Immunogenetic Characterization of Primary Immune Thrombocytopenia (ITP) in Adults: Preliminary Results of the Unit Study.

In adults, ITP displays variable clinical presentation and different response to steroid, Rituximab, and other immune suppressive agents. The pathophysiological differences underlying these different behaviours are mostly unknown and a better knowledge of this biological heterogeneity might help identifying more targeted and rationale treatments for this disease.

To identify immunogenetic features distinguishing ITP patients from control and different subsets of ITP patients based on clinical presentation and responsiveness to steroid or Rituximab. Several biological analyses were based on the model of autoimmune lymphoprolipherative syndrome (ALPS), a pediatric disease due to genetic defects decreasing function of the Fas death receptor involved in shutting off the immune response and cytotoxic cell function. ALPS displays polyclonal lymphoproliferation, peripheral blood (PB) expansion of TCRαβ+ T-cells double negative for CD4 and CD8 (DN T-cells) and autoimmune manifestations frequently including thrombocytopenia. ALPS is mostly due to deleterious mutations of the Fas gene, but the +1239A>C single nucleotide polymorphism (SNP) of the osteopontin gene (OPN) and several variations of the perforin gene (PRF1) can act as disease modifier. Moreover, defective Fas function and these OPN and PRF1 variations are also displayed by subsets of patients with multiple sclerosis, type-I diabetes mellitus, systemic lupus erythematosus, progressive sclerosis, and chronic inflammatory demyelinating polyneuropathy.

Adult patients with ITP and matched controls were selected for the analyses and stratified in different clinical subsets according to the disease severity and responsiveness to therapy (asymptomatic, steroid or Rituximab sensitive vs. steroid or Rituximab refractory). Analyses included evaluation of Fas-mediated apoptosis in T cell cultures; proportions of DN-T cells in PB; typing of the +1239A>C SNP of OPN and sequencing of PRF1. All patients were also investigated for TCR monoclonality, whereas BCR monoclonality was analysed in Rituximab-untreated patients only.

Analysis of Fas function and DN T-cell expansion was assessed in 100 ITP patients and showed that they displayed higher frequency of defective Fas function than the controls (17/100 vs. 5/100; P<0.05). Expansion of DN T-cells was detected in 2/17 (12%) patients displaying defective Fas function and 3/83 (4%) of those with normal Fas function. Analysis of PRF1 and OPN was performed in 64 patients. Sequencing of PRF1 detected three patients carrying two rare variations; two carried the N252S amino acid substitution (previously described in ALPS) and one the novel R385W amino acid substitution. The overall frequency of these rare variations was higher in the patients than in the controls (4.7% vs. 0.8%, P<0.05). By contrast, the OPN +1239A>C SNP displayed a similar distribution in the patients and the controls. TCR monoclonality was assessed in 76 patients and was detected in 4 of them (5%). BCR monoclonality was assessed in 17 patients in PB and bone marrow and it was always absent. No statistical differences of these parameters were detected comparing patients refractory vs. sensitive to either Rituximab or steroid treatments. However, a trend was found for DN T-cell expansion that tended to be more frequent in Rituximab resistant vs. sensitive patients (0/13 vs. 4/20, P=0.13).

These preliminary analyses detected some differences between ITP patients and controls suggesting that defects involved in ALPS development may play a role in adult ITP too. Increasing the patient number is needed to confirm these data and, possibly, to detect differences between clinical subgroups.

Off Label Use: Rituximab in ITP.