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Circulating antibodies reflect a molecular imprint of antigens that are related to autoimmune diseases, cancer or infection. Importantly, serum antibodies are useful clinical markers as they carry diagnostic information from all around the human body. Moreover, the amplification cascade governed by the humoral immune system causes a surplus of circulating antibodies after appearance of the corresponding (low abundance) antigen. In combination with the fact that antibodies are highly stable compared to many other serum proteins, they seem ideal to be implemented in clinical diagnostic assays for the detection of antigen-associated diseases. This review summarises advances in immunoproteomics with respect to technologies for biomarker discovery, with special emphasis on recently developed gel-free MS-based approaches, and looks forward to potential immunoproteomic applications in diagnostic medicine.

BACKGROUND:Intestinal bacteria have long been implicated in colorectal cancer pathology, and many reports point to a close linkage between Streptococcus bovis biotype I (recently renamed Streptococcus gallolyticus) infections and tumors of the human colon. This work aims to investigate the humoral immune response to this bacterium during different stages of colorectal cancer. METHODS: The presence of serum antibodies against S. bovis antigen RpL7/L12, previously assigned as a potential diagnostic antigen, was evaluated in Dutch (n ¼ 209) and American (n ¼ 112) populations using a newly developed enzyme-linked immunosorbent assay. RESULTS: The analyses consistently showed that an immune response against this bacterial antigen was increased in polyp patients and stage I/II colorectal cancer patients as compared with asymptomatic individuals. This was not paralleled by increased antibody production to endotoxin, an intrinsic cell wall component of the majority of intestinal bacteria, which implies that the humoral immune response against RpL7/L12 is not a general phenomenon induced by the loss of colonic barrier function. Notably, increased anti-RpL7/L12 levels were not or were only mildly detected in late stage colorectal cancer patients having lymph node or distant metastasis. CONCLUSIONS: These findings are indicative of an increased exposure to antigen RpL7/L12 during early stages of colon carcinogenesis and suggest that intestinal bacteria such as S. bovis constitute a risk factor for the progression of premalignant lesions into early stage carcinomas. Clearly, the current findings emphasize the necessity for further studies on the possible etiologic relationship between intestinal bacteria and human colorectal cancer. Cancer 2010;116:4014-22.

Streptococcus gallolyticus (formerly known as Streptococcus bovis biotype I) is a low-grade opportunistic pathogen which is considered to be associated with colon cancer. It is thought that colon polyps or tumors are the main portal of entry for this bacterium and that heparan sulfate proteoglycans (HSPGs) at the colon tumor cell surface are involved in bacterial adherence during the first stages of infection. In this study, we have shown that the histone-like protein A (HlpA) of S. gallolyticus is a genuine anchorless bacterial surface protein that binds to lipoteichoic acid (LTA) of the gram-positive cell wall in a growth phase-dependent manner. In addition, HlpA was shown to be one of the major heparin-binding proteins of S. gallolyticus able to bind to the HSPG-expressing colon tumor cell lines HCT116 and HT-29. Strikingly, although wild-type levels of HlpA appeared to contribute to adherence, coating of additional HlpA at the bacterial surface resulted in reduced binding to colon tumor cells. This may be explained by the fact that heparan sulfate and LTA compete for the same binding site in HlpA. Altogether, this study implies that HlpA serves as a fine-tuning factor for bacterial adherence.

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