Plasma Cell Gene-Expression Profiles in Patients with Systemic AL Amyloidosis: Responses to Melphalan and Stem Cell Transplant Are Associated with Differential Expression of Genes Involved in Translation, Protein Degradation and Detoxification.

Systemic AL amyloidosis is a plasma cell dyscrasia and protein conformation disorder in which Ig light chains (Ig V_L) produced by clonal plasma cells form interstitial amyloid in key viscera causing organ dysfunction and death. Patients with AL usually have < 20% marrow plasma cells with low to nil proliferative indices. Paradoxically the plasma cells appear immune to the toxic amyloid-forming Ig V_L possibly due to the robust cytoplasmic protein quality-control processes in plasma cells. A standard approach is to treat with high-dose melphalan and stem cell transplant (SCT) without induction therapy. Post-SCT long-term survival depends on significant reductions of the plasma cell disease and Ig V_L levels as measured by the serum free light chain (FLC) assay. At 3 months post-SCT responses of the plasma cell dyscrasia are defined both by standard criteria and by normalization of abnormal FLC ratios. By standard criteria, post-SCT one-third fail to achieve a > 50% reduction in plasma cell disease (NR), one-third achieve a > 50% reduction (PR), and one-third achieve clearance of marrow plasma cells and a complete or near complete response (CR). By FLC criteria, post-SCT one-third achieve normalization (N) of the FLC ratio and two-thirds do not (A). Many factors contribute to this distribution of responses. In order to identify factors specific to AL plasma cells, gene expression profiles (GEP; Affymetrix U133 PLUS 2.0) were obtained after double amplification of RNA from FACS-sorted CD138+/DAPI- plasma cells from untreated patients with AL pre-SCT. Data were vetted based on plasma cell lineage gene expression; samples contaminated with monocytes were not used. Supervised analyses were performed based on responses at 3 months post-SCT, comparing CR (n=4) to PR (n=7) and CR to NR (n=5), and FLC N (n=5; one with CR) to A (n=7; two with CR). Differentially expressed genes between paired sets were identified using the t-test. Genes with p < 0.01 were examined using EASE, a program that identifies over-represented gene families. In the CR-PR and CR-NR comparisons, genes involved in translation, RNA ligation and protein degradation, particularly aminoacyl tRNA synthetases, were significantly over-represented with Bonferroni-adjusted EASE scores (like p-values) < 0.05. In the CR set, tryptophanyl tRNA synthetase, a protein that can also be anti-angiogenic, and IDE, an insulin-degrading enzyme that also degrades Aβ amyloid peptides, were among the most over-expressed. In the FLC N-A comparison, protein transport and detoxification gene families were also significantly over-represented with Bonferroni-adjusted EASE scores < 0.001. In the N set, CCT subunits (chaperones), UBE2B (a ubiquitination enzyme and RAD6 homolog) and glyoxalase I (detoxification) were among the most over-expressed. Our initial hypothesis is that the CR and FLC N responses to melphalan have similar but distinctive GEP related to protein folding, ER stress and protein degradation. The responses of AL plasma cells to Ig V_L in the ER may influence the patterns observed, possibly modulating melphalan uptake or activity. Further studies of these differences and functional hypothesis testing are currently underway.