Letters to the Editor 725 1 Recombinant Protein Unit, Institut Pasteur de Montevideo, 6 Sevov M, Kaderi M, Kanduri M, Mansouri, Buhl A, Cahill N et al. A comparative Montevideo, Uruguay; study of RNA-based markers in chronic lymphocytic leukemia reveals LPL as a 2 Immunology Department, Institute of Experimental Medicine (IMEX)- powerful predictor of clinical outcome. Haematologica 2009; 94(Suppl 3): 1–95. CONICET, National Academy of Medicine, Buenos Aires, Argentina; 7 Van Bockstaele F, Pede V, Janssens A, Callewaert F, Offner F, Verhasselt B et al. 3Biophysics Protein Unit, Institut Pasteur de Montevideo, Lipoprotein lipase mRNA expression in whole blood is a prognostic marker in B Montevideo, Uruguay; cell chronic lymphocytic leukemia. Clin Chem 2007; 53: 204–212. 4Service of Hematology and Bone Marrow Transplantation, 8 Kaderi MA, Kanduri M, Buhl AM, Sevov M, Cahill N, Gunnarsson R et al. Hospital Maciel, Montevideo, Uruguay; LPL is the strongest prognostic factor in a comparative analysis of RNA-based 5 markers in early chronic lymphocytic leukemia. Haematologica 2011; 96: Epigenetics of Cancer and Aging Laboratory, Institut Pasteur de 1153–1160. Montevideo, Montevideo, Uruguay; 9 Kolset SO, Salmivirta M. Cell surface heparan sulfate proteoglycans and 6 Molecular Virology Laboratory, School of Science, Universidad de la lipoprotein metabolism. Cell Mol Life Sci 1999; 56: 857–870. Repu´blica, Montevideo, Uruguay and 10 Pallasch CP, Schwamb J, Konigs S, Schulz A, Debey S, Kofler D et al. Targeting lipid 7Immunobiology Department, School of Medicine, Universidad metabolism by the lipoprotein lipase inhibitor orlistat results in apoptosis of B-cell de la Repu´blica, Montevideo, Uruguay chronic lymphocytic leukemia cells. Leukemia 2008; 22: 585–592. E-mail: [email protected] 11 Brenet F, Moh M, Funk P, Feierstein E, Viale AJ, Socci ND et al. DNA methylation of 8These authors contributed equally to this work. the first exon is tightly linked to transcriptional silencing. PLoS One 2011; 6: e14524. 12 Watanabe M, Ogawa Y, Itoh K, Koiwa T, Kadin ME, Watanabe T et al. Hypomethylation of CD30 CpG islands with aberrant JunB expression drives CD30 REFERENCES induction in Hodgkin lymphoma and anaplastic large cell lymphoma. Lab Invest 1 Dighiero G, Hamblin TJ. Chronic lymphocytic leukaemia. Lancet 2008; 371: 2008; 88:48–57. 1017–1029. 13 Bilban M, Heintel D, Scharl T, Woelfel T, Auer MM, Porpaczy E et al. 2 Caligaris-Cappio F. Role of the microenvironment in chronic lymphocytic Deregulated expression of fat and muscle genes in B-cell chronic lymphocytic leukaemia. Br J Haematol 2003; 123: 380–388. leukemia with high lipoprotein lipase expression. Leukemia 2006; 20: 3 Zenz T, Fulda S, Stilgenbauer S. More (on) prognostic factors in chronic 1080–1088. lymphocytic leukemia. Leuk Lymphoma 2010; 51: 5–6. 14 Burger JA. Chemokines and chemokine receptors in chronic lymphocytic leuke- 4 Oppezzo P, Vasconcelos Y, Settegrana C, Jeannel D, Vuillier F, Legarff-Tavernier M mia (CLL): from understanding the basics towards therapeutic targeting. Semin et al. The LPL/ADAM29 expression ratio is a novel prognosis indicator in chronic Cancer Biol 2010; 20: 424–430. lymphocytic leukemia. Blood 2005; 106: 650–657. 15 Palacios F, Moreno P, Morande P, Abreu C, Correa A, Porro V et al. High expression 5 Heintel D, Kienle D, Shehata M, Krober A, Kroemer E, Schwarzinger I et al. High of AID and active class switch recombination might account for a more aggressive expression of lipoprotein lipase in poor risk B-cell chronic lymphocytic leukemia. disease in unmutated CLL patients: link with an activated microenvironment in Leukemia 2005; 19: 1216–1223. CLL disease. Blood 2010; 115: 4488–4496.

Supplementary Information accompanies the paper on the Leukemia website (http://www.nature.com/leu)

A phase 1 study of concomitant high-dose lenalidomide and 5-azacitidine induction in the treatment of AML

Leukemia (2013) 27, 725–728; doi:10.1038/leu.2012.214 the findings of our own institutional phase 1 prospective trial of AZA and high-dose lenalidomide (HDL) as an induction regimen followed by maintenance therapy with standard dose AZA and Lenalidomide at a dose of 10 mg daily is approved for treatment lower dose of lenalidomide. In contrast to Pollyea et al.,6 we of low risk/intermediate risk-1 myelodysplasia (MDS) patients who (1) used escalating doses of AZA while keeping the lenalidomide are red cell transfusion dependent.1,2 We recently reported on an dose constant, (2) used concomitant dosing of AZA and HDL and institutional phase 2 trial treating newly diagnosed elderly acute (3) chose to reduce the dose of lenalidomide after the initial two myeloid leukemia (AML) patients with high-dose single agent cycles of therapy. lenalidomide as an induction treatment (50 mg/day  28 days) Newly diagnosed elderly AML patients X60 years of age with followed by lower dose maintenance (10 mg daily for 12 months). intermediate- or poor-risk cytogenetics, without isolated 5q Thirty percent of patients achieved a complete remission (CR) or abnormalities (elderly AML) and relapsed/refractory AML X18 complete remission with incomplete blood count recovery (CRi) of years of age (relapsed AML) were eligible for this study. 30%.3,4 We and others have hypothesized that the combination of Additional inclusion criteria included: Eastern Cooperative lenalidomide and azacitidine (AZA) may result in higher rates of Oncology Group performance status of 0–2, and adequate renal sustained CR compared with these drugs individually. Sekeres (serum creatinine o1.5  upper limit of normal) and hepatic 5 et al. showed feasibility and efficacy of low-dose lenalidomide function (bilirubin o2.0 mg/dl and aspartate aminotransferase/ (10 mg daily) with AZA in high-risk MDS. We read the article alanine transaminase o5  upper limit of normal). Patients with ‘Safety, efficacy and biological predictors of response to sequential acute promyelocytic leukemia, central nervous system leukemia, AZA and lenalidomide for elderly patients with acute myeloid or prior use of lenalidomide or AZA were excluded. The treatment leukemia’ by Pollyea et al.6 with interest. In that article, the authors schedule consisted of two 28-day induction cycles with HDL, 50 mg report their experience with a phase 1 trial using sequential orally for days 1–28 and AZA given intravenously for days 1–5 therapy with AZA and lenalidomide for elderly patients with AML. at three-dose cohorts (cohort 1, 25 mg/m2 (cohort 1), 50 mg/m2; In the trial, the authors escalated the dose of lenalidomide to cohort 2 and cohort 3, 75 mg/m2). Thereafter, patients were given 50 mg/day with AZA dose fixed at 75 mg/m2. Here we report on maintenance cycles (every 28 days) with lenalidomide 10 mg

Accepted article preview online 25 July 2012; advance online publication, 17 August 2012

& 2013 Macmillan Publishers Limited Leukemia (2013) 718 – 757 726

ekma(03 1 757 – 718 (2013) Leukemia Table 1. Clinical charactersitics and response of the 19 patients treated

Cohort Patient Diagnosis Sex Age WBC Karyotype Prior chemotherapy Pre-Tx Cycles BM BM Days Best Reason for (years) count BM completed blast blast on response treatment ( Â 1012 blast post post study discontinuation cell/l) 2I 2M

1 1 New elderly F 74 20.9 46,XX None 76 2 52 NA 79 PR Progressive AML disease 2 Refractory F 72 1.5 47,XX, þ 8 Decitabine 23 17 (2I þ 15M) 1 0 566 CR Continuing on AML study 3 Relapsed F 64 5.8 46,XX,del7q 7 þ 3, high-dose 22 o1 NA NA 8 PD DLT AML cytarabine 4 Relapsed M 72 2.3 46,XY Decitabine, CLAG 2 o1 NA NA 14 PD Patient AML/ prior withdrew MDS from study 5 Relapsed F 64 0.9 46,XX 7 þ 3, high-dose 58 o1 NA NA 14 PD Patient AML cytarabine withdrew from study 6 New elderly M 70 2.2 46,XY, À 7 None 6 7 (2I þ 5M) 3 2 218 SD Patient

AML/ prior withdrew Editor the to Letters MDS from study 7 Relapsed F 65 NA Complex 7 þ 3, HiDAC, 78 0 NA NA N/A NA Did not start AML Clofarabine, Decitabine treatment 8 New elderly F 68 4.5 46,XX None 81 1 NA NA 24 SD Patient AML withdrew from study 9 Relpased M 74 2.2 46,XY 7 þ 3, high-dose 11 5 (2I þ 3M) 45 12 171 SD Patient AML cytarabine withdrew from study 2 10 New elderly M 77 0.8 Complex LBH þ Decitabine, 2a 6 (2I þ 4M) 5 5 198 CRi Progressive AML/ prior Decitabine disease MDS 11 Relapse M 63 NA Complex 7 þ 3 þ plerixafor 22 0 NA NA 23 TF Decided to go AML to hospice 12 New elderly M 65 2.4 46,XY Decitabine 20-30 3 (2I þ 1M) 12 NA 139 PR Progressive AML/ prior disease MDS 13 Refractory F 70 3.3 45,XX, À 77þ 3 23 2 34 NA 77 PD Progressive AML disease 3 14 New elderly M 75 6.6 46,XY,inv9 None 17 4 (2I þ 2M) 0 0 155 CR Continuing on AML/ prior study MDS & 15 New elderly F 76 4.5 46,XX Decitabine 10 o1 NA NA 18 SD Patient 03McilnPbihr Limited Publishers Macmillan 2013 AML/ prior withdrew MDS from study 16 Relapsed F 81 1.1 46,XX Aurora kinase inhibitor, 10 4 (2I þ 2M) 4 7 226 CR Continuing on AML Decitabine þ bexoratene study 17 Relapsed F 74 0.9 Complex LBH þ Decitabine, 31 3 (2I þ 1M) 18 NA 144 PR Progressive AML Decitabine disease 18 New elderly M 77 1.5 46,XY,del20q None 63 1 NA NA 62 PD Progressive AML disease 19 New elderly M 65 0.9 47,XY, þ 2,del20 None 16 2 8 NA 65 PD Progressive AML disease Abbreviations: AML, acute myeloid leukemia; BM, bone marrow; CLAG, cladribine cytarabine and GCSF-based chemotherapy; CR, complete remission; CRi, complete remission with partial count recovery; DLT, dose-limiting toxicity; I, induction cycle; LBH, panobinostat; M, maintenance cycle; MDS, myelodysplasia; NA, not available; PD, progressive disease; PR; partial remission; Pre-Tx, pre treatment; SD, stable disease; WBC, white blood cell; 7 þ 3, cytarabine and idarubicin-based induction. aExtramedullary disease. Letters to the Editor 727 Table 2. Grade 3/4 toxicity

Grade Cohort 1 Cohort 2 Cohort 3

3453345

Cardiovascular Edema 1 Hypertension 1 Hypotension 3

Constitutional Fatigue 12 Fever—no infection 2 Hemorrhage 1 Itching 1 Weakness 2 Wound 1

Dermatological Rash 1 (DLT) 2

Gastrointestinal Dehydration 1

Hepatic Bilirubin 1 1

Infection Infection 5 1 1 2 2

Metabolic/laboratory Hypocalcemia 1 Hypokalemia 1 Natremia high 1 Pt/Ptt 1

Neurological Generalized pain 3

Pulmonary Cough 1 Dyspnea (sob) 2 2 Hypoxia 2 1 Pneumonitis/pulmonary infiltrates 2

Renal/genito-urinary Acute renal failure 1 1 Creatinine 1 1

Other Leukemia cutis 1 Abbreviation: DLT, dose-limiting toxicity. orally for days 1–28 and AZA 75 mg/m2 intravenously for days 1–5 during maintenance therapy. All adverse events were evaluated intravenously for up to 12 cycles. Lenalidomide and AZA were and coded using the National Cancer Institute Common provided by the manufacturer (Celgene Corporation, Summit, NJ, Terminology Criteria for Adverse Events, Version 4.0. Responses USA). All patients except those with progressive disease after the were assessed using the International Working Group criteria.7 The two induction cycles continued with the maintenance cycles. primary endpoint was to determine the maximum tolerated dose Induction cycles were not interrupted nor were doses modified for (MTD) and toxicity profile. expected hematological toxicities, but therapy could be inter- A total of 19 patients were enrolled in the study (Table 1). The rupted for febrile neutropenia, grade X3 nonhematological median age was 72 years (range 63–81); 9 males (47.4%), 10 toxicities or bone marrow aplasia. During maintenance cycles, females (52.6%); 10 (52.6%) with newly diagnosed elderly AML doses were delayed or reduced for febrile neutropenia, grade X3 and 9 (47.4%) with relapsed or refractory AML. The median WBC and prolonged hematological toxicities. Patients continued to count was 2200 Â 109 cell/l (range 800 Â 109–20 900 Â 109 cell/l). receive maintenance cycles until an unacceptable adverse event The median bone marrow blast percentage was 23% (range or disease progression. Growth factors were not administered 2–81%). Cytogenetics showed a normal karyotype in 8 (42.1%), a routinely but were allowed as clinically indicated. Response was complex karyotype in 4 (21.1%), monosomy 7/del 7q in 3 (15.7%), assessed by bone marrow biopsy and complete blood count at trisomy 8 in 1 (5.2%) and other clonal abnormalities in 3 (15.7%). the completion of induction cycles 1 and 2, after completion of Patients in each cohort who did not complete one induction cycle maintenance cycles 2, 4, 6 and 12 and as clinically indicated were replaced. In all, 13 (68.4%) patients completed at least one

& 2013 Macmillan Publishers Limited Leukemia (2013) 718 – 757 Letters to the Editor 728 induction cycle, 11 (57.9%) patients completed two induction G Ramsingh, P Westervelt, AF Cashen, GL Uy, cycles and 8 (42.1%) patients went on to receive maintenance K Stockerl-Goldstein, CN Abboud, N Bernabe, R Monahan, therapy. Patients remained on therapy for a median of 77 days JF DiPersio and R Vij (range 0–566). Section of Bone Marrow Transplant and Leukemia, Dose-limiting toxicities (DLT) observed included grade 3 rash Washington University in St Louis, St Louis, MO, USA in cohort 1 leading to expansion of the cohort to include three E-mail: [email protected] additional patients (Table 2). To date, grade 3/4 non-DLT hematological toxicity was seen in 12 (92.3%) out of 13 patients. The most common 3/4 non-DLT hematological toxicity was REFERENCES thrombocytopenia seen in 10 (76.9%) out of 13 patients. The most 1 List A, Kurtin S, Roe DJ, Buresh A, Mahadevan D, Fuchs D et al. Efficacy of lenali- common grade 1/2 toxicity was fatigue in 6 (42.9%) out of 14. None domide in myelodysplastic syndromes. N Engl J Med 2005; 352: 549–557. of the 6 patients in cohort 3 developed DLT, and hence the MTD 2 Raza A, Reeves JA, Feldman EJ, Dewald GW, Bennett JM, Deeg HJ et al. Phase 2 study was not reached in this study. Of the 13 patients, 7 (53.8%) patients of lenalidomide in transfusion-dependent, low-risk, and intermediate-1 risk myelo- have died, all owing to treatment failure due to resistant disease. dysplastic syndromes with karyotypes other than deletion 5q. Blood 2008; 111:86–93. Of the 13 evaluable patients, 7 (53.8%) responded to treatment, 3 Fehniger TA, Larson S, Trinkaus K, Siegel MJ, Cashen AF, Blum KA et al. A phase 2 multicenter study of lenalidomide in relapsed or refractory classical Hodgkin with CR/CRi in 4 (30.8%) and partial remission in 3 (23.1%), with a lymphoma. Blood 2011; 118: 5119–5125. median duration of response is 3 months (range 1–17). 4 Fehniger TA, Byrd JC, Marcucci G, Abboud CN, Kefauver C, Payton JE et al. Single- 6 Our data complements the findings of Pollyea et al. and agent lenalidomide induces complete remission of acute myeloid leukemia in demonstrates that the combination of HDL and AZA sequentially patients with isolated trisomy 13. Blood 2009; 113: 1002–1005. or concomitantly is well tolerated. The best strategy for post- 5 Sekeres MA, List AF, Cuthbertson D, Paquette R, Ganetzky R, Latham D et al. Phase I induction treatment with these agents should be explored further. combination trial of lenalidomide and azacitidine in patients with higher-risk Reducing to low-dose lenalidomide after the induction phase may myelodysplastic syndromes. J Clin Oncol 2010; 28: 2253–2258. reduce the potential for myelosuppression. The continuing phase 6 Pollyea DA, Kohrt HE, Gallegos L, Figueroa ME, Abdel-Wahab O, Zhang B et al. 2 expansion cohorts of both studies should provide additional Safety, efficacy and biological predictors of response to sequential azacitidine and lenalidomide for elderly patients with acute myeloid leukemia. Leukemia 2012; 26: information on this issue. 893–901. 7 Creutzig U, Kaspers GJ. Revised recommendations of the International Working Group for diagnosis, standardization of response criteria, treatment outcomes, and CONFLICT OF INTEREST reporting standards for therapeutic trials in acute myeloid leukemia. J Clin Oncol The authors declare no conflict of interest. 2004; 22: 3432–3433.

The proportion of CD34 þ CD38low or neg myeloblasts, but not side population frequency, predicts initial response to induction therapy in patients with newly diagnosed acute myeloid leukemia

Leukemia (2013) 27, 728–731; doi:10.1038/leu.2012.217 The relationship between SP and CD34 þ CD38dim cells has not been previously explored in AML. Three studies demonstrated that SP could be isolated from AML patients. The first study of 16 Normal hematopoiesis is sustained by a relatively small pool of patients showed that presence of the SP was not related to self-renewing primitive hematopoietic stem cells (HSCs). In murine expression of the multidrug resistance gene (MDR-1), expressed hematopoietic repopulation studies, human cells with bright variable CD38, was present with different types of cytogenetics expression of CD34, but no expression of CD38, are highly abnormalities, and variably engrafted immunodeficient mice.11 enriched in HSC activity.1 Cells with similar immunophenotype are The second study demonstrated that the SP was present in 80% of implicated as leukemia stem cells (LSCs),2 the initiating and 61 patients, that 3 out of 28 cases could engraft NODscid mice and repopulating reservoir for acute myeloid leukemia (AML). that 11/11 exhibited abnormal cytogenetics.12 The third study CD34 þ CD38neg cell frequency at diagnosis correlates with the highlighted the heterogeneity of the SP derived from 48 patients, incidence of minimal residual disease following achievement of although there was uniform expression of the stem cell marker complete remission (CR) in adult AML3 and with worse event-free CLL-1.13 survival in pediatric AML.4 A similar, but not identical, population LSCs appear to provide a reservoir for relapse of AML following of cells known as ‘side population’ (SP) has been defined based on chemotherapy.14 Chemoresistance may reflect the relative ability to efflux fluorescent dye Hoechst 33342 via the ABCG2 quiescent state of the LSC and/or increased expression of drug multidrug resistance-mediating transporter that can be inhibited efflux mediating transporters that extrude chemotherapy drugs.15 by fungal toxin Fumitremorgin C (FumC).5 The SP was originally The latter possibility motivated us to determine whether SP characterized in mice6 and was demonstrated to contain correlated with outcome in AML. In particular, we investigated the primitive HSCs. The stem cell reservoir in nearly all normal relationship between the frequencies of CD34 þ CD38neg, tissues resides in the SP,7 and it also contains the primitive CD34 þ CD38low and SP blasts and clinical prognostic variables, stem cells for solid tumors, including breast, prostate and lung such as age, cytogenetics, achievement and duration of first CR, in cancer.8–10 patients with newly diagnosed AML.

Accepted article preview online 31 July 2012; advacne online publication, 28 August 2012

Leukemia (2013) 718 – 757 & 2013 Macmillan Publishers Limited