Pegylated interferon for PV & ET Josef Prchal, University of Utah, Salt Lake City Educated at School of Medicine Karlova Universita, Praha
Studies of X-inactivation: MPNs are clonal disorders
Clonal development in polycythemia vera JAK2V617F
Beneficial effect of Peg-IFNα in PV and ET patients Cytoreductive Rx treatment in PV/ET can: hematological remission & decrease thromboses but clonal hematopoiesis persists (i.e. normal HSCs still suppressed) Only IFNα can induce: hematological remission reduction of JAK2 V617F allelic burden return of polyclonal hematopoiesis (suppression of the pre-JAK2V617F PV clone) Liu at all, Blood 2003 Kiladjian, Blood, 2008, Quintas, Cardama, JCO, 2009 Liu, Blood, 2003
Is this salutary effect of Peg-IFNα achieved by stimulation of an immune response against the PV clone ? preferential activation of normal dormant stem cells (HSCs) ? suppression of the PV clone by Peg-IFNα? Via TNFa?
Studied patients (MPD-RC 111 and MPD-RC 112) Phase 3 randomized trial high-risk polycythemia vera (PV) and essential thrombocythemia (ET): Peg-IFNα versus hydroxyurea MPD-RC 111 PV and ET patients intolerant or refractory to hydroxyurea and patients with Budd Chiari syndrome Patients who do not participate in the protocols
Effect of PegasysTM in PV and ET research methods Determination of JAK2V617F allelic burden in clonal granulocytes and CD34+ stem cells Response of PV/ET clones to Peg-IFNα in informative females (q-TCA clonality assay based on X-chromosomal inactivation) Correlation with circulating regulatory T cells Correlation with HSC cell cycle status Correlation with HSC TNFa levels
Molecular response to Peg-IFNα We studied 23 patients (11 females and 12 males) with allelic burden of JAK2V617F at enrollment (1.2% - 99.8%) Following treatment at 6 mo, 14 patients (60.8%) demonstrated a decrease in JAK2V617F allelic burden Females patients with reversion from monoclonal to polyclonal hematopoiesis JAK2617F allelic burden in % P1-PV P2-PV P3-PV Females patients with persistence of clonal hematopoiesis after treatment with Peg-IFNα JAK2617F allelic burden in % P4-PV P5-PV P6-PV P7-PV P8-PV P9-ET P10-PV P11-PV Before treatment GNC Before treatment CD34+ After Peg-IFNα GNC After Peg-IFNα CD34+
Correlation between the absolute number of peripheral blood Tregs and the JAK2V617F allelic burden in PV patients during the treatment with Peg-IFNα JAK2V617F Treqs Peripheral blood Tregs (fold difference) JAK2 V617F (%) Peripheral blood Tregs (fold difference) JAK2 V617F (%) Treqs JAK2V617F Days Days Peg-IFNα: mobilizes marrow Tregs to the periphery, decreasing their immunosuppressive and tumor promoting influence in marrow microenvironment? effects on Tregs may be associated with suppression of pre-JAK2V617F and JAK2V617F –positive clonal hematopoiesis?
Peg-IFNα and HSC in PV ET Peggy Goodell Katherine King
Total HSC number and proliferation Peg-IFNα: decreases the percentage of HSCs in G0 (quiescence) and increases the number of HSCs in G2/MS-phase- suggesting that Peg-IFNα therapy promotes normal HSC proliferation/differentiation n=8 n=13
Myeloid Differentiation Cyclin genes Expression Peg-IFNα: increases cell cycle associated genes expression Myeloid Differentiation Peg-IFNα: increases colony-forming units in methylcellulose, reflecting a greater number of functional progenitor cells –correlation with decreased CBC?? n=10 N=19
TNFα transcript before and after Peg-IFNα in BM CD34+ cells Relative expression of TNFα Relative expression of TNFα
Interaction plot for treatment modality and time on log (TNF-relative expression) and JAK2V617F mutation
define the immune response to PV and ET Ongoing Studies define the immune response to PV and ET The humoral response induced by Peg-IFNα to Tumor Associated Ag (TAA) expressed by Hematopoeitic Stem Cells (HSC). T-Cell response:multi color flow cytometry effector CD4+ T cells, CD4+Foxp3+ Treg and CD8+ T cells. Using surface markers, (CD3, CD4, CD8, CD25, CD38, CD39, CD45RO, CD197, CTLA-4 and HLA-DR) and intracellular markers (Foxp3, Helios and Ki-67) we will be able to determine the frequency as well as the phenotype of different T cells in the PB and BM and compare them pre and post Peg-IFNα treatment Quantitate the expression of immune response regulatory checkpoint molecules (PD-1, CTLA-4, OX40 and 4-1BB) at the surface of T cells Correlate with clonality, JAK2V617F allelic burden, and TNFa before and after Peg-IFNα (with Vladimir Divoky)
Stem cells components in PV and ET patients Prchal, Blood 2011
Summary We conclude that Peg-IFNα can suppress both pre-JAK2V617F and JAK2V617F PV/ET clones in some PV/ET patients, as demonstrated by evaluation of HSCs, granulocytes and platelets, but this often occurs asynchronously In some patients hematological and molecular response delayed as long as 2 years, as yet in study of >30 patients none of the patients with dramatic decrease of JAK2V617F allelic burden (<3 %) achieved full molecular remission Our data demonstrate that Peg-IFNα therapy is associated with an increase in HSCs cycling and mobilization of Tregs to the circulation After Peg-IFNα therapy dramatic decrease of TNF-α in stem cells of responders Is benefit of Peg-IFNα justified by >side-effects and increased costs compared to hydoxyurea Rx?