No Development of Neutralizing Antibodies Against Recombinant Interferon-Alpha in Ph-Negative Myeloproliferative Neoplasms - a Prospective Study
Authors: Ocias Lukas Frans , Hansen Dennis Lund , Kristensen Thomas Kielsgaard , de Stricker Karin , El Fassi Daniel , Stentoft Jesper , Starklint Jørn , Frederiksen Mikael , Severinsen Marianne Tang , Bjerrum Ole Weis , Møller Peter , Mourits-Andersen Torben , Overgaard Ulrik Malthe , Hasselbalch Hans Carl , Bendtzen Klaus , Larsen Thomas Stauffer 
Published: November 2019

Abstract
Background
Treatment of Philadelphia chromosome negative chronic myeloproliferative neoplasms (MPNs) with recombinant pegylated interferon alpha2a/b (rIFN-alpha) has proven effective. It is well known that prolonged therapy with recombinant type 1 interferons (IFN-alpha and IFN-beta) may induce neutralizing antibodies (nAbs) against the drug leading to treatment failure. Most data on type 1 IFN immunogenicity are available from studies of patients with multiple sclerosis treated with rIFN-beta, and patients with hepatitis C treated with rIFN-alpha. A few reports have demonstrated nAbs in MPN patients not responding adequately to rIFN-alpha treatment, but the phenomenon has not been thoroughly investigated in MPNs.
Patients and Methods
Newly diagnosed patients with MPNs enrolled in the Danish multicenter trial - DALIAH (Low-dose rIFN-alpha versus Hydroxyurea in The Treatment of Ph-Negative MPNs) were included. Patients were randomized to treatment with either rIFN-alpha 2a or 2b at a starting dose of 45 and 35 mikrograms once weekly, respectively. The occurrence of neutralizing Abs (nAbs) against rIFN-alpha was investigated at baseline, month 12 and month 24 by reporter gene assays (iLiteTM alphabeta and iLiteTM antialpha, Biomonitor A/S, Copenhagen, Denmark). JAK2 V617F quantitative mutation analyses were performed as previously described (Larsen TS, BJH 2007). Statistical analyses were performed using STATA version 9.0.
Results
Ninety-two patients on sustained treatment with rIFN-alpha2a (n=48) and rIFN-alpha2b (n=44) for 12 months were analyzed for this study. Forty-five patients had ET, 39 patients had PV and 8 patients had proliferative PMF. Thirty-six out of 39 (92%) PV patients, 22 out of 45 (49%) ET patients and 4 out of 8 (50%) PMF patients were JAK2V617F mutated. Hematological responses at 12 months: ET: 67% CR, 29 % PR; PV: 64% CR, 31% PR (ELN 2009 criteria); PMF: 50% had at least a minor response (EUMNET).
The median serum concentration of bioactive IFN-alpha at 12 months was 12,4 (range <2,4-86,4) and 2,6 (range <2,4-12,8) IU/mL serum, for patients treated with rIFN-alpha 2a and -2b respectively. No significant association between hematological or molecular response and serum IFN-alpha activity was found. Serum from 92 patients was analyzed at 12 months and 33 patients were analyzed at both 12 and 24 months and no occurrence of nAbs was seen during treatment. Twenty-four patients had pre-treatment levels of IFN nAbs measured. Notably, one patient was tested positive for the presence of nAbs before rIFN-alpha exposure. This autoAb-positive patient remained positive throughout the study and has shown low IFN serum activity (< 2,4 IU/mL) and only partial hematological and molecular response after 24 months of treatment.
Conclusions
Development of nAbs in MPN patients completing treatment for 12 months with rIFN-alpha seems exceedingly rare as no patients, neither complete responders nor patients not meeting criteria for complete hematological remission developed nAbs after 12 (24) months of therapy. Its apparent rarity does not justify a routine investigation of nAbs in patients not responding to rIFN-alpha treatment. There was no significant correlation between serum concentration of rIFN-alpha 2a and -2b and clinical or molecular responses. The intriguing finding that one of 24 patients had pre-existing cross reacting nAbs against rIFN-alpha 2a and 2b before commencing rIFN-alpha treatment is interesting and was associated with an insufficient response.

Disclosures
Off Label Use: Recombinant interferon-alpha 2a and -2b in the treatment of chronic Philadelphia-negative myeloproliferative neoplasms.. El Fassi:Novartis Denmark: Honoraria, Other: Have conducted an educational session for Novartis Denmark, regarding MPNs and ruxolitinib, for this a honorarium was received.. Bjerrum:Bristoll Myers Squibb, Novartis and Pfizer: Other: educational activities. Hasselbalch:Novartis: Research Funding. Bendtzen:Pfizer: Honoraria; Eurodiagnostica AB: Equity Ownership; Novo-Nordisk: Equity Ownership.