Soluble plasma HLA peptidome as a potential source for cancer biomarkers

Abstract

The HLA molecules are membrane-bound transporters that carry peptides from the cytoplasm to the cell surface for surveillance by circulating T lymphocytes. Although low levels of soluble HLA molecules (sHLA) are normally released into the blood, many types of tumor cells release larger amounts of these sHLA molecules, presumably to counter immune surveillance by T cells. Here we demonstrate that these sHLA molecules are still bound with their authentic peptide repertoires, similar to those of the membranal HLA molecules (mHLA). Therefore, a single immunoaffinity purification of the plasma sHLA molecules, starting with a few milliliters of patients' blood, allows for identification of very large sHLA peptidomes by mass spectrometry, forming a foundation for development of a simple and universal blood-based cancer diagnosis. The new methodology was validated using plasma and tumor cells of multiple-myeloma and leukemia patients, plasma of healthy controls, and with cultured cancer cells. The analyses identified thousands of sHLA peptides, including some cancer-related peptides, present among the sHLA peptidomes of the cancer patients. Furthermore, because the HLA peptides are the degradation products of the cellular proteins, this sHLA peptidomics approach opens the way for investigation of the patterns of protein synthesis and degradation within the tumor cells.

The nature and extent of contributions by defective ribosome products to the HLA peptidome

“…The cell lysate was spun at 18,000 × g for 30 min, and the supernatant was passed through a column containing the W6/32 antibody covalently bound to AminoLink resin (Pierce), as described in ref. 57. The columns were washed with 10 volumes of 400 mM NaCl and with another 10 volumes of 20 mM Tris·HCl, pH 8.…”

Analysis of Major Histocompatibility Complex (MHC) Immunopeptidomes Using Mass Spectrometry*

“…1, middle panel). In fact, this method can be used for the isolation of both class I and class II MHC peptides from a range of biological sources such as cell lines, primary tissues and plasma (36,(51)(52)(53). Moreover, up to ϳ90% of immunopurified peptides were reported to be specific for the MHC molecules (51,54).…”

Mining the plasma immunopeptidome for cancer peptides as biomarkers and beyond

“…HPLC separation followed by MS/MS, while rapid compared with prior technology, is still labor intensive, technically challenging, and expensive. After these technical hurdles are surmounted (as they surely will be given the rapid advances in MS hardware and software), Bassani-Sternberg et al (4) provide an exciting approach for targeting, screening, and monitoring cancer and other common human diseases.…”

“…Insightfully, Bassani-Sternberg et al (4) recognize that the soluble immunopeptidome (i.e., peptides bound to plasma sHLA) represents a window into ongoing disease processes that potentially of- Fig. 1.…”

“…Bassani-Sternberg et al (4) exploit this phenomenon to describe a method to characterize peptide repertoire bound to MHC class I molecules (the immunopeptidome) (13) present in plasma. In addition to being expressed on the plasma membrane, class I molecules are released (through membrane shedding or metalloprotease cleavage) (14) or actively secreted from cells (by generation of splice variants lacking the transmembrane domain) (15) and are present in human plasma, which was first reported in 1970 (16).…”