Autoantibodies: On Patrol for Cancer

Cancer diagnosis often involves a multitude of tests after symptom onset, complicating treatment for tumors that are no longer localized. While genetic analysis indicates cancer risk, it lacks direct disease relevance. Attention has now turned toward discovering novel protein and autoantibody (AAb) biomarkers that offer real-time biological insights [1]. With recent evidence suggesting that tumor antigenicity occurs at the protein level, AAb profiling is a rapidly emerging trend to identify tumor-relevant proteins, gain direct insight into the humoral immune response to cancer, and discover novel biomarkers that are more stable than mRNA and protein [2].

The Humoral Immune System and Cancer

Cancer causes changes at the protein level that may stimulate the production of AAbs [3]. These changes can include aberrant folding; altered expression in terms of location and level; and post-translational modifications (PTMs). For example, Type I MAGE proteins exhibit normal expression in the male mammalian germline during development. However, in men and women with cancer, these cancer-testis antigens (CTAs) are expressed abnormally in non-germ cells, leading to the generation of AAbs [4-6]. In a study by Patel et al., 13 AAbs in patients with resected non-small cell lung cancer were discovered that predicted poor survival rates with a sensitivity and specificity of 84% and 74%, respectively, in a validation cohort. Six of the 13 AAbs targeted CTAs [5].

Benefits of Autoantibody Biomarkers in Cancer

AAbs often predate disease diagnosis by years, enabling disease monitoring and early detection before tumors metastasize [7-10]. Furthermore, collecting AAbs from blood draws is less invasive and faster than biopsies, yielding quicker results. Since they are more stable than mRNA and protein, AAbs are excellent biomarkers for retrospective and longitudinal studies. Population screening in rural and low-income areas is possible because antibodies on dried blood spot collection cards are stable for one month at room temperature and 200 days when stored at -20 °C [11, 12].

Antibodies are also uniquely positioned to identify multiple tumor targets. Zaenker et al. identified a signature of 10 novel AAbs present in clinically diagnosed melanoma patients with a sensitivity of 79% and a specificity of 84%. Among the 10 autoantigens, kinases, transcription factors, and regulatory proteins were represented, expressed in a variety of cells and tissues including the immune and nervous systems [13]. Kinases that phosphorylate proteins are heavily targeted for therapeutics. Recently, histone citrullination has been associated with a variety of cancers [14, 15].

Research Tools to Identify Disease-Relevant Antigens

Protein profiling technologies like mass spectrometry help identify proteins involved in cancer pathology to understand the molecular mechanisms of disease and develop targeted therapeutics. However, protein profiles have traditionally shown a weak correlation with early manifestation of cancer and have struggled to accurately predict disease outcome. Also, these methods commonly analyze proteins in plasma and serum, which only contain proteins that have been secreted or released into the bloodstream. This means that proteins at the tumor site, which could be crucial for understanding cancer, are often missed. Additionally, proteins that cause the disease cannot be distinguished from proteins that correlate to the disease by protein profiling alone [16]. Furthermore, these methods do not provide information about the immunogenicity of cancer-associated proteins, which is vital for understanding how the body’s immune response targets cancer cells.

Functional protein microarrays help fill the knowledge gaps left by protein profiling techniques. Consuming only a small volume (i.e., < 15 mL) of serum and other biological matrices, antibodies can be screened against hundreds to thousands of proteins simultaneously to identify proteins that are modified and immunogenic in cancer. In addition, AAbs obtained from blood samples represent the full repertoire of antibodies from various tissues, as they circulate system-wide. Multiple antibody isotypes can also be analyzed in parallel, providing valuable and direct information about the immune response. For instance, a study of human papillomavirus found that anti-viral IgA antibodies present shortly after infection correlated with a reduced risk of cervical cancer, indicating a protective role for IgA antibodies against cancer [17]. Similarly, research by Zhang et al. identified a panel of IgG and IgM AAbs linked with colorectal cancer [2].

The Importance of Protein Folding in Immunoprofiling

With 90% of humoral antibodies binding conformational epitopes, screening tools with correctly-folded proteins are required for accurate and meaningful results [18]. Sengenics arrays with full-length proteins leverage their patented KREX® technology to ensure proper protein folding for highly specific antibody binding [19]. Moreover, proteins are immobilized to a hydrogel substrate that provides an aqueous environment for the proteins to float naturally. Notably, linear, denatured, and improperly folded proteins can expose hydrophobic sequences that are extremely “sticky,” leading to nonspecific antibody binding and inaccurate data (Figure 1).



Figure 1. KREX Technology. Each protein is tagged with a biotin carboxy carrier protein (BCCP), which binds to biotin only when the protein and BCCP are correctly folded. Subsequently, the protein is captured to the array surface via a stable biotin-streptavidin interaction. Misfolded proteins cannot bind to biotin and will be washed away, thus excluded from analysis.


Concluding Remarks

New advances in immunoproteomics are crucial to advance cancer research in detecting disease earlier, predicting outcomes more accurately, and revealing new protein targets for treatment.


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The editorial staff had no role in this post's creation.