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Objectives Treatment-free remission (TFR) has emerged as a treatment goal in chronic myeloid leukemia in the chronic phase (CML-CP). Attempts to increase proportion of patients achieving TFR include combination of tyrosine kinase inhibitors (TKI) and other drugs. Interferon-α in addition to TKI has shown promising efficacy but with dose-dependent toxicity and discontinuations. NordCML007 was initiated to study the efficacy and safety of low dose pegylated IFN-α (PegIFN-α) in combination with dasatinib (DAS) in CML-CP. Methods Forty patients with newly diagnosed CML-CP were given DAS upfront. After month 3 (M3) 15 μg/wk of PegIFN-α was added and increased to 25 μg/wk from M7 until M15. DAS treatment was continued and adverse events and BCR-ABL1 qRT-PCR values were reported yearly after M24. Results from M1 to M18 have previously been published, and here we present long-term data. Results After 5 years of follow-up, there were no suspected unexpected serious adverse reactions, no increase in serosal effusions, no disease progressions and no CML-related deaths. Rates of MR3.0 (MMR), MR4.0 and MR4.5 were 84.6%, 64.1% and 51.3% respectively at M60, and 95% of patients reached MMR at some point during the study. Conclusion Initial addition of PegIFN-α to DAS shows good long-term efficacy without increased toxicity. Title in Web of Science: Long-term tolerability and efficacy after initial PegIFN-alpha addition to dasatinib in CML-CP: Five-year follow-up of the NordCML007 study

Two decades after the introduction of tyrosine kinase inhibitors (TKI), a sizeable portion of patients with chronic myeloid leukemia (CML) in chronic phase (CP) still undergo allogeneic stem cell transplantation (allo-HSCT). We investigated the indications for allo-HSCT, clinical outcome, management of relapse, and post-transplant TKI treatment in a population-based setting using the Swedish CML registry. Of 118 CML patients transplanted between 2002 and 2017, 56 (47.4%) received allo-HSCT in first CP, among whom TKI resistance was the most common transplant indication (62.5%). For patients diagnosed with CML in CP at65 years of age, the cumulative probability of undergoing allo-HSCT within 5 years was 9.7%. Overall 5-year survival was 96.2%, 70.1% and 36.9% when transplanted in first CP, second or later CP, and in accelerated phase or blast crisis, respectively. Risk factors for relapse were EBMT score2 and reduced intensity conditioning, and for death, CP2 at time point of allo-HSCT only. Non-relapse mortality for patients transplanted in CP was 11.6%. Our data indicate that allo-HSCT still constitutes a reasonable therapeutic option for patients with CML in first CP, especially those resistant to TKI treatment, providing high long-term survival and low non-relapse mortality.

Clinical trials show that tyrosine kinase inhibitor (TKI) treatment can be discontinued in selected patients with chronic myeloid leukaemia (CML). Although updated CML guidelines support such procedure in clinical routine, data on TKI stopping outside clinical trials are limited. In this retrospective study utilising the Swedish CML registry, we examined TKI discontinuation in a population-based setting. Out of 584 patients diagnosed with chronic-phase CML (CML-CP) in 2007-2012, 548 had evaluable information on TKI discontinuation. With a median follow-up of nine years from diagnosis, 128 (23%) discontinued TKI therapy (>= 1 month) due to achieving a DMR (deep molecular response) and 107 (20%) due to other causes (adverse events, allogeneic stem cell transplant, pregnancy, etc). Among those stopping in DMR, 49% re-initiated TKI treatment (median time to restart 4 center dot 8 months). In all, 38 patients stopped TKI within a clinical study and 90 outside a study. After 24 months 41 center dot 1% of patients discontinuing outside a study had re-initiated TKI treatment. TKI treatment duration pre-stop was longer and proportion treated with second-generation TKI slightly higher outside studies, conceivably affecting the clinical outcome. In summary we show that TKI discontinuation in CML in clinical practice is common and feasible and may be just as successful as when performed within a clinical trial.

n/a Funding Agencies|Lund University; Skane University Hospital

Objectives: The use of patient-reported outcome (PRO) measured outside clinical trials is not well defined. We report the first analysis of the prospective PRO study within the Swedish acute myeloid leukemia (AML) and the acute lymphoblastic leukemia (ALL) registries. Methods: PRO was requested 6 months after diagnosis. The EORTC Quality of life Questionnaire Core 30-item, the Patient Health Questionnaire-8 (PHQ-8), and questions from a Swedish National Cancer Questionnaire were used. Results: An invitation letter was sent to 398 patients; 255 (64%) responded, 60% web-based, and 40% on paper. The ALL cohort had lower physical, role and social functioning, higher symptom burden, and more financial difficulties compared to the AML cohort. A PHQ-8 score ≥ 10p, which indicates depression, was reported in 18% of the patients; 33% of these patients reported being prescribed antidepressants. The patients' overall experience of care was satisfying, but more psychological and practical support was desired. There was no difference in survival between patients who reported their PRO and those who did not. Follow-up at 2 and 4 years is ongoing. Conclusions: PRO collected in a registry-based setting is feasible, but the selection of time points and questionnaires are delicate in a diverse patient population. (Less)

TO THE EDITOR: Myeloid (or granulocytic) sarcoma (MS), that is, an extramedullary tumor consisting of myeloid blastic cells, is a unique clinical presentation of any type of acute myeloid leukemia (AML), according to the World Health Organization.[1][1] It could present (1) de novo (isolated MS), (

Objectives As new, effective therapies emerge for acute lymphoblastic leukaemia (ALL), the results of clinical trials need to relate to standard of care. Methods We used the population-based Swedish ALL Registry to evaluate characteristics, treatment and long-term outcome in 933 patients with diagnosis between 1997 and 2015. Results The median age was 53 years. The frequency of Philadelphia (Ph)-positive leukaemia was 34% of examined B-ALL with a peak incidence at 50-59 years. Five-year overall survival (OS) improved between 1997-2006 and 2007-2015; in patients 18-45 years from 50% (95% CI 43-57) to 65% (95% CI 58-72), 46-65 years from 25% (95% CI 18-32) to 46% (95% CI 37-55) and amp;gt;65 years from 7% (95% CI 2.6-11) to 11% (95% CI 5.9-16) (P amp;lt; 0.05). Men with Ph-neg B-ALL 46-65 years had inferior OS compared with women (P amp;lt; 0.01). Standardised mortality ratio was 5.7 (95% CI 5.0-6.3) for patients who survived 5 years from diagnosis. In multivariable analysis, Ph-positive disease was not associated with impaired prognosis but with lower risk of death in 2007-2015. Conclusions In a population-based cohort, OS has improved in adult ALL, especially for Ph-positive disease but for middle-aged men with Ph-negative B-ALL outcome was poor. Cure without late toxicity or relapse is still desired. Funding Agencies|Lions Cancer Foundation Uppsala; Uppsala University; Uppsala County Council/Region agreement on medical training and clinical research (ALF)

The primary goal in management of chronic phase (CP) chronic myeloid leukaemia (CML) is to prevent disease progression to accelerated phase (AP) or blast crisis (BC). We have evaluated progression rates in a decentralised healthcare setting and characterised patients progressing to AP/BC on TKI treatment.Using data from the Swedish CML register, we identified CP-CML patients diagnosed 2007-2011 who progressed to AP/BC within 2 yrs from diagnosis (n = 18) as well as patients diagnosed in advanced phase during 2007-2012 (n = 36) from a total of 544 newly diagnosed CML cases. We evaluated baseline characteristics, progression rates, outcome and adherence to guidelines for monitoring and treatment.The cumulative progression rate at 2 yrs was 4.3%. All 18 progression cases had been treated with imatinib, and six progressed within 6 months. High-risk EUTOS score was associated to a higher risk of progression. Insufficient cytogenetic and/or molecular monitoring was found in 33%. Median survival after transformation during TKI treatment was 1.4 yrs. In those presenting with BC and AP, median survival was 1.6 yrs and not reached, respectively.In this population-based setting, progression rates appear comparable to that reported from clinical trials, with similar dismal patient outcome. Improved adherence to CML guidelines may minimise the risk of disease progression.

Heterozygous mutations in GATA2 underlie different syndromes, previously described as monocytopenia and mycobacterial avium complex infection (MonoMAC); dendritic cell, monocytes, B- and NK lymphocytes deficiency (DCML); lymphedema, deafness and myelodysplasia (Emberger syndrome) and familiar myelodysplastic syndrome/acute myeloid leukemia (MDS / AML). Onset and severity of clinical symptoms vary and preceding cytopenias are not always present. We describe a case of symptomatic DCML deficiency and rather discrete bone marrow findings due to GATA2 mutation. Exome sequencing revealed a somatic ASXL1 mutation and the patient underwent allogeneic stem cell transplantation successfully., Highlights • Allogeneic stem cell transplantation was performed for DCML caused by GATA2 mutation. • Genetic diagnostics were done by Sanger sequencing and whole exome sequencing. • We identified an ASXL1 mutation associated with high risk for leukemic transformation.

Recent studies suggest that a proportion of chronic myeloid leukemia (CML) patients in deep molecular remission can discontinue the tyrosine kinase inhibitor (TKI) treatment without disease relapse. In this multi-center, prospective clinical trial (EURO-SKI, NCT01596114) we analyzed the function and phenotype of T and NK cells and their relation to successful TKI cessation. Lymphocyte subclasses were measured from 100 imatinib-treated patients at baseline and 1 month after the discontinuation, and functional characterization of NK and T cells was done from 45 patients. The proportion of NK cells was associated with the molecular relapse-free survival as patients with higher than median NK-cell percentage at the time of drug discontinuation had better probability to stay in remission. Similar association was not found with T or B cells or their subsets. In non-relapsing patients the NK-cell phenotype was mature, whereas patients with more naive CD56(bright) NK cells had decreased relapse-free survival. In addition, the TNF-alpha/IFN-gamma cytokine secretion by NK cells correlated with the successful drug discontinuation. Our results highlight the role of NK cells in sustaining remission and strengthen the status of CML as an immunogenic tumor warranting novel clinical trials with immunomodulating agents. Funding Agencies|Nordic Cancer Union; Finnish Society of Hematology; Biomedicum Helsinki Foundation; Research Foundation of Blood Diseases in Finland; Academy of Finland; Finnish Cancer Organizations; Signe and Ane Gyllenberg Foundation; Finnish Cancer Institute; Doctoral Programme in Clinical Research in the University of Helsinki

Musculoskeletal pain in patients with chronic myeloid leukemia after discontinuation of Imatinib : a Tyrosine Kinase Inhibitor withdrawal syndrome?

Although several cytokines have been demonstrated to be critical regulators of development of multiple blood cell lineages, it remains disputed to what degree they act through instructive or permissive mechanisms. Signaling through the FMS-like tyrosine kinase 3 (FLT3) receptor and the hematopoietin IL-7 receptor alpha (IL-7Ralpha) has been demonstrated to be of critical importance for sustained thymopoiesis. Signaling triggered by IL-7 and thymic stromal lymphopoietin (TSLP) is dependent on IL-7Ralpha, and both ligands have been implicated in T-cell development. However, we demonstrate that, whereas thymopoiesis is abolished in adult mice doubly deficient in IL-7 and FLT3 ligand (FLT3L), TSLP does not play a key role in IL-7-independent or FLT3L-independent T lymphopoiesis. Furthermore, whereas previous studies implicated that the role of other cytokine tyrosine kinase receptors in T lymphopoiesis might not involve permissive actions, we demonstrate that ectopic expression of BCL2 is sufficient not only to partially correct the T-cell phenotype of Flt3l(-/-) mice but also to rescue the virtually complete loss of all discernable stages of early T lymphopoiesis in Flt3l(-/-)Il7r(-/-) mice. These findings implicate a permissive role of cytokine receptors of the hematopoietin and tyrosine kinase families in early T lymphopoiesis.

Prolonged pre-transplant IFN administration in patients with chronic myeloid leukaemia has been associated with an increased risk of fatal transplant-related complications and an inferior outcome after allogeneic bone marrow transplantation. Using a two-stage long-term bone marrow culture as an in vitro model we were able to show a negative effect of higher doses of IFN pre-treatment of bone marrow stroma on haematopoesis. This correlated with a decline in the number of monocytes-macrophages. While monocytes-macrophages itself produce cytokines, a vicious circle may lead to an ineffective function of the microenvironment resulting in ineffective haematopoieses and increased transplant-related complications.

Phenotypically and functionally distinct progenitors and developmental pathways have been proposed to exist for fetally derived B-1 and conventional B-2 cells. Although IL-7 appears to be the primary cytokine regulator of fetal and adult B lymphopoiesis in mice, considerable fetal B lymphopoiesis and postnatal B cells are sustained in the absence of IL-7; in humans, B-cell generation is suggested to be largely IL-7–independent, as severe combined immune-deficient patients with IL-7 deficiency appear to have normal B-cell numbers. However, the role of other cytokines in IL-7–independent B lymphopoiesis remains to be established. Although thymic stromal lymphopoietin (TSLP) has been proposed to be the main factor driving IL-7–independent B lymphopoiesis and to distinguish fetal from adult B-cell progenitor development in mice, recent studies failed to support a primary role of TSLP in IL-7–independent fetal B-cell development. However, the role of TSLP in IL-7–independent adult B lymphopoiesis and in particular in regulation of B-1 cells remains to be established. Here we demonstrate that, rather than TSLP, IL-7 and FLT3 ligand are combined responsible for all B-cell generation in mice, including recently identified B-1–specified cell progenitors. Thus, the same IL-7– and FLT3 ligand–mediated signal-ing regulates alternative pathways of fetal and adult B-1 and B-2 lymphopoiesis.

The WHO classification (2008) defines "myeloid/lymphoid neoplasm with eosinophilia and rearrangement of PDGFRA, PDGFRB or FGFR1" as a rare subtype of myeloid neoplasms. Whilst patients with PDGFRA or PDGFRB rearrangements respond very well to imatinib, the optimal therapy for patients with FGFR1 rearrangements, which we refer to as FGFR1 fusion gene positive MLN-eo (FGFR1+ MLN-eo), remains to be defined. Encouraging in-vitro data using inhibitors of the FGFR1 tyrosine kinase prompted the implementation of ponatinib, which inhibits FGFR1, into therapeutical strategies. In a recent report, the clinical activity of ponatinib was reported in a single patient who concomitantly received high-dose chemotherapy and allogeneic stem cell transplantation (ASCT, Khodadoust et al, Leukemia 2015). We sought to evaluate efficacy of ponatinib in seven consecutive FGFR1+ MLN-eo patients. Median age was 52 years (range, 48-74) with a male predominance (n=5). Median observation time after diagnosis was 10 months (range, 5-36). All patients presented with left-shifted leukocytosis but only three patients [all with t(8;13)] had eosinophilia of >0.5 x 109/l. Bone marrow biopsy revealed a hypercellular marrow consistent with myeloproliferative neoplasm in all patients. Five patients presented with concomitantly diagnosed lymphoid neoplasms, i.e. T-lymphoblastic lymphoma (T-LBL, n=3), biclonal accelerated phase (n=1) or lymphoid blast phase of MPN/B-cell acute lymphoblastic leukemia (B-ALL, n=1). Cytogenetic analysis revealed a reciprocal translocation with involvement of chromosome band 8p11 in all patients [t(8;13)(p11;q12), n=3; t(8;22)(p11;q11), n=2; t(1;8;22)(?;p11;q11), n=1; t(6;8)(q27;p11), n=1]. On molecular level, RT-PCR identified the associated fusion genes ZMYM2-FGFR1 (n=3), BCR-FGFR1 (n=3), and FGFR1OP-FGFR1 (n=1), respectively. In one patient with T-LBL, the FGFR1 rearrangement was revealed by FISH analysis in 80% of lymph node cells indicating an origin of both MPN and T-LBL from the same progenitor/stem cell (myeloid/lymphoid stem cell neoplasm) and T-LBL as a feature of extramedullary lymphoid blast phase. All patients were initially treated with chemotherapy-based regimens including hydroxyurea (n=4) and/or high-dose chemotherapy (n=3), the latter exclusively in patients with concomitant aggressive lymphoid neoplasms. Lack of complete response, e.g. persisting features of MPN, relapse or progression led to the off-label use of ponatinib at a dose of 30mg/day (n=2) or 45mg/day (n=5). Median duration of treatment was 8 weeks (range, 2-52). A temporary partial hematologic response (control of peripheral blood cell count) was observed in 6 of 7 patients. One patient did not respond at all and died within a few weeks while on ponatinib due to progressive disease. Three of the 6 responders had cytogenetic analysis at a median of 3 months after the start of ponatinib. One patient with t(8;13) achieved a partial cytogenetic response (50% of metaphases positive after 3 months of treatment); in all other patients no cytogenetic response was observed. Four patients underwent ASCT and are in complete molecular remission and alive after a median time of 19 months (range, 8-36) after diagnosis and 13 months (range, 4-29) after ASCT. For one patient with BCR-FGFR1-positive MLN-eo without concomitant lymphoid disease ASCT is planned. One patient is on supportive care. Conclusion: Unexpectedly, response to standard dose ponatinib in FGFR1+ MLN-eo has been poor. There was either progressive disease or no evidence for sustained hematologic or cytogenetic response. However, there was also no evidence for a sustained complete remission on intensive chemotherapy in patients with full myeloid/lymphoid phenotype. Hence, ASCT currently remains the only option to achieve long-term remission and possibly cure in FGFR1+ MLN-eo. Disclosures Off Label Use: ponatinib, used as FGFR1 inhibitor. Bommer:Alexion Pharmaceuticals: Honoraria. Cross:Ariad: Consultancy, Honoraria, Research Funding; Qiagen: Consultancy, Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding. Hochhaus:Bristol-Myers Squibb: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding.

Background: Tyrosine kinase inhibitors (TKIs) have significantly improved the treatment of CML. Even though TKI treatment is generally not considered curative, recent studies have shown that nearly half of CML patients who have achieve good and durable responses are able to stop the TKI treatment. However, patients who have successfully discontinued TKI treatment still have residual disease. We hypothesized that the immune system plays a role in treatment free remission (TFR), and our preliminary results in the EURO-SKI trial showed that patients who relapse early after imatinib discontinuation have decreased numbers and frequencies of NK cells. In EURO-SKI trial relapse was defined as the loss of major molecular response (MMR). We now aimed to analyze in more detail the phenotype and function of the NK cells in order to understand their role in TFR. Methods: Lymphocyte subclass analysis (the number of NK-, T- and B-cells) was performed at the time of therapy discontinuation and 1 month after the imatinib discontinuation in patients participating in the EURO-SKI stopping trial in the Nordic countries (n=105, results are presented from patients who have reached 6 months follow-up). More detailed immune phenotype and functional assays (NK-cell degranulation and secretion of Th1 type of cytokines IFN-γ/TNF-α) were analyzed from a proportion of patients (n=31). Results: Imatinib treated patients remaining in remission for 6 months (non-relapsing, n=48, median age 60,5 years) displayed an increased amount of NK cells at the time of drug discontinuation (18.6% vs. 11.0%, p=0.02, NK-cell count 0.25 x109 cells/L vs. 0.184 x109 cells/L m, p=0.059) compared to patients who relapsed early (before 5 months, n=29, median age 60,5 years). Furthermore, the NK cell frequency in non-relapsing patients was even higher than in healthy controls (11.5%, n=48, p=0.001). T and B cell counts and frequencies showed no differences between the groups. Detailed analysis of the NK cell compartment displayed a more mature phenotype for the NK cells in non-relapsing patients. Larger frequencies of NK cells from early relapsing patients was CD56bright compared to non-relapsing patients (4.8% vs. 2.7% of CD56 NK cells, p=0.04). Furthermore, patients who had higher frequencies of CD56bright NK cells than median had decreased TFR at 6 months (42%) compared to patients with lower frequency (70%, p=0.01). In addition, there was a trend towards more CD57pos (78% (n=21) vs. 66% (n=10), p=0.09) CD56dim NK cells in non-relapsing patients. To further study the mature NK cells in non-relapsing patients, recently identified markers (FceRgneg, PLZFneg, SYKneg, EAT-2neg) for adaptive NK cells were analyzed. Interestingly, there was a trend that non-relapsing patients had higher frequencies of adaptive-like NK cells. For example, non-relapsing patients had more CD56dim NK cells that had down regulated EAT-2 (2.8% (n=6) vs. 1.3% (n=5) of lymphocytes, p=0.03) and more CD56dim NK cells expressing NKG2D (11.2% vs. 2.6% of lymphocytes, p=0.02) and NKp46 (13.6% vs. 3.9% of lymphocytes, p=0.05). Moreover, after imatinib discontinuation the expression of transcription factor Eomes increased in the CD56dim NK cells of the early relapsing group (baseline MFI 2045 vs. 1 month 3480, p=0.06), while in non-relapsing group it seemed to even decrease (baseline MFI 2273 vs. 1 month 1980, p=0.13) pointing towards an adaptive phenotype. No significant differences between the groups were observed when degranulation against K562 cell line was studied. However, CD16neg NK cells from non-relapsing patients responded to K562 stimulation by secreting more TNFα/IFNγ compared to the early relapsing patients (21% vs. 13% of CD56pos CD16neg NK cells, p=0.01). Furthermore, patients whose CD16neg NK cells had higher than median TNFα/IFNγ secretion when stimulated with K562 cells showed an increased TFR at 6 months (78%) compared to patients who had lower TNFα/IFNγ secretion than median (37%, p=0.005). Conclusions: CML patients who successfully discontinued imatinib therapy displayed a higher number and frequency of peripheral blood mature, adaptive-like NK cells capable of secreting cytokines TNFα/IFNγ relative to relapsing patients. How such NK cells may contribute to maintenance of treatment free remission is still unknown. Nonetheless, our results warrant further clinical studies with NK-cell modulating agents. Disclosures Muller: Novartis: Honoraria, Other: Consulting or Advisory Role, Research Funding; ARIAD Pharmaceuticals Inc.: Honoraria, Other: Consulting & Advisory Role, Research Funding; BMS: Honoraria, Other: Consulting or Advisory Role, Research Funding. Hjorth-Hansen:Novartis: Honoraria; Ariad: Honoraria; Bristol-Myers Squibb: Research Funding; Pfizer: Honoraria, Research Funding. Saussele:Pfizer: Honoraria, Other: Travel grant; BMS: Honoraria, Other: Travel grant, Research Funding; Novartis Pharma: Honoraria, Other: Travel grant, Research Funding; ARIAD: Honoraria. Mahon:ARIAD: Consultancy; Novartis: Consultancy, Honoraria; Bristol-Myers Squibb: Consultancy, Honoraria; Pfizer: Consultancy. Porkka:Bristol-Myers Squibb: Honoraria; Celgene: Honoraria; Novartis: Honoraria; Pfizer: Honoraria. Richter:Ariad: Honoraria; Bristol-Myers Squibb: Honoraria; Novartis: Honoraria. Mustjoki:the Finnish Cancer Societies: Research Funding; Pfizer: Honoraria, Research Funding; Academy of Finland: Research Funding; Sigrid Juselius Foundation: Research Funding; Finnish Cancer Institute: Research Funding; Signe and Ane Gyllenberg Foundation: Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

Rationale: Dasatinib (DAS) and interferon have different modes of action and may have synergistic activity in CML, due to both antineoplastic and immunostimulatory mechanisms. Addition of pegylated interferon (PegIFN) to imatinib therapy in CP-CML has in previous clinical trials (French SPIRIT and NordCML002) resulted in deeper molecular responses. Thus, an optimal combination of DAS and PegIFN may increase the proportion of patients who reach deep molecular response with potential for treatment-free remission (TFR). Design: Newly diagnosed CP-CML patients were treated with DAS (Sprycel, BMS) 100 mg OD as single drug for three months. Thereafter weekly subcutaneous injections of Peg-IFN α2b (PegIntron, MSD) were added to DAS; from end of month 3 (M3) to M6, 15µg/week, thereafter 25µg/week until M15. Primary end points were safety and the rate of MMR at M12. The doses of PegIFN were lower than in the SPIRIT and NordCML002 studies to increase adherence. Population: Forty patients were included at 14 university centers. One patient was lost to follow-up after M6. All patients were included in analysis up to M12. Mean and median age was 48 years (range 19-71). The proportions of high risk patients were 25% (Sokal), 15% (Hasford), and 15% (EUTOS). Safety and dosing: Treatment was well tolerated with expected DAS and PegIFN related side effects. Six patients had seven serious adverse events (AEs), all hospitalizations. 1 episode each of bradycardia/atrial fibrillation (possibly PegIFN-related), headache (DAS), fever (PegIFN), anaphylaxis-like reaction (PegIFN), fever/malaise/headache (PegIFN), pneumonia and a knee effusion (both unrelated). One pleural effusion occurred (grade 2, 3%). Grade 3-4 neutropenia and thrombocytopenia occurred in 6 and 9 patients respectively. Prolonged hematological toxicity (>2 months) occurred in 8 patients, causing dosing problems in 5. One patient suffered grade 3 depression. Grade 3 flu-like symptoms occurred in 2 patients. One patient had lipase elevation grade 3 and one patient developed hypothyroidism attributed to PegIFN. Grade 2 dermal AEs like rash and acne occurred in about 20%, attributable to both drugs. 94% (DAS) and 76% (PegIFN) of assigned dose was given. Dose reductions occurred in 19 patients for DAS and 20 patients for PegIFN. Two patients discontinued DAS and switched to nilotinib, 1 for headache at M3 and 1 at M12 for lack of efficacy/hematological toxicity. Two patients could not start PegIFN for hematological toxicity (one lost to follow-up after M6). PegIFN was discontinued because of bradycardia/atrial fibrillation (1 patient), anaphylaxis (1 patient), flu-like syndrome (2 patients) and long-term hematological toxicity (2 patients). At 12 months 31/38 pats (82%) were still on PegIFN, a higher proportion than in the French Spirit or NordCML002 studies. Efficacy: We have used the DAS arm of the Dasision study (Kantarjian NEJM 2010) as a historical control. Early response at M3 was very similar between studies. In the present and the Dasision cohorts respectively, 18% vs 16% missed the 10% BCR-ABLIS landmark, 66% vs 56% achieved a CCyR and 8% vs 8% achieved MMR. At M6, three months after introduction of PegIFN, a steep increase in MMR rate was observed compared with Dasision. This was also reflected in deep responses, MR4.0 (see tables) and MR4.5 at M12, 18% vs 5%. The primary efficacy endpoint was MMR at M12, 82% vs 46%. Table 1.MMRDAS+PegIFN (%)DAS (Dasision)(%)Difference (%)M3880M6532726M9663927M12824636Table 2.MR4.0DAS+PegIFN (%)DAS (Dasision) (%)Difference (%)M3303M620614M938830M12481236 Progressions and treatment failure defined by ELN 2013: Failures: No progression was noted. At M3, 2 patients still had >95% Ph+ metaphases (MF). At M6, four patients (11%) had > 35% Ph+MF or >10% BCR-ABL levels. At M12, one patient failed CCgR and two more patients failed Conclusion: The combination of DAS and low dose PegIFN could be safely administered in newly diagnosed CP CML. No unexpected autoimmune phenomena were observed, and pleural effusions were rare. Efficacy appears very promising with high early MMR rates and deep molecular responses. A randomized comparison DAS +/- PegIFN is warranted. Support: Study drug from BMS and MSD. Grant from BMS. Figure 1. Figure 1. Disclosures Hjorth-Hansen: Ariad: Honoraria; Novartis: Honoraria; Pfizer: Honoraria, Research Funding; Bristol-Myers Squibb: Research Funding. Off Label Use: Dasatinib and Pegylated IFN combination in CML. Richter:Ariad: Honoraria; Bristol-Myers Squibb: Honoraria; Novartis: Honoraria. Porkka:Bristol-Myers Squibb: Honoraria; Celgene: Honoraria; Novartis: Honoraria; Pfizer: Honoraria. Mustjoki:Bristol-Myers Squibb: Honoraria, Research Funding.

Background: Recent reports suggest that approximately 40% of CML patients who have achieved sustained complete molecular remission are able to stop TKI treatment without disease relapse. However, there are no predictive markers for successful therapy discontinuation. Therefore, we set up an immunological sub-study in the ongoing pan-European EURO-SKI stopping study. Our aim was to identify predictive biomarkers for relapse/non-relapse and to understand more on the mechanisms of immune surveillance in CML. Methods: The EURO-SKI study started in 2012, and patients included were at least three years on TKI and at least one year in MR4 or deeper before the study entry. Basic lymphocyte immunophenotyping (the number of NK-, T- and B-cells) was performed at the time of therapy discontinuation and 1, 6, and 12 months after the TKI stop and in case of relapse (defined as loss of MMR, BCR-ABL1>0.1% IS). In addition, from a proportion of patients more detailed immunophenotypic and functional analyses (cytotoxicity of NK-cells and secretion of Th1 type of cytokines IFN-γ/TNF-α) were done at the same times. Results: Thus far 119 Nordic patients (imatinib n=105, dasatinib n=12, nilotinib n=2) who have discontinued TKI treatment within the EURO-SKI study have been included in the lymphocyte subclass analysis (results are presented from patients who have reached 6 months follow-up). Immunophenotyping analysis demonstrates that imatinib treated patients who were able to maintain remission for 6 months (n=36) had increased NK-cell counts (0.26 vs. 0.15x109cells/L, p=0.01, NK-cell proportion 18.9% vs. 11%, p=0.005) at the time of drug discontinuation compared to patients who relapsed early (before 5 months n=22). Furthermore, the phenotype of NK-cells was more cytotoxic (more CD57+ and CD16+cells and less CD62L+cells), and also their IFN-γ/TNF-α secretion was enhanced (19.2% vs. 13%, p=0.02). Surprisingly, patients who relapsed more slowly (after 5 months, n=16) had similar baseline NK-cell counts (0.37x109cells/L), NK-cell proportion (21.2%), and phenotype and function as patients, who were able to stay in remission. No differences in the NK-cell counts were observed between patients who had detectable or undetectable BCR-ABL1 transcripts at the baseline (0.22 x109cells/L vs. 0.31 x109cells/L, p=0.61). Interestingly, NK-cell count was higher in patients with low Sokal risk score than in patients with intermediate risk (0.33 x109cells/L vs. 0.20 x109cells/L, p=0.04). Furthermore, there was a trend that male patients had a higher proportion of NK-cells than females (21.6% vs. 15.7%, p=0.06). Pretreatment with IFN-α or the duration of imatinib treatment did not have an effect on NK-cell count or proportion. In comparison to the imatinib group, dasatinib treated patients had higher NK-cell counts at the baseline (median 0.52x109cells/L vs. 0.26x109cells/L, p=0.02), and also the proportion of CD27 (median 50% vs. 16%, p=0.01) and CD57 expressing (median 79% vs. 74%, p=0.05) NK-cells was higher. The follow-up time of dasatinib treated patients is not yet long enough to correlate the NK-cell counts with the success of the treatment discontinuation. The absolute number of T-cells or their function did not differ significantly between relapsing and non-relapsing patients at the time of treatment discontinuation. However, both CD4+ and CD8+ T-cells tended to be more mature in patients who stayed in remission compared to patients who relapsed early (CD4+CD57+CD62L- median 5.7% vs. 2.4%, p=0.06, CD8+CD62L+CD45RA+ 13% vs. 26.7%, p=0.05). The analysis of follow-up samples showed that in patients who stayed in remission the Th1 type cytokine (IFN-γ/TNF-α) secretion of CD8+T-cells increased at 6 months compared to baseline (23.6 vs. 18.5%, p=0.07). Same phenomenon was observed in the late relapsing group at relapse compared to baseline (37.9 vs. 13.5%, p=0.03). No similar increase was observed in the early relapsing group. Conclusions: Low NK-cell numbers and poor cytokine secretion may predict early disease relapse after TKI discontinuation. However, patients who relapse later have high numbers of normally functioning NK-cells. Further research (detailed phenotypic analysis of NK- and T-cells including activating and inhibitory receptors and immune checkpoint molecules) and correlation of biomarker data with clinical parameters are ongoing to understand the ultimate determining factors of relapse. Disclosures Själander: Novartis: Honoraria. Hjorth-Hansen:Novartis: Honoraria; Bristol-myers Squibb: Honoraria; Ariad: Honoraria; Pfizer: Honoraria. Porkka:BMS: Honoraria; BMS: Research Funding; Novartis: Honoraria; Novartis: Research Funding; Pfizer: Research Funding. Mustjoki:Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

Background The inhibition of oncogenic BCR-ABL1 kinase with tyrosine kinase inhibitors (TKIs) has significantly improved the prognosis of CML. Recent reports suggest that approximately 40 % of CML patients who have achieved optimal therapy response (complete molecular remission, CMR) can stop imatinib treatment without recurrence of detectable BCR-ABL1 transcripts. However, no predictive prognostic factors for successful therapy discontinuation have yet been identified. We therefore set up an immunological substudy in the ongoing pan-European EURO-SKI stopping study. We aimed to identify predictive biomarkers for relapse and non-relapse after TKI discontinuation. In addition, we aimed to understand more on the mechanisms of immune surveillance in CML and to study the effects of TKI treatment on the immune system. Materials and methods Patients in deep molecular remission (MR4, BCR-ABL < 0,01% IS) for at least one year and with TKI treatment for at least 3 years were eligible for the clinical study. Basic lymphocyte immunophenotyping (the proportions and absolute numbers of NK-, T- and B-cells) was performed at the university hospital laboratories at the time of therapy discontinuation, and 1, 6, and 12 months after the TKI discontinuation. In a proportion of patients a more detailed immunophenotypic (analysis of CD45RA, CD57, CD27 and CD62L expressions) and functional analyses were done from fresh blood samples in a central immunology laboratory (Helsinki) at the same time points. The cytotoxicity of NK-cells was studied by measuring the direct killing of target cells (K562) and by the degranulation assay (CD107a/b expression). The secretion of Th1 type of cytokines IFN-γ/TNF-α was studied from both T- and NK-cells. Results Thus far the basic lymphocyte subclass measurement has been analyzed from 62 patients who have discontinued TKI treatment within the EURO-SKI study. Functional analyses have been performed from 30 patients. 60 patients have used imatinib before treatment discontinuation and 2 patients dasatinib. At baseline, before the treatment discontinuation both CD4+ and CD8+ T-cell counts were within the normal range (median CD4+ 0.73x 109/L, range 0.11-2.4x 109/L; CD8+ 0.35x 109/L, 0.07-1.92 x 109/L). The TKI stop had no significant numerical or functional effect on T-cells, and at 1 month time-point the median T-cell counts were unchanged (CD4+ 0.73x 109/L; CD8+ 0.35x 109/L). Similarly, at the baseline, the median NK-cell count was within a normal range (0.26 x 109/L, range 0.04-1.04 x 109/L), and no significant change was observed 1 month after stopping the treatment (median 0.29 x 109/L). Furthermore, at the baseline and at the 1-month time-point the cytotoxicity of NK-cells and the cytokine secretion of T- and NK-cells did not significantly differ from the healthy controls when all patients were considered as a one group. However, when patients were divided in two groups based on the relapse status, the patients who eventually relapsed had significantly fewer NK-cells already at the baseline (Figure A; absolute count 0.18x 109/L vs. 0.32 109/L, p=0.008; proportions 11% vs. 21%, p=0.001). The phenotype of NK-cells also differed between the two groups, and the patients who relapsed had less NK-cells expressing CD57 (median 58% vs. 69%, p=0.046) and CD16 (median 67% vs. 83%, p=0.018) on the cell surface. Furthermore, the cytotoxicity of NK-cells was impaired in patients who failed to discontinue the TKI treatment successfully and no killing activity was observed in their samples (Figure B; alive K652 cells after co-incubation with effector cells 100% vs. 88%, p=0.07). No clear differences were observed in the function or the numbers of T-cells between relapsing and non-relapsing patients. Conclusions The NK-cell numbers and their function may predict disease relapse after TKI discontinuation. This may have impact on the future stopping trials. In addition, it further illustrates the importance of the immune system in the successful long-term treatment of CML. Disclosures: Ekblom: Novartis: Honoraria; Bristol-Myers Squibb: Honoraria. Hjorth-Hansen:Pfizer, BMS: Honoraria, Travel expenses Other. Porkka:BMS: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Richter:Bristol-Myers Squibb: Consultancy, Speakers Bureau; Novartis: Consultancy, Research Funding, Speakers Bureau. Mustjoki:Novartis: Honoraria; BMS: Honoraria, Research Funding.