- Email: [email protected]
Sa1278 Bortezomib Restores Defective Apoptosis by Upregulation of Noxa in Enteropathy-Associated T-Cell Lymphoma
AGA Abstracts
Sa1278 Bortezomib Restores Defective Apoptosis by Upregulation of Noxa in Enteropathy-Associated T-Cell Lymphoma Laura R. de Baaij, Marijn Radersma, Jolanda M. van de Water, Nathalie J. Hijmering, Laura Moesbergen, Otto Visser, Joost J. Oudejans, Chris J. Mulder, Chris J. Meijer, Saskia A. Cillessen Enteropathy-associated T-cell lymphoma (EATL) is a rare intestinal lymphoma that arises from intraepithelial lymphocytes. Clinical outcome of patients with EATL is very poor, due to chemotherapy-resistance and high relapse rates. Therefore, new therapeutic options for EATL are urgently needed. Many studies in other types of lymphoma have shown that inhibition of apoptosis may cause chemotherapy-resistance and that restoration of defective apoptosis can induce cell death in these lymphomas. Preliminary data in EATL samples have demonstrated an increased expression of a fraction of NF- κB target genes, suggesting upregulation of NF-κB activity in EATL tumor cells. The proteasome inhibitor bortezomib inhibits NF-κB activity and can induce apoptosis via upregulation of the pro-apoptotic BH3only protein Noxa. In the present study, we evaluated if the intrinsic apoptosis pathway is disrupted in EATL and if bortezomib can restore apoptosis in cultured EATL cells. Lasercapture microdissection was applied to 19 frozen EATL samples to obtain purified tumor cells for RNA isolation. Intraepithelial lymphocytes (IEL) of healthy controls were obtained from fresh duodenal biopsies and isolated by FACS cell sorting. RT-MLPA analysis revealed that expression of the pro-apoptotic BH3-only gene Noxa was significantly downregulated in EATL cells compared to healthy donor IEL. Treatment with bortezomib resulted in induction of apoptosis in all EATL samples tested. The lethal dose (LD50) varied between 5 nM and 15 nM after 36 and 48 hours of incubation. Bortezomib-induced cell death in EATL cells was caspase-9 mediated. mRNA and protein expression analyses showed upregulation of Noxa after incubation with bortezomib. Downregulation of Noxa using siRNA analysis decreased bortezomib-induced apoptosis in EATL cells. In conclusion, our study showed that bortezomib induces apoptosis by upregulation of Noxa in EATL cells. Therefore, bortezomib should be considered as a potential drug in the treatment of patients with EATL to improve their prognosis.
Sa1280 Adult-Onset Autoimmune Enteropathy: An European Single Center Experience Roy L. van Wanrooij, Chris J. Mulder, Andra Neefjes-Borst, Marco Schreurs, Boudewina M. von Blomberg, Gerd Bouma Background & Aims: Autoimmune enteropathy (AIE) is a very rare cause of chronic adultonset diarrhea and occurs as the consequence of a diffuse autoimmune disorder of the gastrointestinal system. AIE is related to the immunodysregulation polyendocrinopathy enteropathy X-linked (IPEX)-syndrome that occurs in infants. Very few data on the clinical characteristics, treatment strategies and prognosis exist. We here report these characteristics in a relatively large series of adult-onset AIE patients diagnosed at our institute. Material & Methods: Nine adults were diagnosed with AIE between 2005-2012. Diagnosis of adultonset AIE was based on the Akram-criteria: (1) Adult-onset diarrhoea for more than 6 weeks (2) Malabsortion (3) Compatible histological abnormalities of the duodenum (4) Exclusion of other causes of villous atrophy (5) Anti-enterocyte or anti-goblet antibodies. Clinical, histological and serological characteristics as well as treatment response were reported. Results: All patients were Caucasians, seven female, with a median age at diagnosis of 52 years (range 23 to 78 years). Median time since onset of symptoms to time of diagnosis was 3 months. One patient was previously diagnosed with type 1 diabetes and two with celiac disease. Histological evaluation revealed total villous atrophy in 4 patients, while in 2 and 3 patients subtotal and partial villous atrophy was noted, respectively. Epithelial lymphocytosis was observed in 5/9 patients. No significant aberrant (cytCD3- sCD3+) intraepithelial lymphocyte population was identified in any of the patients. Anti-enterocyte antibodies were found in 8/9 patients, and in one patient anti-goblet cells were observed. Aspecific inflammation was seen in the colon of 5 patients, while in all patients a chronic helicobacter-negative gastric inflammation was observed, with anti-parietal cell antibodies being positive in 3 patients. At least one serological autoimmune-associated antibody was found in every patient. Seven patients required parenteral feeding. All patients received steroids for induction therapy. For maintenance therapy, rituximab failed in one out of two patients while this patient subsequently did respond to 6-thioguanine. Three other patients treated with 6-mercaptopurine or 6-thioguanine showed clinical, histological and serological response with a median follow-up of 3 years, while one patient showed a partial response. In addition, a severely ill 23 year aged patient successfully underwent autologous stem cell transplantation. Conclusions: Adult onset AID is a rare disease that in our center mainly affects females of varying age. This data provides strong support that AIE should be considered as a diffuse gastrointestinal autoimmune disorder with systemic immune-dysregulation. Furthermore, 6-thioguanine showed promising results in the treatment of adult-onset AIE.
Sa1279 "Non Celiac Gluten Sensitivity" (NCGS) Is Uncommon in Patients Spontaneously Adhering to Gluten Free Diet (GFD) Without Medical Necessity Barbara Zanini, Roberta Baschè, Alice Ferraresi, Chiara Ricci, Francesco Lanzarotto, Alberto Lanzini Background. Adverse gastrointestinal and extra intestinal reaction to gluten is reported by many patients that spontaneously adhere to a gluten free diet (GFD) without medical necessity because of exclusion of celiac disease (CD). It is unknown how many of these patients fulfil the criteria for NCGS, a still poorly defined clinical entity of uncertain pathogenesis. Aims. To assess gluten dependence of symptoms (NCGS) in patients spontaneously adhering to GFD for self diagnosed adverse reaction to gluten. Methods. Patients spontaneously adherent to GFD without medical necessity referred to our Clinic in years 2008-2011 were recalled and randomized to a double blind cross over study involving, while continuing GFD, daily administration of 10 g gluten Vs 10 g gluten free flour for 10 days each with 2 weeks washout in between. Endpoints: ability to clearly identify the gluten phase of the study, changes of Gastrointestinal Symptoms Rating Scale (GSRS) and of visual analogue fatigue scale, changes of antigliadin (AGA) IgA/IgG, anti tissue transglutaminase (t-TG) IgA and antiendomysial (EMA) IgA/IgG antibodies. Results. Out of 86 patients recalled, 64 agreed to followup visit. On review 45 patients persisting on GFD fulfilled exclusion criteria for CD (no villous atrophy and negative CD related serology while on gluten containing diet). 26 patients (age 42 + 9 years, M/F = 2/24, 11 HLA DQ2/8 positive, 13 negative, 2 unknown, all t-TG, EMA and AGA negative) signed the informed consent and were randomized. The gluten phase of the study was correctly, incorrectly, or not clearly identified by 7, 11 and 5 patients, respectively. In the 7 patients recognizing the gluten phase (3 with HLA DQ2/8), scores for the 5 dimensions of GSRS, but not those for fatigue scale, were higher during the gluten compared with no gluten phase (figure) and with baseline values (not shown). Scores were similar in the other patients not recognizing the gluten phase. No change in AGA, t-TG and EMA level was observed at any time point (serology currently available in 12 patients). Conclusions. NCGS is diagnosed by double blind challenge in a minority only of patients with self reported adverse reaction to gluten spontaneously adhering to GFD. Components other than gluten, such as fermentable carbohydrates of wheat, may potentially account for symptoms in patients testing negative at gluten challenge than may suffer of non-celiac wheat sensitivity.
AGA Abstracts
Sa1281 Long Term Adherence to a Gluten Free Diet Javier A. Villafuerte-Galvez, Rupa Mukherjee, Rohini R. Vanga, Toufic A. Kabbani, Melinda Dennis, Joshua Hansen, Ciaran P. Kelly, Daniel A. Leffler Background: The gluten free diet (GFD) is the mainstay of therapy for celiac disease, yet there is a paucity of studies addressing long term GFD adherence in the literature. Methods: A survey was sent to 777 individuals who had been diagnosed with biopsy confirmed celiac disease ≥ 5 years previously. The survey included the validated Celiac Dietary Adherence Test (CDAT) and items assessing demographics and clinical variables. Demographics of respondents were compared to non-respondents. All other variables were analyzed using the CDAT score as the dependent variable. Results: At the time of analysis 351 (45.2%) patients had responded. The mean age of the respondents vs. non-respondents was marginally higher (53.4±15.4 vs. 51.1±16.4, p=0.043). The gender, state or region of residence did not differ significantly between the two groups. The median time on a GFD for responders was 8 years (Range 5 to 54 years). The mean age at diagnosis was 41.6±15.0 years. Demographics of the respondent population were as follows: 78.1% female, 70.2% married, 88.5% ≥ college educated and 96.3% resided in New England. The median CDAT score was 10 (Range 7-30), where a higher score indicates worse adherence and 31.1% had a CDAT score of ≥ 13, consistent with inadequate adherence. We found no difference in median CDAT scores by gender (p=0.36), marital status (p=0.70), or state of residence (p=0.46). There were statistically significant correlations between better GFD adherence as measured by CDAT score and level of education ( ρ=-0.123, p=0.025), respondent's perceived long term GFD adherence (ρ=-0.358, p,0.001), perception of knowledge of the GFD ( ρ=-0.348, p,0.0001) and with the perception that cost of the GFD is an obstacle to adherence ( ρ= 0.211, p,0.001). Those who perceived that a GFD is unhelpful (n=44, 12.5%) had a higher median CDAT score than those who perceive it as helpful (12±7 vs. 10±2, p=0.013).
S-250
Sa1278 Bortezomib Restores Defective Apoptosis by Upregulation of Noxa in Enteropathy-Associated T-Cell Lymphoma Laura R. de Baaij, Marijn Radersma, Jolanda M. van de Water, Nathalie J. Hijmering, Laura Moesbergen, Otto Visser, Joost J. Oudejans, Chris J. Mulder, Chris J. Meijer, Saskia A. Cillessen Enteropathy-associated T-cell lymphoma (EATL) is a rare intestinal lymphoma that arises from intraepithelial lymphocytes. Clinical outcome of patients with EATL is very poor, due to chemotherapy-resistance and high relapse rates. Therefore, new therapeutic options for EATL are urgently needed. Many studies in other types of lymphoma have shown that inhibition of apoptosis may cause chemotherapy-resistance and that restoration of defective apoptosis can induce cell death in these lymphomas. Preliminary data in EATL samples have demonstrated an increased expression of a fraction of NF- κB target genes, suggesting upregulation of NF-κB activity in EATL tumor cells. The proteasome inhibitor bortezomib inhibits NF-κB activity and can induce apoptosis via upregulation of the pro-apoptotic BH3only protein Noxa. In the present study, we evaluated if the intrinsic apoptosis pathway is disrupted in EATL and if bortezomib can restore apoptosis in cultured EATL cells. Lasercapture microdissection was applied to 19 frozen EATL samples to obtain purified tumor cells for RNA isolation. Intraepithelial lymphocytes (IEL) of healthy controls were obtained from fresh duodenal biopsies and isolated by FACS cell sorting. RT-MLPA analysis revealed that expression of the pro-apoptotic BH3-only gene Noxa was significantly downregulated in EATL cells compared to healthy donor IEL. Treatment with bortezomib resulted in induction of apoptosis in all EATL samples tested. The lethal dose (LD50) varied between 5 nM and 15 nM after 36 and 48 hours of incubation. Bortezomib-induced cell death in EATL cells was caspase-9 mediated. mRNA and protein expression analyses showed upregulation of Noxa after incubation with bortezomib. Downregulation of Noxa using siRNA analysis decreased bortezomib-induced apoptosis in EATL cells. In conclusion, our study showed that bortezomib induces apoptosis by upregulation of Noxa in EATL cells. Therefore, bortezomib should be considered as a potential drug in the treatment of patients with EATL to improve their prognosis.
Sa1280 Adult-Onset Autoimmune Enteropathy: An European Single Center Experience Roy L. van Wanrooij, Chris J. Mulder, Andra Neefjes-Borst, Marco Schreurs, Boudewina M. von Blomberg, Gerd Bouma Background & Aims: Autoimmune enteropathy (AIE) is a very rare cause of chronic adultonset diarrhea and occurs as the consequence of a diffuse autoimmune disorder of the gastrointestinal system. AIE is related to the immunodysregulation polyendocrinopathy enteropathy X-linked (IPEX)-syndrome that occurs in infants. Very few data on the clinical characteristics, treatment strategies and prognosis exist. We here report these characteristics in a relatively large series of adult-onset AIE patients diagnosed at our institute. Material & Methods: Nine adults were diagnosed with AIE between 2005-2012. Diagnosis of adultonset AIE was based on the Akram-criteria: (1) Adult-onset diarrhoea for more than 6 weeks (2) Malabsortion (3) Compatible histological abnormalities of the duodenum (4) Exclusion of other causes of villous atrophy (5) Anti-enterocyte or anti-goblet antibodies. Clinical, histological and serological characteristics as well as treatment response were reported. Results: All patients were Caucasians, seven female, with a median age at diagnosis of 52 years (range 23 to 78 years). Median time since onset of symptoms to time of diagnosis was 3 months. One patient was previously diagnosed with type 1 diabetes and two with celiac disease. Histological evaluation revealed total villous atrophy in 4 patients, while in 2 and 3 patients subtotal and partial villous atrophy was noted, respectively. Epithelial lymphocytosis was observed in 5/9 patients. No significant aberrant (cytCD3- sCD3+) intraepithelial lymphocyte population was identified in any of the patients. Anti-enterocyte antibodies were found in 8/9 patients, and in one patient anti-goblet cells were observed. Aspecific inflammation was seen in the colon of 5 patients, while in all patients a chronic helicobacter-negative gastric inflammation was observed, with anti-parietal cell antibodies being positive in 3 patients. At least one serological autoimmune-associated antibody was found in every patient. Seven patients required parenteral feeding. All patients received steroids for induction therapy. For maintenance therapy, rituximab failed in one out of two patients while this patient subsequently did respond to 6-thioguanine. Three other patients treated with 6-mercaptopurine or 6-thioguanine showed clinical, histological and serological response with a median follow-up of 3 years, while one patient showed a partial response. In addition, a severely ill 23 year aged patient successfully underwent autologous stem cell transplantation. Conclusions: Adult onset AID is a rare disease that in our center mainly affects females of varying age. This data provides strong support that AIE should be considered as a diffuse gastrointestinal autoimmune disorder with systemic immune-dysregulation. Furthermore, 6-thioguanine showed promising results in the treatment of adult-onset AIE.
Sa1279 "Non Celiac Gluten Sensitivity" (NCGS) Is Uncommon in Patients Spontaneously Adhering to Gluten Free Diet (GFD) Without Medical Necessity Barbara Zanini, Roberta Baschè, Alice Ferraresi, Chiara Ricci, Francesco Lanzarotto, Alberto Lanzini Background. Adverse gastrointestinal and extra intestinal reaction to gluten is reported by many patients that spontaneously adhere to a gluten free diet (GFD) without medical necessity because of exclusion of celiac disease (CD). It is unknown how many of these patients fulfil the criteria for NCGS, a still poorly defined clinical entity of uncertain pathogenesis. Aims. To assess gluten dependence of symptoms (NCGS) in patients spontaneously adhering to GFD for self diagnosed adverse reaction to gluten. Methods. Patients spontaneously adherent to GFD without medical necessity referred to our Clinic in years 2008-2011 were recalled and randomized to a double blind cross over study involving, while continuing GFD, daily administration of 10 g gluten Vs 10 g gluten free flour for 10 days each with 2 weeks washout in between. Endpoints: ability to clearly identify the gluten phase of the study, changes of Gastrointestinal Symptoms Rating Scale (GSRS) and of visual analogue fatigue scale, changes of antigliadin (AGA) IgA/IgG, anti tissue transglutaminase (t-TG) IgA and antiendomysial (EMA) IgA/IgG antibodies. Results. Out of 86 patients recalled, 64 agreed to followup visit. On review 45 patients persisting on GFD fulfilled exclusion criteria for CD (no villous atrophy and negative CD related serology while on gluten containing diet). 26 patients (age 42 + 9 years, M/F = 2/24, 11 HLA DQ2/8 positive, 13 negative, 2 unknown, all t-TG, EMA and AGA negative) signed the informed consent and were randomized. The gluten phase of the study was correctly, incorrectly, or not clearly identified by 7, 11 and 5 patients, respectively. In the 7 patients recognizing the gluten phase (3 with HLA DQ2/8), scores for the 5 dimensions of GSRS, but not those for fatigue scale, were higher during the gluten compared with no gluten phase (figure) and with baseline values (not shown). Scores were similar in the other patients not recognizing the gluten phase. No change in AGA, t-TG and EMA level was observed at any time point (serology currently available in 12 patients). Conclusions. NCGS is diagnosed by double blind challenge in a minority only of patients with self reported adverse reaction to gluten spontaneously adhering to GFD. Components other than gluten, such as fermentable carbohydrates of wheat, may potentially account for symptoms in patients testing negative at gluten challenge than may suffer of non-celiac wheat sensitivity.
AGA Abstracts
Sa1281 Long Term Adherence to a Gluten Free Diet Javier A. Villafuerte-Galvez, Rupa Mukherjee, Rohini R. Vanga, Toufic A. Kabbani, Melinda Dennis, Joshua Hansen, Ciaran P. Kelly, Daniel A. Leffler Background: The gluten free diet (GFD) is the mainstay of therapy for celiac disease, yet there is a paucity of studies addressing long term GFD adherence in the literature. Methods: A survey was sent to 777 individuals who had been diagnosed with biopsy confirmed celiac disease ≥ 5 years previously. The survey included the validated Celiac Dietary Adherence Test (CDAT) and items assessing demographics and clinical variables. Demographics of respondents were compared to non-respondents. All other variables were analyzed using the CDAT score as the dependent variable. Results: At the time of analysis 351 (45.2%) patients had responded. The mean age of the respondents vs. non-respondents was marginally higher (53.4±15.4 vs. 51.1±16.4, p=0.043). The gender, state or region of residence did not differ significantly between the two groups. The median time on a GFD for responders was 8 years (Range 5 to 54 years). The mean age at diagnosis was 41.6±15.0 years. Demographics of the respondent population were as follows: 78.1% female, 70.2% married, 88.5% ≥ college educated and 96.3% resided in New England. The median CDAT score was 10 (Range 7-30), where a higher score indicates worse adherence and 31.1% had a CDAT score of ≥ 13, consistent with inadequate adherence. We found no difference in median CDAT scores by gender (p=0.36), marital status (p=0.70), or state of residence (p=0.46). There were statistically significant correlations between better GFD adherence as measured by CDAT score and level of education ( ρ=-0.123, p=0.025), respondent's perceived long term GFD adherence (ρ=-0.358, p,0.001), perception of knowledge of the GFD ( ρ=-0.348, p,0.0001) and with the perception that cost of the GFD is an obstacle to adherence ( ρ= 0.211, p,0.001). Those who perceived that a GFD is unhelpful (n=44, 12.5%) had a higher median CDAT score than those who perceive it as helpful (12±7 vs. 10±2, p=0.013).
S-250