Finding
n
Proximal Advanced Neoplasia, % (CI)
No Adenoma
515
5.2 (3.3, 7.2)
>lTA
764
5.5(3.9,7.1)
> 1 TA -> 1 cm
850
5.7 (4.1, 7.2)
Villous Features
815
12.2 (9.9, 14.4)
CONCLUSIONS: The risk of proximal advanced neoplasia in an average risk subject is not inconsequential. However, the risk is the same in a subject with a negative screening sigmoidoscopy as in one with a distal TA. These findings support a strategy of colonoscopy only if a polyp with villous or more advanced features is found on screening sigmoidoscopy in the sixth decade of life. The increase in proximal findings with age suggest the consideration of screening colonoscopy later, perhaps in the latter part of the seventh decade. Dr. Levin is supported by an ADHF Outcomes Research Training Award. • G2615 REG I PROTEIN LOCALIZATION IN COLONIC MUCOSA AT RISK FOR NEOPLASIA. ~ , J Kilcullen, JJ Steinberg, and ME Zenilman. Departments of Surgery and Pathology, Albert Einstein College of Medicine, Montefiore Medical Center, Bronx, N.Y. Pancreatic regenerating gene (reg I) messenger RNA is overexprassed within the pancreas following injury, and its expression corresponds to cellular dedifferentiation. It is rarely expressed in normal colon tissue. While only 50% of colon cancers express reg, 100% of the transition zones between normal and neoplastic colonic tissue ectopically express reg. In this study we mapped the expression of reg in colorectal mucosa at high risk for malignant transformation. We retrospectively evaluated colonic tissue from 14 patients who underwent total colectomy, 11 for familial adenomatous polyposis (FAP), and from 4 for ulcerative colitis (UC). Paraffin-fixed tissue was immnnohistochemically stained with a monoclonal antibody to human reg, mabD4, and examined using light microscopy. Of the 11 FAP patients, 6 patients were male and 5 were female. Their ages ranged from 16 to 55 years (mean=33, median = 35 years). Of the UC patients, 2 were male and 2 were female. Their ages ranged from 45 to 71 (mean=61). Multiple adenomas were seen in the FAP tissue, but no dysplastic changes were seen on any of
the UC specimens. While all normal, hamartomatous and hyperplastic polyp tissue were negative for reg, all adenomatous polyps and adenomatous tissue stained positive for reg. By contrast, only one of the 4 UC patients' tissue was positive for reg. We conclude that reg protein is ectopically expressed in adenomatous colonic mucosa, a n d not in normal, hyperplastic or inflammatory (UC) colonic mucosa. It is not expressed in all colonic tissue at risk for neoplasia. These data suggest that reg I expression in colon tissue is timed close to the development of adenomatous change. G2616 REGRESSION OF LOCALIZED LOW GRADE MALT GASTRIC LYMPHOMA UNDER ANTI-HELICOBACTER PYLORI TREATMENT. M L6vy (1), C Traull6 (2), C Copie-Bergman (1), JC Delchier (1) and le Groupe d'Etudes des Lymphomes de l'Adulte (GELA). (1) HSpital Henri Mondor, 94000 Cr6teil, (2) H6pital Nord, 80054 Amiens, France.
Helicobacter pylori (Hp) eradication may induce remission of low grade MALT gastric lymphoma (MGL). The aim of this work was to study prospectively the influence of anti-Hp treatment on localized types of MGL and the predictive factors on tumoral response. Patients: 25 patients with localized MGL (stage IE) and infected with Hp were included. Histological study showed score 5 lesions in the Wotherspoon's classification in all cases. The treatment consisted in a 7-day tri-therapy. Tumoral response was evaluated after 7 months (2 to 12) by gastroscopy (G) and endoscopic ultrasonography (EUS). Median follow-up was 11 months (2 to 25). Complete remission (CR) was defined by disappearance of endoscopic and histological lesions and partial remission (PR) by regression of endoscopic lesions and persistence of histological abnormalities but decrease of the Wotherspoon score. Results: n
Hp
CR
PR
Persistent disease 4 2 1 5 l 5
Eradicated 22 11 7 Not eradicated 31 1 0 G Gastritis 102 5 4 Large ulcerated folds 15 7 3 EUS Infiltration < 5 mm 13 6 6 Infiltration > 5 mm 10 5 0 (6 to 17 ram) 1These patients remained infected despite 3 different treatments. 2 EUS was not performed in two of these patients.
Conclusion: 1) Anti-Hp treatment in patients with localized low grade MGL induced a tumoral response in 76% with a CR in 48% at 7 months. 2) Factors of treatment failure were Hp eradication failures, large ulcerated folds and tumoral infiltration > 5 mm. • G2617 PROTEIN KINASE C 8 DOWN-REGULATION MEDIATES APOPTOSIS IN THE ADENOCARCINOMA CELL LINE C O L t 205. AE Lewis. MJS Langman, BCY Wong, D Watters*, MC Eggo; Dept of Medicine, University of Birmingham, B 1 5 2 T H , UK and *Queensland Institute of Medical Research, Brisbane, Australia. Background Apoptosis results in the progressive and complete selfdestruction of targeted cells and is essential for normal homeostasis in the colon. Various signalling pathways are involved in this process, including that of protein kinase C (PKC). The effects induced by PKC activators can either be due to PKC activation or to PKC inactivation/down-regulation which succeeds activation. We have examined the role of activation/down regulation of protein kinase C 8, one of the eleven PKC isozymes, in apoptosis in C O L t 205 cells. We used bistratene A (Bis A), a marine polyether which is reported to selectively activate PKC8 (Watters et al (1996) Exp Cell Res 22_99 327-335) and rottlerin, a compound from Mallotus phillippinensis (Gschwendt et al (1994) BBRC 19993-98) which is reported to specifically inhibit PKC5 at ~M concentrations. Methods COLt 205 cells were incubated in serum-free medium for 24h and treated with Bis A (1-100ng/ml) and/or rottlerin (10-8-104M) for a further 24h. Apoptosis was assessed by i) cell cycle analysis using fluorescence activated cell sorting of propidium iodide-labeled cells and by ii) DNA fragmentation. Results Bis A increased the apoptotic fraction to 22.6 ± 2.5% (n=3 + SD) compared with control values of 10.5 -+2.5%. The maximum effect was seen at 10ng/ml. Rottlerin at concentrations which specifically inhibit PKC8 (3-6~tM) produced an increase in apoptosis of 24.9 + 0.1%. Rottlerin was not able to block the effects of Bis A and their effects on apoptosis were additive. Conclusions PKC8 activation with Bis A and inhibition with rottlerin induce apoptosis in COLt 205 cells. Since PKC inhibition with rottlerin is not able to prevent Bis A effects and is additive with Bis we conclude that PKC8 down-regulation mediates apoptosis in these ceils.