Is there a role for capsule endoscopy in the staging work-up of patients with gastric marginal zone B-cell lymphoma of MALT?

INTRODUCTION: In earlier studies, involvement of the small intestine was reported in patients with gastrointestinal lymphoma. Prospective data on the involvement of the small intestine in patients suffering from gastric extranodal MZBCL of MALT do not exist so far. In this study, we investigated the frequency of the involvement of the small intestine and the role of capsule endoscopy in patients with gastrointestinal extranodal MZBCL of MALT and of follicular lymphoma. PATIENTS AND METHODS: 40 consecutive patients with gastrointestinal extranodal MZBCL of MALT (26 men, 14 women, aged 27 - 80 years), and 7 patients with known follicular lymphoma of the small intestine (5 men, 2 women, aged 34 - 63 years) underwent capsule endoscopy. RESULTS: Involvement of the small intestine was identified by capsule endoscopy in all 7 patients with known follicular lymphoma of the small intestine. In 6 of 40 patients with gastric extranodal MZBCL of MALT abnormal findings could be observed, three of these findings indicative for lymphoma involvement of the small intestine. However, in each of these 3 cases, intestinal involvement had been already diagnosed by conventional GI endoscopy before capsule endoscopy. CONCLUSIONS: Capsule endoscopy is able to detect involvement of the small intestine in patients with gastrointestinal lymphoma. However, involvement of the small intestine seems to be rare in patients with gastric extranodal MZBCL of MALT. In summary, routine diagnostic work-up of the small intestine, e. g. by capsule endoscopy seems unnecessary because of the rare involvement of the small intestine and an excellent long-term outcome irrespective of a possible intestinal manifestation.

[Rectal goblet cell carcinoid. Primary tumor or metastasis?]. / Becherzellkarzinoid des Rektums. Primarius oder Metastase?

Goblet cell carcinoids are biphasic neoplasms of the gastrointestinal tract composed of a glandular and neuroendocrine differentiation. Typically, goblet cell carcinoids are localized in the vermiform appendix. We report the case of a 60-year-old female patient with the diagnosis of a 1.2-cm rectal goblet cell carcinoid tumor discovered during prophylactic proctocolonoscopy. Because of the known aggressive behavior of this entity, a rectosigmoidectomy was performed. The preoperative staging revealed neither local nor systemic spread. After 8 months, the patient is in good health. As a primary tumor of the extraappendiceal gastrointestinal tract, goblet cell carcinoids are a rarity. It is generally recommended to exclude metastasis of a primary appendiceal neoplasm. However, since the patient underwent an appendectomy in 1974, primary origin in the rectum is favored.

[Littoral cell angioma of the spleen. Association with colon and hepatocellular carcinoma]. / Littoralzellangiom der Milz. Assoziation mit Kolon- und hepatozellulärem Karzinom.

Littoral cell angioma is a rare vascular tumor of the spleen. An increased association with concomitant malignancies has been described. We report the case of a 67-year-old man suffering from colon carcinoma and presenting with a tumor of the spleen which could be seen using radiography. Splenectomy was performed and histology revealed a 2.1 cm vascular tumor with a characteristic immunophenotype (CD31/68+). In addition a hepatocellular carcinoma was found. Due to the relationship with malignant tumors littoral cell angioma should be considered in every case of a splenic tumoral space-occupying mass, particularly in carcinoma patients.

A randomized multicenter phase II study comparing capecitabine with irinotecan or cisplatin in metastatic adenocarcinoma of the stomach or esophagogastric junction.

BACKGROUND: The combination of irinotecan with 5-fluorouracil demonstrates efficacy with tolerable safety in the first-line treatment of metastatic gastroesophageal cancer (mGC). This randomized phase II trial compared for the first time capecitabine with irinotecan or cisplatin in this setting. PATIENTS AND METHODS: Patients were randomly assigned to receive 3-week cycles of capecitabine 1000 mg/m(2), twice daily for 14 days, with on day 1 either irinotecan 250 mg/m(2) (XI) or cisplatin 80 mg/m(2) (XP). The primary end point was overall response rate (ORR) and secondary end points included progression-free survival (PFS), overall survival (OS) and safety. RESULTS: Of 118 patients recruited, 112 were eligible for safety analysis and 103 for efficacy analysis. In the XI and XP treatment arms, there were no marked differences in ORR, 37.7% versus 42.0%, and median PFS, 4.2 versus 4.8 months, although median OS was longer, 10.2 versus 7.9 months, respectively. Grade 3/4 toxicity was higher in the XP regimen for thrombocytes (18.2% versus 1.8%), nausea (23.6% versus12.3%) and vomiting (16.4% versus 1.8%) and in the XI arm for diarrhea (22.8% versus 7.3%). CONCLUSION: The comparable activity and safety of the XI and XP regimens establish XI as a relevant platinum-free first-line treatment choice for patients with mGC.

[MALT-lymphoma]. / MALT-Lymphom.

Gastric lymphoma is most frequently an extranodal marginal zone-B-cell-lymphoma of mucosa-associated lymphoid tissue (MALT). The diagnosis is established by biopsy of the tumor. Computed-tomography, endosonography and recently capsule-endoscopy are used for staging. It is a relative new finding that MALT-lymphomas of the stomach are induced by Helicobacter pylori. A side-effect-poor breakthrough has been achieved by eradication of this bacterium with antibiotics. Refractory cases are amenable to radiotherapy, chemotherapy or surgical resection. The rarer aggressive lymphomas of the stomach are treated primarily by chemotherapy and radiotherapy. For both entities a further improvement may be achieved in ongoing prospective clinical trials by addition of the monoclonal antibody rituximab, which targets CD20 on lymphoma cells.

Capsule endoscopy in gastrointestinal lymphomas.

BACKGROUND AND STUDY AIMS: Capsule endoscopy (CE) is a new procedure for small-bowel imaging. The potential contribution of this method to the diagnosis and staging of gastrointestinal lymphomas has not yet been evaluated. The aim of this prospective study was to assess the frequency and morphology of different forms of intestinal pathology in patients with gastrointestinal lymphomas. PATIENTS AND METHODS: Commercially available capsule video endoscopes were administered to 27 patients (16 men and 11 women, aged 27-77) with known gastrointestinal lymphomas. CE was also carried out in 30 control individuals. RESULTS: All seven patients with primary intestinal lymphomas who were examined were found to have pathological findings on CE--four with ulcerations, four with nodes, three with villous atrophy, and one with plaques/white villi. One patient with severe diarrhea was examined three times before and after chemotherapy, and improvement of the lesions was evident, as well as resolution of diarrhea. Twenty patients with gastric lymphoma were examined (16 with low-grade and four with high-grade B cell lymphomas). The capsule did not leave the stomach in three patients, suggesting impaired motility. The small-bowel transit times were 261+/-41 min for intestinal lymphoma, 245+/-62 min for gastric lymphoma, and 224+/-82 min for controls (P>0.05). Five of the 20 patients with gastric lymphoma had pathological findings in the intestine--three with plaques/white villi, two with nodes, and two with villous atrophy. In two patients, subsequent biopsies revealed secondary follicular lymphoma and high-grade lymphoma, respectively. CONCLUSIONS: CE is a valuable diagnostic tool for defining the extent of bowel involvement and assessing the efficacy of treatment in patients with gastrointestinal lymphoma.

Randomised phase II evaluation of irinotecan plus high-dose 5-fluorouracil and leucovorin (ILF) vs 5-fluorouracil, leucovorin, and etoposide (ELF) in untreated metastatic gastric cancer.

An open-label randomised comparison of efficacy and tolerability of irinotecan plus high-dose 5-fluorouracil (5-FU) and leucovorin (LV) (ILF) with etoposide plus 5-FU/LV (ELF) in patients with untreated metastatic or locally advanced gastric cancer. One cycle of ILF comprised six once-weekly infusions of irinotecan 80 mg m(-2), LV 500 mg m(-2), 24-h 5-FU 2000 mg m(-2), and ELF comprised three once-daily doses of etoposide 120 mg m(-2), LV 300 mg m(-2), 5-FU 500 mg m(-2). In all, 56 patients received ILF and 58 ELF. Median age was 62 years, Karnofsky performance 90%, and disease status was comparable for both arms. The objective clinical response rates after 14 weeks treatment (primary end point) were 30% for ILF and 17% for ELF (risk ratio (RR) 0.57, 95% confidence interval (CI) 0.29-1.13, P = 0.0766). Overall response rates over the entire treatment period for ILF and ELF were 43 and 24%, respectively (RR 0.56, 95% CI 0.33-0.97; P = 0.0467). For ILF and ELF, respectively, median progression-free survival was 4.5 vs 2.3 months, time to treatment failure was 3.6 vs 2.2 months (P = 0.4542), and overall survival was 10.8 vs 8.3 months (P = 0.2818). Both regimens were well tolerated, the main grade 3/4 toxicities being diarrhoea (18%, ILF) and neutropenia (57%, ELF). The data from this randomised phase II study indicate that ILF provides a better response rate than ELF, and that ILF should be investigated further for the treatment of metastatic gastric cancer.

[Palliative treatment for colorectal cancer]. / Kolorektales Karzinom.

Over 50,000 new cases of colorectal cancer are diagnosed in Germany every year. About half of these patients will be cured by surgery. The other reoccur or present primarily with advanced disease. Recently, the survival of patients with metastasized disease has been prolonged from about 6 months with best supportive care alone to more than 20 months with combination chemotherapy. Therefore, adequate palliation is reasonable. Irresectable liver metastasis can be treated by locally ablative procedures such as radiofrequency ablation (RFA) or laserinduced thermotherapy (LITT). In the individual case resection of lung or brain metastasis should be discussed. Practitioners have to deal with plenty of supportive opportunities e.g. analgesics, bisphosphonates, central acting drugs, nutrition but also with drug side effects. In centers different endoscopic or interventional radiologic procedures are offered. Considering the variety of therapeutic options, it is prudent to discuss individual treatment plans in an interdisciplinary "tumor board" and involve the practitioners in the decision making.

Better acceptance of Fecal Occult Blood Test (FOBT) for colorectal cancer screening during hospitalization.

INTRODUCTION: Colorectal cancer (CRC) is a leading cause of illness and death in the Western world. Screening with fecal occult blood test (FOBT) significantly reduces the death rate and the incidence from CRC but these tests are not widely accepted. We investigated the possible contribution of hospitalization to a better acceptance of CRC screening. PATIENTS AND METHODS: From October 1998 through September 2000, 721 consecutive patients between 45 and 75 years of age admitted for various reasons were asked for participation in the study. They were asked to participate in FOBT-screening. In case of refusal of FOBT they were asked a second time after detailed information. In patients who accepted 3 consecutive FOBT's were performed. In case of positive FOBT results colonoscopy and gastroscopy were performed. RESULTS: 149 (82 male/67 female) patients were included. 94 (63.5%) of them agreed to undergo FOBT primarily and 10 (6.8%) secondarily after detailed information. The total acceptance rate of the FOBT was 69.8% (m/w : 69.1%/71.6%). In one of 5 cases with a positive FOBT result colorectal cancer (CRC) was diagnosed. Information on repetition of FOBT after one year could be obtained from 82 patients (55%). 37 patients (45%) had undergone repeated FOBT. None of the 37 patients was motivated by the FOBT screening during hospitalization. CONCLUSIONS: Staying in a hospital offers a good chance to achieve a higher acceptance of the FOBT. Therefore, hospitalization may contribute to a better colorectal cancer prevention. However, motivation to regularly repeat screening does not last in all patients. Therefore, public campaigns as well as medical counseling need to continuously stress the necessity of CRC screening procedures.

Natural cytotoxicity and antibody-dependent cellular cytotoxicity (ADCC) is not impaired in patients suffering from chronic hepatitis C.

BACKGROUND/AIMS: Recent observations suggest that natural killer (NK) cell activity might be impaired in chronic hepatitis C. However, to date antibody-dependent cellular cytotoxicity (ADCC) has not been studied in chronic hepatitis C in detail. METHODS: Therefore, we investigated spontaneous and cytokine-induced (interleukin-2 and interferon-gamma) natural cytotoxicity and ADCC in 29 patients suffering from chronic hepatitis C and 19 healthy controls. Cytotoxicity was determined with a flow-cytometric assay, which can also assess monocyte cytotoxicity. As target cells we used the colorectal tumor cell line HT29 and the lymphoma cell line Raji. RESULTS: We found no significant differences with respect to spontaneous cytotoxicity (HCV versus healthy controls (32 vs. 46%) and 17-1A specific ADCC (59 vs. 48%), even if isolated monocytes or NK cells were studied. Preincubation and stimulation of effector cells with cytokines increased both natural cytotoxicity and ADCC by 20-30%. However, natural cytotoxicity and ADCC after stimulation did not differ between the two groups. CONCLUSIONS: Our data obtained with a long-term cytotoxicity assay do not reveal impaired cytolytic capacity of the innate immune system in chronic hepatitis C, even when isolated monocytes and NK cells were studied as effector cells.

Therapeutic vaccination against metastatic carcinoma by expression-modulated and immunomodified autologous tumor cells: a first clinical phase I/II trial.

Therapeutic vaccination of tumor patients with cytokine gene-transfected tumor cells leads to tumor regression in animal models but has so far not resulted in significant clinical benefit. We and others demonstrated that tumor cells transfected to mediate overexpression of a cytokine gene activate immunologic effector cells for an improved proliferation rate and significantly higher antitumoral cytotoxic activity. Here, we performed a pilot study of therapeutic vaccination in patients with metastatic disease. Autologous tumor cells were simultaneously transfected with novel minimalistic, immunogenically defined, gene expression constructs (MIDGE) for overexpression of the two cytokines interleukin 7 (IL-7) and GM-CSF and newly designed double stem-loop immunomodulating oligodeoxyribonucleotides (d-SLIM) as a Th1-promoting and NK cell-stimulating adjuvant. Transfection was performed ex vivo by ballistomagnetic gene transfer. Patients received four subcutaneous injections of at least 1 x 10(6) of their expression-modulated and immunomodified autologous tumor cells. Ten patients have been enrolled in the study protocol. In all patients no adverse effects could be detected. IL-7 and interferon gamma levels were elevated in the serum of the patients after treatment. Interestingly, cytotoxicity of patient-derived PBLs increased significantly during treatment. All 10 patients had progressive disease when entering our protocol. One complete, one partial, and one mixed response with progression of abdominal metastases and regression of lung metastases were observed. Two patients showed a stable disease after treatment and five patients remained in progressive disease. Our observations confirm the capability of autologous expression-modified and immunomodulated tumor cell vaccines to stimulate a strong immune response in patients with metastatic cancer even in the presence of a large tumor burden.

Influence of cytokines, monoclonal antibodies and chemotherapeutic drugs on epithelial cell adhesion molecule (EpCAM) and LewisY antigen expression.

MoAbs against tumour-associated antigens (TAA) may be useful for the treatment of colorectal cancer. Since an increased expression of TAA may lead to enhanced antibody-dependent cellular cytotoxicity we examined whether the cytokines IL-2, IL-4, IL-6, IL-10, IL-12, interferon-alpha (IFN-alpha), IFN-gamma, granulocyte-macrophage colony-stimulating factor, macrophage colony-stimulating factor and tumour necrosis factor-alpha can influence EpCAM and LewisY expression on the surface of the colorectal carcinoma cell lines HT29, LoVo and SW480. We found that only IFN-alpha increased significantly whereas IL-4 decreased both EpCAM and LewisY expression. IFN-gamma significantly increased LewisY expression only. When tumour cells were treated with MoAb, the LewisY-specific MoAb BR55-2 down-regulated LewisY antigen expression, whereas MoAb 17-1A, which binds to EpCAM, up-regulated this TAA after 3 days of culture. The cytokines IFN-alpha or IFN-gamma combined with MoAb 17-1A enhanced further slightly the expression of EpCAM. In additional experiments with chemotherapeutic drugs commonly used for the treatment of colorectal cancer, we found that 5-fluorouracil, mitomycin-C and oxaliplatin up-regulated EpCAM and LewisY antigen expression. Raltitrexed enhanced LewisY and down-regulated EpCAM expression, whereas CPT-11 had no influence at all. The highest expression for EpCAM on HT29 cells was achieved by the combination of IFN-alpha, 5-fluorouracil and MoAb 17-1A. Our results may be useful for defining combinations of biological and chemotherapeutic drugs for the treatment of colorectal cancer. Further trials should evaluate to what extent these combinations enhance antibody-dependent cellular cytotoxicity.

A bispecific single-chain antibody directed against EpCAM/CD3 in combination with the cytokines interferon alpha and interleukin-2 efficiently retargets T and CD3+CD56+ natural-killer-like T lymphocytes to EpCAM-expressing tumor cells.

Cytokine-induced killer cells (CIK), generated in vitro from peripheral blood mononuclear cells (PBMC) by addition of interferon gamma (IFNgamma), interleukin-2 (IL-2), IL-1 and a monoclonal antibody (mAb) against CD3, are highly efficient cytotoxic effector cells with the CD3+CD56+ phenotype. In this study, we evaluated whether the cytotoxicity of these natural-killer-like T lymphocytes against the colorectal tumor cell line HT29 can be enhanced by the addition of a bispecific single-chain antibody (bsAb) directed against EpCAM/CD3. For determination of bsAb-redirected cellular cytotoxicity we used a new flow-cytometric assay, which directly counts viable tumor cells and can assess long-term cytotoxicity. We found that this bsAb induced distinct cytotoxicity at a concentration above 100 ng/ml with both PBMC and CIK at an effector-to-target cell ratio as low as 1:1. CIK cells revealed higher bsAb-redirected cytotoxicity than PBMC. Cellular cytotoxicity appeared after 24 h whereas PBMC showed the highest bsAb-redirected cytotoxicity after 72 h. The addition of the cytokines IL-2 and IFNalpha but not granulocyte/macrophage-colony-stimulating factor enhanced bsAb-redirected cytotoxicity of both PBMC and CIK. When the bsAb was combined with the murine mAb BR55-2, which recognizes the Lewis(Y) antigen, bsAb-redirected cytotoxicity was partly augmented, whereas murine mAb 17-1A, which binds to EpCAM as well, slightly suppressed bsAb-redirected cytotoxicity induced by the bsAb. We conclude that CIK generated in vitro or in vivo combined with this new EpCAM/CD3 bsAb and the cytokine IL-2 should be evaluated for the treatment of EpCAM-expressing tumors.

Prestimulation of monocytes by the cytokines GM-CSF or IL-2 enhances the antibody dependent cellular cytotoxicity of monoclonal antibody 17-1A.

Previously, we have shown that the cytokines IFN-alpha, IFN-gamma and IL-2 significantly enhance the antibody dependent cellular cytotoxicity (ADCC) exerted by the monoclonal antibody (mAb) 17-1A which recognizes the tumor associated antigen EpCAM. ADCC was assessed by a new flow cytometric cytotoxicity assay using the PKH2 labeled colorectal tumor cell line HT29 as target cells and peripheral blood mononuclear cells as effectors. Monocytes are assumed to be one of the major effectors for ADCC. However, isolated monocytes have a rather low ADCC capacity while addition of CD4+ lymphocytes optimizes ADCC. Since such an interaction between immune cells may act through cytokines we investigated whether a seven-day-prestimulation of monocytes by the cytokines M-CSF, GM-CSF, IFN-gamma, IFN-alpha and IL-2 enhances ADCC. Thereafter, we added for three days IL-2 and IFN-alpha with or without the mAb 17-1A for terminal activation of monocytes. Interestingly, GM-CSF which was ineffective in terminal activation, significantly enhanced ADCC of monocytes when it was used for prestimulation. Similar results were obtained with IL-2. IFN-gamma and M-CSF were also active but less than GM-CSF. Astonishingly, IFN-gamma and IFN-alpha prestimulation of monocytes suppressed the enhancement of ADCC exerted by GM-CSF and IL-2, respectively. Our experiments suggest that the timing of cytokine application is critical for the induction of optimal ADCC. Subcutaneous pretreatment with GM-CSF or IL-2 followed by the combination of IL-2/IFN-alpha/17-1A should be evaluated in a phase I clinical trial in patients with colorectal cancer.

Mechanism of cytotoxicity induced by chimeric mouse human monoclonal antibody IDEC-C2B8 in CD20-expressing lymphoma cell lines.

With a new flow cytometric cytotoxicity assay, we examined the mechanism of action of chimeric mouse human anti-CD20 monoclonal antibody IDEC-C2B8. IDEC-C2B8 alone induced direct cytotoxicity in four of eight examined CD20-expressing lymphoma cell lines (RAJI, DAUDI, JOK-1, and WT100) at a concentration above 100 ng/ml. Moreover, after 4 h incubation in human serum, only a moderate complement-dependent cellular cytotoxicity (CDCC) was observed, whereas cytotoxicity increased markedly after 3 days of culture, indicating that combined direct cytotoxicity and CDCC were responsible. IDEC-C2B8 induced an effective antibody-dependent cellular cytotoxicity (ADCC) in seven of eight tested lymphoma cell lines when peripheral blood mononuclear cells were used as effector cells. ADCC was moderately enhanced by cytokine interleukin-2, whereas interleukin-12, interferon-alpha, and GM-CSF had no influence. Interestingly, we could demonstrate a correlation between CD32 expression on lymphoma cell lines and IDEC-C2B8-induced direct cytotoxicity, indicating that crosslinking of CD20 with CD32 may be involved in the mechanism of cytotoxicity. We propose that direct cytotoxicity, CDCC, and ADCC result in the marked elimination of CD20-expressing tumor cells observed after treatment with IDEC-C2B8.

Augmentation of 17-1A-induced antibody-dependent cellular cytotoxicity by the triple cytokine combination of interferon-alpha, interleukin-2, and interleukin-12.

Previously, interferon-alpha (IFN-alpha), interleukin-2 (IL-2), and interleukin-12 (IL-12) were shown to increase the antibody-dependent cellular cytotoxicity (ADCC) induced by the murine monoclonal antibody 17-1A, which recognizes the tumor-associated antigen EpCAM. In this study, the authors wanted to determine whether the combination of these three cytokines would yield greater cytotoxicity than the single cytokines. For cytotoxicity assessment, a new flow cytometric assay was used that allows the analysis of long-term ADCC exerted by macrophages. Peripheral blood mononuclear cells from healthy donors were used as effector cells against the colorectal carcinoma cell line HT29 at a low effector-to-target ratio of 4.5:1. With this test, the effectiveness of the combinations IL-2 and IFN-alpha, IL-2 and IL-12, and IL-12 and IFN-alpha were compared with each other. The combinations IL-2 plus IL-12 and IFN-alpha plus IL-12 were more potent at the concentrations tested. Furthermore, the triple cytokine combination of IFN-alpha, IL-2, and IL-12 revealed significantly greater ADCC than dual cytokine combinations. Next, CD14+, CD4+, and CD4- cells were isolated by paramagnetic beads and magnetic activated cell sorter (MACS) columns. The CD14+ and CD4- cell populations contained the ADCC effectors. The addition of CD4+ cells to CD14+ or CD4- cells resulted in augmented ADCC, indicating that cooperation between immune cells occurs. These results suggest that multiple cytokine combinations with monoclonal antibodies may be more effective for cancer immunotherapy.

Combinations of the cytokines IL-12, IL-2 and IFN-alpha significantly augment whereas the cytokine IL-4 suppresses the cytokine-induced antibody-dependent cellular cytotoxicity of monoclonal antibodies 17-1A and BR55-2.

Since some cytokines effectively enhance the cytotoxicity of monoclonal antibodies, we investigated whether a combination of cytokines can augment the antibody-dependent cellular cytotoxicity (ADCC) of monoclonal antibodies 17-1A and BR55-2 against the colorectal carcinoma cell line HT29. Since monocytes/macrophages are important effector cells for ADCC, we used a new flow cytometric cytotoxicity assay, which allows the analysis of long-term-ADCC exerted by these cells. In our previous studies with peripheral blood mononuclear cells from normal donors, we found that IL-2, IL-12 and IFN-alpha increase ADCC. Therefore, we examined whether combination of these three cytokines with IL-2, IL-4, IL-6, IL-10, IL-12, IFN-alpha, IFN-gamma, GM-CSF, M-CSF and TNF-alpha may yield higher ADCC than obtained by the application of single cytokines. Indeed, we found that the combinations IL-2/IFN-alpha, IL-2/IL-12 and IL-12/IFN-alpha potentiated ADCC. Interestingly, the ineffective single cytokines TNF-alpha and GM-CSF in the combinations IL-2/TNF-alpha, IFN-alpha/TNF-alpha and IFN-alpha/GM-CSF also proved to enhance ADCC. In contrast, IL-4 significantly suppressed the IL-2, IL-12 and IFN-alpha-induced ADCC. In addition, the immunosuppressive cytokine IL-10 in higher concentrations significantly suppressed the IL-12-induced-ADCC. Our results may be useful to find combinations of cytokines and mAb for the treatment of cancer.

Enhancement of antibody dependent cellular cytotoxicity (ADCC) by combination of cytokines.

Monoclonal antibodies (MAb) specific for tumor-associated antigens (TAA) can induce an immunological cellular attack of tumor cells by a process termed antibody dependent cellular cytotoxicity (ADCC). Cytokines may augment ADCC by direct activation of immune cells or by enhancement of TAA on tumor cells. Thus, we investigated whether ADCC by MAb 17-1A and BR55-2, which recognize TAA on colorectal tumor cells, can be augmented by 3-day incubation with different concentrations of IL-2, IL-4, IL-6, IL-12, IFN-alpha, IFN-gamma, GM-CSF, M-CSF, and TNF-alpha. ADCC was assessed by a new flowcytometric cytotoxicity assay (Flieger et al. Immunol Methods 1995; 180:1-13) using PKH-2 labeled HT29 cells as targets and PKH-26 labeled peripheral blood mononuclear cells from three healthy volunteers as effector cells. We found three reaction patterns with the cytokines tested: (a) cytokines, which increase ADCC (IL-2, IL-12, IFN-alpha, and IFN-gamma, which represent Thl cytokines); (b) cytokines with no effect (GM-CSF, M-CSF, and TNF-alpha); and (c) cytokines, which decrease ADCC (IL-4 and IL-6, which represent Th2 cytokines). Then, we tested cytokines that increase ADCC in combination with the other cytokines. We found that the combinations IL-2/IFN-alpha, IL-2/IFN-gamma, IL-2/IL-12, and IL-12/IFN-alpha potentiated ADCC. By contrast, IL-4 reduced the IL-2, IL-12, and IFN-alpha-induced ADCC. Since the Thl response, cooperation of monocytes and CD4 cells is involved, we plan to elucidate by magnetic cell sorting (MACS) separation techniques, which cells are involved in cytokine-induced ADCC. Our results may be useful for finding combinations of cytokines and MAb for the locoregional treatment of colorectal cancer.

The combination of interleukin-2 and interferon alpha effectively augments the antibody-dependent cellular cytotoxicity of monoclonal antibodies 17-1A and BR55-2 against the colorectal carcinoma cell line HT29.

Monoclonal antibodies (mAb) are promising substances for the treatment of colorectal carcinoma, but the efficiency of this therapy still needs further improvement. We used a flow-cytometric cytotoxicity test to determine the efficacy of the cytokines interferon alpha (IFNalpha) and gamma (IFNgamma), interleukin-2 (IL-2), macrophage-colony-stimulating factor (M-CSF), granulocyte/macrophage-CSF (GM-CSF) and tumor necrosis factor alpha (TNFalpha) in enhancing the antibody-dependent cellular cytoxicity (ADCC) of the mAb 17-1A and the mAb BR55-2 against the colorectal carcinoma cell line HT29. In experiments performed at an effector to target ratio of 9:1, with peripheral blood mononuclear cells from five healthy volunteers as effector cells, we found that IFNalpha, IFNgamma and IL-2 significantly augmented the ADCC of both mAb at concentrations between 3 ng/ml and 30 ng/ml. The other three cytokines were not effective. In further experiments we examined combinations of the three effective cytokines in different concentrations. The combination of IFNalpha and IL-2 proved to be optimal in enhancing ADCC of both mAb. Thus, the examination of ADCC by flow cytometry may reveal potentially useful combinations of cytokines and mAb for the treatment of colorectal carcinoma.