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Background: Despite optimal surgical treatment and modern adjuvant therapies, 50% of patients diagnosed with colorectal cancer die within 5 years. Immunotherapy offers an appealing complement to traditional chemotherapy, with possible long-term protection against tumor recurrences through immunological memory. We have conducted a pilot study of a novel adoptive immunotherapy, using autologous, in vitro expanded lymphocytes isolated from the tumor-draining sentinel lymph node., Study Design: Sentinel nodes were recovered from 16 patients with disseminated or locally advanced, high-risk colorectal cancer. Single-cell suspensions of sentinel-node-acquired lymphocytes were clonally expanded in vitro in the presence of autologous tumor extract and returned as a transfusion. Patients were followed with clinical and radiological evaluations. Long-term survival was compared with traditionally treated controls., Results: Sentinel-node-acquired CD4(+) Th1-lymphocytes could be clonally expanded in vitro and safely administered to all 16 patients without side-effects. In four out of nine stage IV patients, complete tumor regression occurred. Median survival time in the stage IV patients (n = 9) was 2.6 years, as compared with 0.8 years in conventionally treated controls. A dose-dependent effect with regards to reduced tumor burden and long-term survival was observed., Conclusion: Sentinel-node-based adoptive immunotherapy is feasible; the method has shown no apparent side-effects and appears to convey therapeutic antitumor effects. Further studies are justified to determine its efficacy and precise role in the treatment of colorectal cancer.

The immunosurveillance theory argues that the immune system recognizes tumour-specific antigens expressed by transformed cells, which results in the destruction of cancer precursors before they become clinically manifest. As a model for the development of cancer, we set out to study premalignant lesions and immune responses in sentinel lymph nodes from patients with long-standing ulcerative colitis and progression of mucosal dysplasia. Mesenteric lymph nodes draining dysplastic and normal intestinal segments were identified by sentinel node technique during surgery in 13 patients with ulcerative colitis who were subjected to colectomy because of intestinal dysplasia. T cells were extracted from the lymph nodes and analysed by flow cytometry, and lymphocyte proliferation assays were set up in the presence of extracts from dysplastic and normal intestinal mucosa. Increase in CD4/CD8 ratio was observed in sentinel lymph nodes draining dysplastic epithelium compared to normal mucosa. The increase in CD4(+) T cells in relation to CD8(+) T cells correlated with the degree of dysplasia reflected by a significant increase in the ratio against low-grade dysplasia compared to indefinite dysplastic lesions. The T-cell response was specific to antigens from dysplastic epithelial lining as seen in proliferation assays. The observation suggests an important surveillance role for the immune system against premalignant intestinal lesions in patients with long-standing ulcerative colitis.

In colon cancer the presence of metastases in the regional lymph nodes is an important prognostic factor. Currently, examination is based on histological and immunohistochemical evaluations of lymph node sections. However, these methods are time-consuming, labour intensive and micro-metastases might be missed. The sentinel node technique may be used to identify the direct tumour draining lymph node. In this study the possible use of flow cytometry for detection of sentinel node metastases in patients with colon cancer has been investigated. Peripheral blood mononuclear cells (PBMC) with the addition of DLD-1 colon cancer cells were stained intracellularly for cytokeratin 20 (CK20), and for the cell surface markers epithelial cell adhesion molecule (EpCAM) and carbohydrate antigen 19-9 (CA19-9). CK20 positive colon cancer cells were reliably detected with as few as 0.037% events. However, PBMCs from both colon cancer patients and from healthy individuals contained CK20 positive cells. Staining for EpCAM resulted in an almost complete recovery of tumour cells, and as few as 0.037% added cells were detected. Low intra-assay variability was determined for EpCAM (CV 8.8) and CA19-9 (CV 9.7) stainings. The results from staining for CK20 or for EpCAM and CA19-9 concorded, but the degree to which respective antigens were expressed varied. The use of flow cytometry for detection of metastatic colon cancer cells was verified in fourteen sentinel nodes specimens from patients with colon cancer. Herein we have explored the potential of flow cytometry to become a fast, sensitive and reliable method for detection of lymphatic metastases in patients with colon cancer using direct fluorophore-conjugated antibodies against multiple surface antigens. The method seems feasible and further testing is warranted.

Background: We previously identified tumor-reactive lymphocytes in the first lymph nodes that drain the primary tumor. In this study, we performed lymphatic mapping to investigate the possibility of finding the first lymph nodes that drain metastases, and of learning whether these lymph nodes contained tumor-reactive lymphocytes suitable for adoptive immunotherapy., Methods: Nineteen patients were studied. The primary tumor site was colorectal cancer in seven patients, malignant melanoma in four, ovarian cancer and breast cancer in two, and one each with pancreatic cancer, cholangiocarcinoma, leiomyosarcoma, and squamous cellular cancer of the tongue. By injection of Patent blue dye or radioactive tracers around the metastases, we identified draining lymph nodes from liver metastases (n = 9), intra-abdominal local recurrences (n = 3), and regional lymph node metastases (n = 7). In six patients, a preoperative lymphoscintigraphy was performed., Results: We located the first draining lymph node or nodes from metastases or local recurrences; we named them "metinel nodes." Lymphocytes from the metinel nodes proliferated, showed clonal expansion, and produced interferon gamma (via in vitro expansions on stimulation with tumor homogenate) and interleukins, all of which demonstrate the characteristics of tumor-reactive lymphocytes. Eight of the nineteen patients received immunotherapy on the basis of tumor-reactive T cells derived from the metinel nodes., Conclusions: We demonstrate that it is possible to locate the first lymph nodes draining subcutaneous, lymphatic, and visceral metastases, the so-called metinel nodes. Metinel node-derived lymphocytes may be used to treat disseminated solid cancer, and clinical trials should evaluate the effect of such treatment.

The current status of lymphatic mapping and sentinel node biopsy in the treatment of patients with malignant melanoma and breast cancer is described. The possible use of a similar method in patients with colorectal and gastric cancer is outlined. Peroperative lymphatic mapping and identification of sentinel node(s) in patients with gastrointestinal cancer may lead to modified (tailored) resections and extended lymph node dissections only in those patients in whom the sentinel node(s) contains tumour cells. The method offers the possibility of improving staging by identification of patients with early disseminated disease who should be considered for adjuvant treatment or be included in trials of adjuvant treatment to speed up the breakthrough of more effective adjuvant regimens. Large studies are needed to find out if the sentinel node concept is as valid in gastrointestinal cancer as studies so far have shown that it is for malignant melanoma and breast cancer.

The sentinel node is the first lymph node to receive lymphatic drainage from a tumour and is usually the first site of metastases. Today, the sentinel node is used for tumour staging. Here, we focus on its immunological role and investigate lymphocytic function in sentinel nodes, identified intraoperatively by peritumoural dye injection, from 15 patients with colon cancer. Tumour infiltrating lymphocytes, sentinel and nonsentinel lymph node cells and peripheral blood leukocytes were studied by flow cytometry, proliferation assays and interferon-gamma secretion after activation with autologous tumour homogenate. Whereas tumour-infiltrating lymphocytes were nonresponsive in the proliferation assays, lymphocytes from sentinel nodes proliferated dose dependently and secreted interferon-gamma upon stimulation with tumour homogenate. The responses were of varying magnitude and tended to be weaker in metastatic sentinel nodes. Sentinel node lymphocytes represents an enriched source of tumour reactive lymphocytes, and may be useful in future trials of adoptive immunotherapy. [ABSTRACT FROM AUTHOR]