Interferon Alpha Treated Patients with Chronic Myeloid Leukemia (CML) In Prolonged Complete Remission Have Increased Numbers of NK-Cells and Clonal Gamma-Delta T-Cells, and a Distinct Plasma Cytokine Profile.

Before the era of tyrosine kinase inhibitors (TKIs), interferon alpha (IFN-α) was the treatment of choice in CML and prolonged survival of responding patients. Recent studies suggest that combination of IFN-α with TKI improves therapy outcome. Significantly, a proportion of IFN-α treated patients in prolonged complete cytogenetic remission (CCyR) have been able to discontinue treatment without disease relapse (Mahon et al. JCO 2002). The mechanism of action of IFN-α therapy is incompletely understood; the drug exerts both direct cytotoxic and immunomodulatory effects on leukemic cells. The aim of this project was to study the immunomodulatory effects of IFN-α in CML patients in prolonged remission and isolate biological markers predicting response.

The study population consisted of CML patients treated with IFN-α monotherapy (n = 10, median therapy time 146 months, range 63–231 months) and CML patients who had discontinued IFN-α therapy but remained in remission for >2 years (n = 9, median therapy time 120 months, range 80–184; median time without therapy 53 months, range 24–96). None of the patients were previously treated with TKI therapy. In addition, non-CML patients (3 patients with essential trombocythemia and one patient with polycythemia vera) treated with IFN-α and healthy volunteers (n = 43) were included as controls. Lymphocyte populations in all four groups were characterized with comprehensive immunophenotyping panels. The clonality of T-cells was analyzed by a TCR γ/δ rearrangement assay by PCR. Lymphocytes were further sorted into CD3⁺ TCR αβ⁺ and CD3⁺ TCR γδ⁺ populations (n = 8). Additionally, plasma levels of 25 cytokines were measured with a multiplex bead-based cytokine assay (Luminex®).

The proportion of NK-cells from lymphocytes was significantly increased in IFN-α discontinued patients (median 26%, range 18–51%) compared to healthy volunteers (11%, 5–21%) or patients on IFN-α therapy (12%, 6–31%)(P=0.0005). Similarly the proportion of CD8⁺ cells from T-cells was significantly increased in both CML IFN-α groups (55% in IFN-α discontinued patients, 44% in IFN-α treated patients vs. 31% in healthy volunteers; P<0.05 for both groups). Also a larger proportion of T-cells expressed the long-term memory antigen CD45RO in IFN-α patients (74%, 58% vs. 44% in healthy controls, P<0.01). The proportion of regulatory T-cells (CD4+CD25+FoxP3+) was increased in IFN-α groups (6.1%, 5.2% vs. 3.8% in healthy volunteers, P=0.01). Similar changes in immunoprofile were not observed in IFN-treated non-CML patients.

Clonal TCR γ/δ rearrangements were observed in 18 of 19 (95%) IFN-treated CML patients as compared to 3 of 22 (14%) in healthy volunteers (P<0.01). In both IFN-α CML patient groups a unique rearrangement pattern was observed: 14/19 (79%) of patients had the Vγ9 gene clonally rearranged. This clonal rearrangement resided in CD3⁺ γδ⁺ T-cell population, as assessed by cell sorting. Two of four non-CML patients treated with IFN-α had the same clonal rearrangement, as well as one healthy control (1/22; 5%). Similar clonality patterns have not been observed in dasatinib or imatinib treated CML patients (Kreutzman et al. Blood 2010).

IFN-α treatment was associated with a distinct plasma cytokine profile in CML patients. IP-10, IL-6, IL-12, eotaxin, MCP-1, and IFN-γ levels were significantly increased in IFN-α treated CML patients. In particular, eotaxin and MCP-1 levels differed significantly between healthy controls and IFN-α patients who had successfully discontinued IFN-α therapy (428 vs. 1173 pg/ml, P<0.0001 and 107 vs. 459 pg/ml, P=0.0003, respectively). In IFN-α treated non-CML patients, eotaxin or MCP-1 levels were not increased.

Our results show that IFN-α treatment induces distinct changes in the immunoprofile of CML patients. Patients who had successfully discontinued IFN-α therapy differed markedly from healthy controls. IFN-α therapy was associated with increased numbers of NK-cells and clonal γδ⁺ T-cells. These cells possess potent anti-leukemic activity and may contribute to the prolonged therapy responses in this group of patients. Furthermore, plasma cytokine profile could be a helpful biomarker when considering which patients can discontinue the IFN-α treatment without imminent disease relapse.

Faber:BMS, Novartis: Consultancy, Honoraria. Porkka:BMS, Novartis: Consultancy, Honoraria, Research Funding. Mustjoki:BMS, Novartis: Honoraria.