Plasma Proteome Profiling with Monoclonal Antibody Libraries: A Pilot Biomarker Analysis for Nanomedicine-Induced Complement Activation

Abstract

Complement (C) activation-related hypersensitivity reactions (HSRs) represent an unsolved adverse immune effect of many i.v. administered "nanomedicines", such as liposomal doxorubicin (Doxil/Caelyx). Because these pseudoallergic reactions can be severe or even lethal, it is an important clinical objective to find biomarkers for proneness for C activation by reactogenic nanoparticles that will allow the prediction of in vivo reactions by in vitro assays. With this goal in mind we identified a normal human blood donor whose serum consistently showed high sensitivity to Caelyx-induced C activation in vitro (CSS). The plasma of this blood (Caelyx-sensitive plasma, CSP) was subjected to proteome profiling with a library of human plasma proteome specific mAbs. The chip (PlasmaScan-380 TM) contained 380 non-redundant (with respect to epitopes) mAbs. The analysis revealed 8 proteins that were differentially represented in CSP in comparison with Caelyx-insensitive control plasma. These proteins were identified by mass spectrometry and Western blot analyses to represent factor H (decreased in CSP), factor H related protein, serum amyloid P component, fibronectin, complement component C4, Apo B100, prothrombin and alpha-2-HS glycoprotein (all increased in CSP). Some of these protein changes are consistent with proneness for increased C activation, suggesting the potential use of this method in the search for biomarkers for liposome-induced or other types of nanomedicine-induced HSRs.

The possible role of factor H in complement activation-related pseudoallergy (CARPA): a failed attempt to correlate blood levels of FH with liposome-induced hypersensitivity reactions in patients with autoimmune disease

“…Among the significant changes we found that the level of FH decreased 1.9-fold in CSP, while the level of FHR was increased 3.4-fold (Table 1) (18). The decrease in FH was consistent with proneness for increased C activation, while the increase of FHR was difficult to interpret.…”

“…Increased FHR levels could also result in less regulation by FH due to their antagonistic effect on FH function via competition with FH for certain ligands and surfaces. This may explain the association of increased FHR protein level in Caelyx sensitive plasma compared with Caelyx non-sensitive plasma (18).…”

“…Based on the above theory on the potential use of FH as a biomarker for drug-induced CARPA, we recently carried out a study wherein the protein profile in the plasma of a normal human subject was analyzed, who showed proneness for C activation by liposomal doxorubicin (Caelyx) in vitro (referred to as Caelyx-sensitive plasma, CSP) (18). The aim was to find one or more specific changes that could be considered as a biomarker for increased susceptibility for C activation.…”

Multicapillary SDS‐gel electrophoresis for the analysis of fluorescently labeled mAb preparations: A high throughput quality control process for the production of QuantiPlasma and PlasmaScan mAb libraries

“…Here, we report on the comparison of contemporary electrophoresis techniques (SDS‐PAGE, and multicapillary SDS‐CE with light‐emitting diode induced fluorescence (LED‐IF)) for the development of a streamlined and high throughput quality analysis of QuantiPlasma™ and PlasmaScan™ mAb libraries (>1000 mAbs), developed for the mAb proteomics based global plasma proteome profiling at BioSystems International Kft. (Debrecen, Hungary) .…”

Fractionation of the human plasma proteome for monoclonal antibody proteomics‐based biomarker discovery 2: Antigen identification by dot‐blot array screening

“…Such targeted arrays can represent a versatile tool in detecting possible candidate cancer biomarkers; however, the validation of these markers is needed afterwards . mAb proteomics, a novel high throughput, mAb‐based biomarker discovery platform has recently been developed offering the option of mass screening for biomarkers with nascent or cloned (PlasmaScan™ and QuantiPlasma™) mAb libraries. It is a reversed proteomic approach also requiring subsequent identification and validations steps .…”