Shared and individual specificities of immunodominant cytotoxic T-cell clones in paroxysmal nocturnal hemoglobinuria as determined by molecular analysis.
OBJECTIVE
Similar immune mechanisms have been suggested to operate in aplastic anemia (AA) and paroxysmal nocturnal hemoglobinuria (PNH), and the presence of PNH clones in AA may indicate that an immune reaction directed against hematopoietic stem cells may be responsible for the immune selection pressure leading to PNH evolution. We previously described expansions of selective cytotoxic T-lymphocyte (CTL) clones in AA patients.


METHODS
We applied a molecular analysis of the T-cell receptor repertoire to study the characteristics of CTL response in patients with various forms of PNH. Immunodominant T-cell clones were detected using combined flow cytometric and molecular analysis of the variable beta (VB) chain and CDR3 representation, followed by determination of the frequency of individual CDR3 clonotypes. Clonotypic polymerase chain reaction (PCR) was performed to establish clonotypic utilization pattern.


RESULTS
In patients with a past history of AA, and when subgrouped by current blood counts as "hypoproliferative" PNH patients (in contrast to purely hemolytic form of PNH), more pronounced skewing of VB family utilization was found, consistent with T-cell responses involving several immunodominant CTL clones. Sequences of the PNH-derived clonotypes were used to design PCR-based assays for the utilization analysis of individual clones in PNH patients. The clonotypic distribution pattern established by this PCR method indicated that immunodominant T-cell specificities were shared between some patients but also may be found at low frequencies in controls.


CONCLUSIONS
Analysis of the CDR3 sequence pattern as a marker for expanded immunodominant clonotypes may have an application in the study of T-cell responses in PNH.
Experimental Hematology and Hematopoiesis Section, Taussig Cancer Center, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
Magdalena
M
Plasilova
Antonio M
AM
Risitano
Christine L
CL
O'Keefe
Alexander
A
Rodriguez
Marcin
M
Wlodarski
Neal S
NS
Young
Jaroslaw
J
Maciejewski
D000328:Adult; D000368:Aged; D000741:Anemia, Aplastic / Q000150:complications / Q000276:immunology; D001327:Autoimmune Diseases / Q000276:immunology*; D002999:Clone Cells / Q000276:immunology; D022801:Complementarity Determining Regions / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D018450:Disease Progression; D005260:Female; D015333:Gene Rearrangement, beta-Chain T-Cell Antigen Receptor; D017261:Glycosylphosphatidylinositols / Q000172:deficiency; D006457:Hemoglobinuria, Paroxysmal / Q000150:complications / Q000235:genetics / Q000276:immunology*; D006801:Humans; D016056:Immunodominant Epitopes / Q000276:immunology*; D008297:Male; D008565:Membrane Proteins / Q000235:genetics; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000235:genetics; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
D022801:Complementarity Determining Regions; D018076:DNA, Complementary; D017261:Glycosylphosphatidylinositols; D016056:Immunodominant Epitopes; D008565:Membrane Proteins; D016693:Receptors, Antigen, T-Cell, alpha-beta; C080086:phosphatidylinositol glycan-class A protein