Chronic Viral Infection, Virus-Specific Monoclonal Immunoglobulin, and Development of Plasma Cell Malignancy

Chronic infection by B-cell tropic viruses can lead to B-cell malignancy via the direct transformation of infected B-lymphocytes or indirectly via cell transformation consecutive to chronic antigen-driven stimulation. In the context of infection by Hepatitis C virus (HCV), the monoclonal immunoglobulin (Ig mc) that arise are typically directed against the virus (Blood 2008; 112 :4357). Moreover, the two mechanisms may occur simultaneously (New Engl J Med 2003; 348 :178). Here we studied 11 patients diagnosed with monoclonal Ig of undetermined significance (MGUS, 3 cases) or multiple myeloma (MM, 8 cases) for whom HCV and/or Epstein-Barr virus (EBV) serologic status was known.

To determine the specificity of monoclonal Ig of MGUS and MM patients chronically infected with HCV or EBV.

Monoclonal Ig were purified as described (Blood 2008; 112 :4357). Three of the 11 patients were positive for HCV. For the 8 HCV-negative patients, EBV tests showed that 4 had a positive serology (IgG) for both Epstein Barr Nuclear Antigen (EBNA) and Viral Capside Antigen (VCA); the 4 other patients were negative for both EBNA and VCA. Sera and purified monoclonal Ig were tested using novel HCV and EBV specific epitope arrays generated by spotting affinity-purified recombinant proteins (either core and non structural HCV proteins or EBNA and VCA epitopic peptides) in 4 replicates on nitrocellulose-coated slides. After saturation with blocking solution, slides were incubated with serum or purified monoclonal Ig. After washing, slides were incubated with a labelled secondary antibody (Dylight™ 680 Labelled Goat anti-human IgG (H+L) (KPL)) in the dark, washed, scanned and fluorescence signals were detected using a ScanArray Microarray Analysis system (PerkinElmer).

Using the HCV/EBV antigen array described above, purified monoclonal Ig from the 3 HCV-positive patients (1 MGUS, 2 MM) specifically recognized HCV, either the core protein (2 patients) or the NS-4 protein (1 patient). For 3 of the 4 EBV-positive patients (1 MGUS/smoldering myeloma, 2 MM), purified monoclonal Ig specifically recognized EBNA. The HCV/EBV arrays were specific as no significant fluorescent signal was observed in the following control conditions: sera or purified monoclonal Ig from HCV-negative patients, tested on HCV proteins; purified monoclonal Ig of patients with a negative serology for EBNA and VCA, tested for EBV epitopic peptides; EBNA-specific purified monoclonal Ig, tested for EBV VCA epitopes.

Certain monoclonal Ig of EBV-positive MM patients are specific for EBNA, and monoclonal Ig of HCV-infected MGUS and MM patients are directed against HCV. Thus for subsets of MM, the first events triggering progression toward plasma cell malignancy can be viral infection and abnormal –monoclonal- response to it. Since anti-viral therapy could help eradicate the virus-driven plasma cell clone, efforts should be made to identify such patients, preferably at the MGUS stage. The novel HCV and EBV antigen arrays we developed should be tests of choice.

Moreau:Janssen: Advisory board, Honoraria; Millennium Pharmaceuticals, Inc.: Advisory board, Honoraria.