XBP-1 a Selective and Specific Target for Immunotherapy in Myeloma.

Abstract

X-box binding protein 1 (XBP-1) is a transcription factor is essential for the differentiation of plasma cells and the unfolded protein response. XBP-1 is significantly up-regulated in myeloma cells compared to both normal plasma cells and B-cells. Selective and specific requirement of XBP-1 for differentiation of B cells to plasma cells and its further up-regulation in multiple myeloma (MM) cells makes it a promising target for immunotherapy directed at MM. We have evaluated XBP-1 as a target antigen to develop MM-specific immunotherapy. In order to generate XBP-1 antigen-specific cytotoxic T lymphocytes (CTLs), we have identified HLA-A2-specific peptides derived from non-spliced (short form) and spliced (long form) XBP-1 proteins. We have further modified the peptide by altering one amino acid, which is critical for HLA-A2 affinity, to obtain epitope, which has higher stability to HLA-A2 clefts. The modified XBP-1 peptides were able to evoke higher levels of IFN-g release compared to native peptides and were able to induce CTLs highly specific to MM, demonstrated by Calcein-released cytotoxicity assay. Cytotoxic activity of these XBP1 peptide-specific-CTLs against U266, a HLA-A2+/XBP-1+ MM cell line was 60% and 79% by CTLs stimulated by non-spliced peptide and 69% and 88% by CTLs stimulated by spliced peptide at Effector:Target ratios of 20:1 and 60:1, respectively. The CTLs did not lyse HLA-A2− or XBP-1− cells. These results were further confirmed by the novel CD107 cytotoxicity assay, which detects MM-specific cytolytic CD8+ T cells by flow-cytometric analyses. The CTLs generated with XBP-1 peptide displayed distinct phenotypes, showing high CD69+CD45RO+ cell population (62% in non-spliced peptide-CTLs, 67% in spliced peptide-CTLs, 4% in unstimulated control-CTLs) and CD45RA+CCR7+ cell population (1% in non-spliced peptide-CTLs, 1% in spliced peptide-CTLs, 24% in unstimulated control-CTLs). In conclusion, we report the identification of highly immunogenic heteroclitic XBP-1 epitopes that have ability to generate MM-specific CTLs. Confirmation of in vivo activity of these CTLs in SCID mouse model is currently underway prior to its evaluation in clinical studies.

Chemical bonding: From Lewis to atoms in molecules

“…One distinguishes between charge concentrations (CCs) and charge depletions (CDs) determined by the topology of L ( r ), and charge accumulations and charge reductions determined by the differing topology of ρ( r ). L ( r ) exhibits multiple valence maxima that are found to coincide with the number and relative positions of the localized electron pair domains that have been invoked in chemical models 12. The much simpler topology of ρ( r ), while defining the atom and molecular structure, in general exhibits maxima only at the positions of the nuclei 14…”

“…Spatial displays of the density of the Fermi hole, a distribution function in six‐dimensional space, are of limited use 2. What is needed is a clear representation of the pairing of electrons in real space, a requirement fulfilled by the Laplacian of the electron density 12…”

“…Since G ( r ) > 0 and $\cal V ({\bf r}) < 0$ , a negative value for ∇ 2 ρ( r ), as found at a CC in L ( r ), indicates a region where $\cal V ({\bf r})$ overrides the kinetic energy making the electronic potential energy density maximally stabilizing, thus accounting for the role of bonded and nonbonded CCs as sites of nucleophilic activity or Lewis bases. Contrawise, minima in L ( r ), denote sites that perform as Lewis acids and the approach of reactants is determined by the alignment of the CCs and CDs on one reactant with the complement of CDs and CCs on the other, the Laplacian complement of Fischer's ‘lock and key’ analogy of binding 12. A recent example of this property of L ( r ) is the accounting of the structure and formation of donor–acceptor complexes of transition metal carbonyls.…”

Influence of substitution on the strength and nature of CH···N hydrogen bond in XCCH···NH3complexes

“…The Bader theory is a powerful tool not only to characterize the electron properties of the considered species but also to have a better understanding of the nature of the intermolecular interactions 56, 57. The characteristic of the BCP and its Laplacian (∇ 2 ρ BCP ) reveal the nature of the interaction.…”

Functional groups expressed as graphs extracted from the Laplacian of the electron density

“…This article presents a sizeable set of graphs of a variety of functional groups occurring in organic chemistry, drawn from the Laplacian of the electron density, denoted by ∇ 2 ρ. About two decades ago this simple function was proposed 5 as a measure for electron pair localization, an idea that has since led to a considerable number of studies 6–52. In all these articles the Laplacian appears as a central feature but in many more studies 53, 54 it is only evaluated at bond critical points to characterize bonds, increasingly in crystallographically obtained electron densities.…”