Lethal BCG-osis in the context of superficial urothelial bladder carcinoma, diagnosed in autopsy

AbstractIntra-vesicular Bacillus Calmette–Guérin (BCG) immuno-therapy is a cornerstone in the treatment of superficial bladder carcinoma. However, it can rarely cause BCG-osis, a systemic infection, potentially fatal. A case of a 73year old man, diagnosed with superficial urothelial bladder carcinoma five months ago, who died with clinical features of septic shock one day after BCG therapeutic maintenance infusion, is presented. The cause of death, BCG-osis, was revealed during autopsy

Frequent high-level expression of the immunotherapeutic target Ep-CAM in colon, stomach, prostate and lung cancers

Epithelial cell adhesion molecule (Ep-CAM; CD326) is used as a target by many immunotherapeutic approaches, but little data are available about Ep-CAM expression in major human malignancies with respect to level, frequency, tumour stage, grade, histologic tumour type and impact on survival. We analysed by immunohistochemical staining tissue microarrays with 4046 primary human carcinoma samples from colon, stomach, prostate and lung cancers for both frequency and intensity of Ep-CAM expression under highly standardised conditions. A total of 3360 samples were analysable. High-level Ep-CAM expression was observed in 97.7% (n=1186) of colon, 90.7% of gastric (n=473), and 87.2% of prostate cancers (n=414), and in 63.9% of lung cancers (n=1287). No detectable Ep-CAM staining was found with only 0.4% of colon, 2.5% of gastric, 1.9% of prostate cancers, and 13.5% of lung cancers. The only significant correlation of Ep-CAM expression with tumour grading was observed in colon cancer where high-level Ep-CAM expression on grade 3 tumours was down to 92.1% (P<0.0001). Adenosquamous and squamous carcinomas of the lung had a lower percentage of high-level Ep-CAM expression compared to adenocarcinomas with 35.4 and 53.6%, respectively, and with 45.5 and 17.3% of tumours being Ep-CAM negative. With the exception of moderately differentiated colon carcinoma, where patients not expressing Ep-CAM on their tumours showed an inferior survival (P=0.0014), correlation of Ep-CAM expression with survival did not reach statistical significance for any of the other cancer indications and subgroups. In conclusion, the data strongly support the notion that Ep-CAM is a prime target for immunotherapies in major human malignancies. This is because the most common human cancers show (i) a low frequency of Ep-CAM-negative tumours, (ii) a high frequency of Ep-CAM expression on cells of a given tumour, and (iii) for most cancers, an insignificant influence of tumour staging, grading and histology on Ep-CAM expression

Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries

The identification of novel T cell antigens is central to basic and translational research in autoimmunity, tumor immunology, transplant immunology, and vaccine design for infectious disease. However, current methods for T cell antigen discovery are low throughput, and fail to explore a wide range of potential antigen-receptor interactions. To overcome these limitations, we developed a method in which programmable microarrays are used to cost-effectively synthesize complex libraries of thousands of minigenes that collectively encode the content of hundreds of candidate protein targets. Minigene-derived mRNA are transfected into autologous antigen presenting cells and used to challenge complex populations of purified peripheral blood CD8+ T cells in multiplex, parallel ELISPOT assays. In this proof-of-concept study, we apply synthetic minigene screening to identify two novel pancreatic islet autoantigens targeted in a patient with Type I Diabetes. To our knowledge, this is the first successful screen of a highly complex, synthetic minigene library for identification of a T cell antigen. In principle, responses against the full protein complement of any tissue or pathogen can be assayed by this approach, suggesting that further optimization of synthetic libraries holds promise for high throughput antigen discovery

Natural and induced immunity against the tumour-associated antigen, Ep-CAM

The tumour-associated antigen (TAA), Ep-CAM is overexpressed on various human carcinomas, including colorectal carcinoma (CRC). TAAs or their immunodominant epitopes that are spontaneously recognised by the immune system might constitute a suitable target for immunotherapy. Fifteen % of sera of CRC patients with no previous immunotherapy elicited IgG antibodies against Ep-CAM. No Ep-CAM specific antibodies were detected in healthy controls or patients with Crohn´s disease or colitis ulcerosa. Further analyses revelaed that 50% of the Ep-CAM-reactive sera bound to peptide residues 29-46 of Ep-CAM. The results provide evidence for spontaneous immune recognition of Ep-CAM in CRC patients and identify an immunodominant B cell epitope of human Ep-CAM. Anti-idiotypic antibodies (anti-Id) may serve as surrogate TAAs for vaccination. The optimal design of an anti-Id vaccine, however remains unclear. Moreover, whether vaccination with anti-Id or the original antigen is superior is controversial. SM262 is a human anti-Id raised against mAb 17-1A that recognises Ep-CAM. Vaccination of mice with anti-Id induced antibodies that shared idiotopes with mAb 17-1A and recognised Ep-CAM. Fusion of GM-CSF to anti-Id enhanced the magnitude of the antibody responses, while xenogeneic Fc domain had no significant modulatory effect. Recombinant anti-Id protein vaccine evoked a more potent humoral immunity as compared to DNA delivered by gene gun. Our study provides the fist evidence that immune tolerance in mice expressing the transgene for human Ep-CAM can be circumvented by anti-Id vaccination. The results may have implications for future anti-Id vaccine design. Vaccination of CRC patients with recombinant Ep-CAM protein, in combination with GM-CSF, induced Ep-CAM specific T and NK-like T cells producing cytotoxic cytokines. In addition, a long-lasting Th1 biased humoral and proliferative T cell response was elicited against Ep-CAM. The original antigen, Ep-CAM induced a more potent overall immune response as compared to anti-Id mimicking Ep-CAM. Analysis of TCR BV gene repertoire revealed that BV19+ CD8+ T cells might be involved in the vaccine induced anti-Ep-CAM immune response. The results collectively suggest that immunisation with Ep-CAM protein may serve as a novel approach to CRC immunotherapy. Furthermore, immunogenic MHC class I and II restricted Ep-CAM epitopes were identified that may provide new opportunities for developing effective multiepitope cancer vaccines targeting Ep-CAM. Vaccination with a recombinant canarypox virus (ALVAC) encoding human Ep-CAM in combination with GM-CSF induced a potent Ep-CAM specific, type 1 cellular immune response in CRC patients. However, no anti-Ep-CAM antibody or proliferative T cell responses were elicited. Combining ALVAC-Ep-CAM and recombinant Ep-CAM in a prime-boost vaccination approach may represent an effective strategy to induce a coordinated antigen specific cellular and humoral immune response. In conclusion, the results suggest that Ep-CAM is a promising target structure for immunotherapy. The present studies may form a basis for further enlarged clinical trials targeting Ep-CAM by active specific vaccination