Lenalidomide and azacitidine are active in MDS patients, and may complement each other by targeting the bone marrow microenvironment and the malignant clone. A recent Phase I trial testing the lenalidomide and azacitidine combination yielded encouraging results; however, lenalidomide's contribution was unclear. In this study, 18 higher-risk MDS patients were treated with the combination for seven cycles, after which lenalidomide was discontinued in eight patients who achieved a complete response, with azacitidine monotherapy continuing until disease progression. We report on three patients who relapsed on monotherapy with excess blasts at 12, 19, and 24 months, in whom lenalidomide was then resumed in combination with azacitidine. Each patient, one with normal cytogenetics at relapse; one with a +8 abnormality; and one with del(4q25), recaptured a complete response that was sustained for 5, 7, and 7+ months. We conclude that the addition of lenalidomide to azacitidine provides additional clinical benefit over azacitidine monotherapy.
The myelodysplastic syndromes (MDS) comprise a spectrum of bone marrow disorders associated with cytopenias, cytogenetic abnormalities, and in higher-risk subtypes (patients with excess myeloblasts or an International Prognostic Scoring System (IPSS) risk score >1.0), a high likelihood of transformation to acute myeloid leukemia (AML) [1–5]. Three drugs, azacitidine, decitabine and lenalidomide, are approved for the treatment of MDS [6–8]. Lenalidomide selectively suppresses deletion 5q MDS clones through inhibition of haplodeficient cell cycle regulatory targets coded within the common deleted region (CDR) [9, 10] complemented by effects on the bone marrow microenvironment. Azacitidine and decitabine exert their effects via DNA methyltrasferase inhibition and also direct cytotoxicity,  and azacitidine monotherapy improves overall survival in higher-risk MDS patients compared to conventional care .
We recently reported results from a Phase I study of combination lenalidomide and azacitidine in 18 higher-risk MDS patients, in which a Phase II dose was established: azacitidine administered at 75 mg/m2 daily for five consecutive days, and lenalidomide 10 mg daily for 21 days, of a 28-day cycle . The overall response rate was 67%, including a complete response (CR) rate of 44%. Patients were treated with combination therapy for seven cycles, after which patients achieving a CR were continued on azacitidine monotherapy, administered for five to seven consecutive days at 75 mg/m2 daily, repeated every four to six weeks. As this was a single-arm study, it was unknown whether combination therapy provided additional benefit over single-agent azacitidine.
We report herein three patients with normal cytogenetics who achieved a CR with a lenalidomide and azacitidine combination regimen; continued on azacitidine monotherapy until disease relapse, with two patients demonstrating distinct cytogenetic abnormalities; and recaptured their CR status with the reinitiation of lenalidomide combined with azacitidine (see Fig. 1).
This multicenter, single-arm, open-label, Phase I study of combination therapy with lenalidomide and azacitidine received local Institutional Review Board approval from all participating sites and from the Data Safety and Monitoring Board of the Rare Diseases Branch of the National Institutes of Health, and was registered with http//:clinicaltrials.gov (NCT00352001). The study opened in May, 2005 and the last subject was enrolled in May, 2008. Responses were defined according to 2006 International Working Group criteria .
This 70-year-old man with newly diagnosed RAEB-2 (15% blasts), normal cytogenetics, no somatic lesions by single nucleotide polymorphism array (SNP-A) karyotyping, and an IPSS score of 1.5, started combination therapy with lenalidomide (5 mg daily for 21 days) and azacitidine (75 mg/m2 daily for 5 days) in May, 2007 (Table I). He achieved a CR in September, 2007, and completed the study in December, 2007. Maintenance therapy was initiated with azacitidine monotherapy, 75 mg/m2 daily for six days, repeated every four to six weeks. Nineteen months later (April, 2009) he relapsed, with decreased peripheral blood counts and 7% blasts, with normal cytogenetics. Lenalidomide was resumed at a daily dose of 10 mg for 21 days combined with azacitidine. His counts normalized, and a bone marrow biopsy in August, 2009, confirmed CR recovery, with 1% blasts and normal peripheral counts. A microdeletion of 4q25, spanning nucleotides 109226732-109731065, was detected using SNP-A. He remained in a CR until February, 2010, when he progressed to AML with persistence of the 4q25 SNP-A-detected microdeletion. He is undergoing cytotoxic induction therapy.
Table I. Characteristics of Subjects
|Age at study entry (years)||66||72||62|
|Baseline: Hgb (g/dl)||8.1||12||9.2|
| Plt (×103/ml)||243||94||37|
| ANC (×103/ml)||4.28||0.88||0.32|
| Blast %||15||7||16|
|Initial combination regimen:||AZA 75mg/m2 days 1-5; LEN 5mg days 1-21||AZA 50mg/m2 days 1-5, 8-12; LEN 5mg days 1-14||AZA 75mg/m2 days 1-5; LEN 10mg days 1-21|
|Time to initial CR (months)||4||4||4|
|1st CR duration (months)||19||12||24|
|Cytogenetic lesions at 1st CR||ND||ND||ND|
|Cytogenetic lesions at relapse||Del(4q25)||+8||ND|
|2nd CR Duration (months)||5||7||7+ (NR)|
|Cytogenetic lesions at 2nd CR||Del(4q25)||ND||ND|
This 72-year-old man presented with newly-diagnosed RAEB-1 (7% blasts), normal cytogenetics, with no additional somatic lesions by SNP-A-based karyotyping, and an IPSS score of 1.0. He started combination therapy in September, 2007, with lenalidomide (5 mg daily for 14 days) and azacitidine (50 mg/m2 daily Days 1–5 and 8–12). He achieved CR in January, 2008, and completed the study in April, 2008, then continuing azacitidine monotherapy, 75 mg/m2 daily for six days, repeated every four to six weeks. He relapsed nine months later, in January, 2009, with decreased platelet and neutrophil counts and 8% bone marrow blasts, with a new +8 cytogenetic abnormality. He continued on azacitidine, with lenalidomide added at 10 mg daily for 21 days. His peripheral blood counts normalized, and a second CR was confirmed by bone marrow biopsy in May, 2009, with 3% blasts and normal cytogenetics. SNP confirmed no somatic lesions. He progressed to AML in January, 2010, and achieved a third CR following remission induction chemotherapy.
This 60-year-old man presented with newly diagnosed RAEB-2 (16% blasts), normal cytogenetics, and an IPSS score of 2.0. He began combination therapy with lenalidomide (10 mg daily for 21 days) and azacitidine (75 mg/m2 daily for 5 days) in September, 2007. A CR was documented in November, 2007, and he completed the combination study in March, 2008, continuing therapy with azacitidine monotherapy, at a dose of 75 mg/m2 daily for five to seven days, repeated every four to six weeks. Karyotyping using SNP-A identified no somatic lesions. Twenty months later, in November, 2009, he developed neutropenia and thrombocytopenia, and a bone marrow biopsy confirmed relapsed disease, with 6% blasts and normal cytogenetics, and no additional somatic lesions identified by SNP-A analysis. He continued on azacitidine at the same dose and schedule, with the addition of lenalidomide at 10 mg daily for 21 days. A bone marrow biopsy performed in April, 2010, demonstrated a reduction in blast percentage, to 2%, and no somatic lesions detected by SNP-A karyotyping. He continues on the combination therapy.
Our Phase I study was the first trial to combine two FDA-approved drugs for MDS, lenalidomide, and azacitidine. Here we report evidence supporting the additive benefit of lenalidomide in this combination strategy. Three patients served as their own internal controls, achieving a CR with the combination regimen; relapsing after prolonged azacitidine monotherapy maintenance; and then recapturing CR with lenalidomide readministration. In so doing, we believe we satisfied Koch's postulates for causality,  when applied to disease in general, as agents were introduced and evoked a response; one was withdrawn, with loss of that response; and was then reintroduced, with the same response resulting.
The combination design of the initial Phase I study was intended to complement the antiproliferative effects of azacitidine on the MDS clone with the modulatory effects of lenalidomide on the bone marrow microenvironment. These cases provide unambiguous evidence for the contribution of lenalidomide to the disease modifying effect of the combination regimen, and show that remissions were not maintained by treatment with azacitidine alone. All three patients had normal cytogenetics at diagnosis, confirmed at the molecular level by 250k SNP-A analysis, with specific molecular testing for lesions involving c-Cbl, b-Cbl, JAK2, and TET2 genes. Two patients acquired new genetic lesions during relapse; one involving trisomy 8 (Case 2), which disappeared upon achieving a second CR; the second involving a microdeletion 4q25 (Case 3) detected by SNP-A karyotyping, which persisted at the time of pathologic CR recapture. Second CRs were not durable, and two patients progressed to AML. It is possible that such cryptic genetic lesions involving the TET2 gene may be linked to lenalidomide responsiveness, as has been speculated recently as lenalidomide relates to histone methylation in Namalwa cell lines . The latter will be investigated in a comparative study of azacitidine vs. the combination to verify the benefit of this novel combination regimen.
In conclusion, the combination of azacitidine and lenalidomide produced clinical benefit compared with single agent azacitidine alone, when patients were used as their own controls.
M.A.S. designed the research, performed the research, analyzed the data, and wrote the manuscript. C.O. analyzed data and edited the manuscript. A.F.L. designed the research, performed the research, and edited the manuscript. K.P. analyzed the data and edited the manuscript. M.A. performed the research and edited the manuscript. R.E. performed the research and edited the manuscript. J.P.M. designed the research, performed the research, analyzed the data, and wrote the manuscript.
Mikkael A. Sekeres* , Christine O'Keefe, Alan F. List, Katarina Paulic*, Manuel Afable II*, Ricki Englehaupt*, Jaroslaw P. Maciejewski, * Department of Hematologic Oncology and Blood Disorders, Leukemia Program, Cleveland Clinic Taussig Cancer Institute, Cleveland, Ohio, Translational Hematology and Oncology Research, Cleveland Clinic Taussig Cancer Institute, Cleveland, Ohio, H. Lee Moffitt Cancer Center & Research Institute, Tampa, Florida.