Leukemia stemness and co-occurring mutations drive resistance to IDH inhibitors in acute myeloid leukemia.

Allosteric inhibitors of mutant IDH1 or IDH2 induce terminal differentiation of the mutant leukemic blasts and provide durable clinical responses in approximately 40% of acute myeloid leukemia (AML) patients with the mutations. However, primary resistance and acquired resistance to the drugs are major clinical issues. To understand the molecular underpinnings of clinical resistance to IDH inhibitors (IDHi), we perform multipronged genomic analyses (DNA sequencing, RNA sequencing and cytosine methylation profiling) in longitudinally collected specimens from 60 IDH1- or IDH2-mutant AML patients treated with the inhibitors. The analysis reveals that leukemia stemness is a major driver of primary resistance to IDHi, whereas selection of mutations in RUNX1/CEBPA or RAS-RTK pathway genes is the main driver of acquired resistance to IDHi, along with BCOR, homologous IDH gene, and TET2. These data suggest that targeting stemness and certain high-risk co-occurring mutations may overcome resistance to IDHi in AML.

Combination of azacitidine and enasidenib enhances leukemic cell differentiation and cooperatively hypomethylates DNA.

Azacitidine and enasidenib are two therapies available for treatment of acute myelogenous leukemia (AML), and the mechanisms of action of these drugs involve alteration of aberrant DNA methylation. We hypothesized that a combination of these agents could have interactive effects on DNA methylation and enhance differentiation in mIDH2 cells. Combination treatment enhanced cellular differentiation in TF-1 cells overexpressing IDJ2R140Q through increased hemoglobinization and increased hemoglobin γ RNA expression compared with the effects of single agents. Furthermore, in primary AML samples (IDH2R140Q or R172K), combination treatment reduced CD34+ cells and increased CD15+ cells to a greater extent than attained with single agents. To explore the mechanism of enhanced differentiation with combination treatment, the TF-1 epigenome was analyzed by profiling 5-hydroxymethylcytosine (5hmC) and 5-methylcytosine (5mC) DNA methylation changes. Enasidenib treatment alone increased 5hmC, consistent with reactivation of ten-eleven-translocation (TET) enzyme activity. Compared with treatment with azacitidine alone, combination treatment reduced 5mC levels at greater numbers of sites and these loci were significantly enriched in regions with increased 5hMC (25.8% vs. 7.4%). Results are consistent with a model in which enasidenib-mediated reactivation of ten-eleven-translocation enzymes cooperates with azacitidine-mediated inhibition of DNA methyltransferase enzymes, leading to greater reductions in DNA methylation and enhanced erythroid differentiation.

TP53 mutations in myelodysplastic syndromes and secondary AML confer an immunosuppressive phenotype.

Somatic gene mutations are key determinants of outcome in patients with myelodysplastic syndromes (MDS) and secondary AML (sAML). In particular, patients with TP53 mutations represent a distinct molecular cohort with uniformly poor prognosis. The precise pathogenetic mechanisms underlying these inferior outcomes have not been delineated. In this study, we characterized the immunological features of the malignant clone and alterations in the immune microenvironment in patients with TP53-mutant and wild-type MDS or sAML. Notably, PDL1 expression is significantly increased in hematopoietic stem cells of patients with TP53 mutations, which is associated with MYC upregulation and marked downregulation of MYC's negative regulator miR-34a, a p53 transcription target. Notably, patients with TP53 mutations display significantly reduced numbers of bone marrow-infiltrating OX40+ cytotoxic T cells and helper T cells, as well as decreased ICOS+ and 4-1BB+ natural killer cells. Further, highly immunosuppressive regulatory T cells (Tregs) (ie, ICOShigh/PD-1-) and myeloid-derived suppressor cells (PD-1low) are expanded in cases with TP53 mutations. Finally, a higher proportion of bone marrow-infiltrating ICOShigh/PD-1- Treg cells is a highly significant independent predictor of overall survival. We conclude that the microenvironment of TP53 mutant MDS and sAML has an immune-privileged, evasive phenotype that may be a primary driver of poor outcomes and submit that immunomodulatory therapeutic strategies may offer a benefit for this molecularly defined subpopulation.

Enasidenib in patients with mutant IDH2 myelodysplastic syndromes: a phase 1 subgroup analysis of the multicentre, AG221-C-001 trial.

BACKGROUND: Mutations in isocitrate dehydrogenase-2 (IDH2) occur in around 5% of patients with myelodysplastic syndromes. Neomorphic activity of mutant IDH2 proteins results in hypermethylation of DNA and histones, leading to blocked haemopoietic differentiation. Enasidenib, an inhibitor of mutated IDH2 proteins, induces responses in patients with IDH2-mutated, relapsed or refractory acute myeloid leukaemia. We aimed to establish the clinical outcomes of enasidenib monotherapy in a subgroup of patients with myelodysplastic syndromes harbouring mutations in IDH2 from the AG221-C-001 trial. METHODS: The multicentre, open-label, phase 1-2 AG221-C-001 trial enrolled patients with advanced haematological malignancies (2008 WHO criteria) harbouring an IDH2 mutation. The present study is a subgroup analysis of patients with IDH2-mutated myelodysplastic syndromes in the phase 1 dose-escalation and expansion portions of the trial. Patients with myelodysplastic syndromes were aged 18 years or older with an ECOG performance status score of 2 or lower, and were relapsed or refractory to, or ineligible for, standard treatments. Patients received oral doses of enasidenib at 60-300 mg per day in repeated 28-day treatment cycles. In this subgroup analysis, we focused on the safety and activity of enasidenib as main outcomes. Overall response rate, duration of response, and overall and event-free survival analyses were by intention-to-treat. Safety was assessed in all participants who received at least one dose of study drug in terms of treatment-emergent adverse events. The AG221-C-001 trial is registered on ClinicalTrials.gov, NCT01915498, status ongoing but closed to recruitment. FINDINGS: 17 patients with myelodysplastic syndromes harbouring an IDH2 mutation (median age, 67·0 years [IQR 60·5-73·0]) were enrolled between Feb 18, 2014, and Sept 1, 2015. At data cutoff (Oct 1, 2018), after a median follow-up of 11·0 months (IQR 6·8-23·0), all patients had discontinued enasidenib, with a median of 3 treatment cycles (2-15) for all patients (five [29%] received ≥12 cycles). At entry, three (18%) patients had relapsed after allogeneic stem-cell transplants, 13 (76%) had previously received therapy with hypomethylating agents, and ten (59%) had received at least two previous therapies. No dose-limiting toxicities were reported. The most common treatment-emergent adverse events were diarrhoea and nausea (in nine [53%] patients each). Most common grade 3-4 treatment-emergent adverse events were indirect hyperbilirubinaemia (in six [35%] patients), pneumonia (in five [29%] patients), and thrombocytopaenia (in four [24%] patients). Serious treatment-emergent adverse events in more than one patient were pneumonia (in five [29% patients); tumor lysis syndrome (in three [18%] patients); and sepsis, atrial flutter, indirect hyperbilirubinaemia, cerebral hemorrhage, and mental status change (in two [12%] patients each). No treatment-related deaths occurred. An overall response was achieved in 9 patients (53% [95% CI 28-77]), with a median duration of response of 9·2 months (95% CI 1·0-not reached). Six (46%) of 13 patients previously treated with hypomethylating agents responded. Median overall survival was 16·9 months (95% CI 1·5-32·3), and median event-free survival was 11·0 months (1·5-16·7). INTERPRETATION: Enasidenib is generally well tolerated and can induce responses in patients with mutant IDH2 myelodysplastic syndromes, including in those who have had previous therapy with hypomethylating agents. Testing for IDH2 mutations in myelodysplastic syndromes is essential for identifying patients who might benefit from enasidenib therapy, including those patients in whom conventional treatments have been unsuccessful. FUNDING: Celgene and Agios Pharmaceuticals.

Enasidenib, an inhibitor of mutant IDH2 proteins, induces durable remissions in older patients with newly diagnosed acute myeloid leukemia.

Older adults with acute myeloid leukemia (AML) who are not fit for standard chemotherapy historically have poor outcomes. Approximately 12-15% of older patients with AML harbor isocitrate dehydrogenase 2 (IDH2) gene mutations. Enasidenib is an oral inhibitor of mutant IDH2 proteins. Among 39 patients with newly diagnosed mutant-IDH2 AML who received enasidenib monotherapy in this phase I/II trial, median age was 77 years (range 58-87) and 23 patients (59%) had had an antecedent hematologic disorder. The median number of enasidenib treatment cycles was 6.0 (range 1-35). The most common treatment-related adverse events were indirect hyperbilirubinemia (31%), nausea (23%), and fatigue, decreased appetite, and rash (18% each). Treatment-related grade 3-4 cytopenias were reported for eight patients (21%); there was no treatment-related grade 3-4 infections. Twelve patients achieved a response (overall response rate 30.8% [95% CI 17.0%, 47.6%]), including seven patients (18%) who attained complete remission. At a median follow-up of 8.4 months, the median duration of any response was not reached (NR). Median overall survival for all patients was 11.3 months (95% CI 5.7, 15.1), and was NR for responders. Oral, outpatient targeted treatment with enasidenib may benefit older adults with newly diagnosed mutant-IDH2 AML who are not candidates for cytotoxic regimens.

Randomised phase 2 study of pembrolizumab plus CC-486 versus pembrolizumab plus placebo in patients with previously treated advanced non-small cell lung cancer.

INTRODUCTION: Preclinical and early clinical studies suggest that combining epigenetic agents with checkpoint inhibitors can potentially improve outcomes in patients with previously treated advanced non-small cell lung cancer (NSCLC). This phase 2 trial examined second-line pembrolizumab + CC-486 (oral azacitidine) in patients with advanced NSCLC. METHODS: Patients with one prior line of platinum-containing therapy were randomised in a ratio of 1:1 to CC-486 or placebo, on days 1-14, in combination with pembrolizumab on day 1 of a 21-day cycle. The primary end-point was progression-free survival (PFS). Key secondary end-points included overall survival (OS), overall response rate (ORR) and safety. RESULTS: Among 100 patients randomised (pembrolizumab + CC-486: 51; pembrolizumab + placebo: 49), most were male (57.0%), were white (87.0%) and had Eastern Cooperative Oncology Group performance status 1 (68.0%). No significant difference in PFS was observed between the pembrolizumab + CC-486 and pembrolizumab + placebo arms (median, 2.9 and 4.0 months, respectively; hazard ratio [HR], 1.374; 90% confidence interval [CI], 0.926-2.038; P = 0.1789). Median OS was 11.9 months versus not estimable (HR, 1.375; 90% CI, 0.830-2.276; P = 0.2968); ORR was 20% versus 14%. Median treatment duration was shorter (15.0 versus 24.1 weeks), and the number of cycles was lower (5.0 versus 7.0) with pembrolizumab + CC-486 versus pembrolizumab + placebo. No new safety signals for CC-486 or pembrolizumab were detected. Treatment-emergent adverse events were more common in the pembrolizumab + CC-486 arm, particularly gastrointestinal, potentially impacting treatment feasibility. CONCLUSIONS: No improvement in PFS was observed with pembrolizumab + CC-486 versus pembrolizumab + placebo. Decreased treatment exposure due to adverse events may have impacted efficacy with pembrolizumab + CC-486.

Molecular remission and response patterns in patients with mutant-IDH2 acute myeloid leukemia treated with enasidenib.

Approximately 8% to 19% of patients with acute myeloid leukemia (AML) have isocitrate dehydrogenase-2 (IDH2) mutations, which occur at active site arginine residues R140 and R172. IDH2 mutations produce an oncometabolite, 2-hydroxyglutarate (2-HG), which leads to DNA and histone hypermethylation and impaired hematopoietic differentiation. Enasidenib is an oral inhibitor of mutant-IDH2 proteins. This first-in-human phase 1/2 study evaluated enasidenib doses of 50 to 650 mg/d, administered in continuous 28-day cycles, in patients with mutant-IDH2 hematologic malignancies. Overall, 214 of 345 patients (62%) with relapsed or refractory (R/R) AML received enasidenib, 100 mg/d. Median age was 68 years. Forty-two patients (19.6%) attained complete remission (CR), 19 patients (10.3%) proceeded to an allogeneic bone marrow transplant, and the overall response rate was 38.8% (95% confidence interval [CI], 32.2-45.7). Median overall survival was 8.8 months (95% CI, 7.7-9.6). Response and survival were comparable among patients with IDH2-R140 or IDH2-R172 mutations. Response rates were similar among patients who, at study entry, were in relapse (37.7%) or were refractory to intensive (37.5%) or nonintensive (43.2%) therapies. Sixty-six (43.1%) red blood cell transfusion-dependent and 53 (40.2%) platelet transfusion-dependent patients achieved transfusion independence. The magnitude of 2-HG reduction on study was associated with CR in IDH2-R172 patients. Clearance of mutant-IDH2 clones was also associated with achievement of CR. Among all 345 patients, the most common grade 3 or 4 treatment-related adverse events were hyperbilirubinemia (10%), thrombocytopenia (7%), and IDH differentiation syndrome (6%). Enasidenib was well tolerated and induced molecular remissions and hematologic responses in patients with AML for whom prior treatments had failed. The study is registered at www.clinicaltrials.gov as #NCT01915498.

Multiomics of azacitidine-treated AML cells reveals variable and convergent targets that remodel the cell-surface proteome.

Myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) are diseases of abnormal hematopoietic differentiation with aberrant epigenetic alterations. Azacitidine (AZA) is a DNA methyltransferase inhibitor widely used to treat MDS and AML, yet the impact of AZA on the cell-surface proteome has not been defined. To identify potential therapeutic targets for use in combination with AZA in AML patients, we investigated the effects of AZA treatment on four AML cell lines representing different stages of differentiation. The effect of AZA treatment on these cell lines was characterized at three levels: the DNA methylome, the transcriptome, and the cell-surface proteome. Untreated AML cell lines showed substantial overlap at all three omics levels; however, while AZA treatment globally reduced DNA methylation in all cell lines, changes in the transcriptome and surface proteome were subtle and differed among the cell lines. Transcriptome analysis identified five commonly up-regulated coding genes upon AZA treatment in all four cell lines, TRPM4 being the only gene encoding a surface protein, and surface proteome analysis found no commonly regulated proteins. Gene set enrichment analysis of differentially regulated RNA and surface proteins showed a decrease in metabolic pathways and an increase in immune defense response pathways. As such, AZA treatment led to diverse effects at the individual gene and protein levels but converged to common responses at the pathway level. Given the heterogeneous responses in the four cell lines, we discuss potential therapeutic strategies for AML in combination with AZA.

Isoform Switching as a Mechanism of Acquired Resistance to Mutant Isocitrate Dehydrogenase Inhibition.

Somatic mutations in cytosolic or mitochondrial isoforms of isocitrate dehydrogenase (IDH1 or IDH2, respectively) contribute to oncogenesis via production of the metabolite 2-hydroxyglutarate (2HG). Isoform-selective IDH inhibitors suppress 2HG production and induce clinical responses in patients with IDH1- and IDH2-mutant malignancies. Despite the promising activity of IDH inhibitors, the mechanisms that mediate resistance to IDH inhibition are poorly understood. Here, we describe four clinical cases that identify mutant IDH isoform switching, either from mutant IDH1 to mutant IDH2 or vice versa, as a mechanism of acquired clinical resistance to IDH inhibition in solid and liquid tumors. SIGNIFICANCE: IDH-mutant cancers can develop resistance to isoform-selective IDH inhibition by "isoform switching" from mutant IDH1 to mutant IDH2 or vice versa, thereby restoring 2HG production by the tumor. These findings underscore a role for continued 2HG production in tumor progression and suggest therapeutic strategies to prevent or overcome resistance.This article is highlighted in the In This Issue feature, p. 1494.

Cytogenetics and gene mutations influence survival in older patients with acute myeloid leukemia treated with azacitidine or conventional care.

Older patients with newly diagnosed acute myeloid leukemia (AML) in the phase 3 AZA-AML-001 study were evaluated at entry for cytogenetic abnormalities, and a subgroup of patients was assessed for gene mutations. Patients received azacitidine 75 mg/m2/day x7 days (n = 240) or conventional care regimens (CCR; n = 245): intensive chemotherapy, low-dose cytarabine, or best supportive care only. Overall survival (OS) was assessed for patients with common (occurring in ≥10% of patients) cytogenetic abnormalities and karyotypes, and for patients with recurring gene mutations. There was a significant OS improvement with azacitidine vs CCR for patients with European LeukemiaNet-defined Adverse karyotype (HR 0.71 [95%CI 0.51-0.99]; P = 0.046). Azacitidine-treated patients with -5/5q-, -7/7q-, or 17p abnormalities, or with monosomal or complex karyotypes, had a 31-46% reduced risk of death vs CCR. The most frequent gene mutations were DNMT3A (27%), TET2 (25%), IDH2 (23% [R140, 15%; R172, 8%]), and TP53 (21%). Compared with wild-type, OS was significantly reduced among CCR-treated patients with TP53 or NRAS mutations and azacitidine-treated patients with FLT3 or TET2 mutations. Azacitidine may be a preferred treatment for older patients with AML with Adverse-risk cytogenetics, particularly those with chromosome 5, 7, and/or 17 abnormalities and complex or monosomal karyotypes. The influence of gene mutations in azacitidine-treated patients warrants further study.

Clonal heterogeneity of acute myeloid leukemia treated with the IDH2 inhibitor enasidenib.

Mutations in the gene encoding isocitrate dehydrogenase 2 (IDH2) occur in several types of cancer, including acute myeloid leukemia (AML). In model systems, mutant IDH2 causes hematopoietic differentiation arrest. Enasidenib, a selective small-molecule inhibitor of mutant IDH2, produces a clinical response in 40% of treated patients with relapsed/refractory AML by promoting leukemic cell differentiation. Here, we studied the clonal basis of response and acquired resistance to enasidenib treatment. Using sequential patient samples, we determined the clonal structure of hematopoietic cell populations at different stages of differentiation. Before therapy, IDH2-mutant clones showed variable differentiation arrest. Enasidenib treatment promoted hematopoietic differentiation from either terminal or ancestral mutant clones; less frequently, treatment promoted differentiation of nonmutant cells. Analysis of paired diagnosis/relapse samples did not identify second-site mutations in IDH2 at relapse. Instead, relapse arose by clonal evolution or selection of terminal or ancestral clones, thus highlighting multiple bypass pathways that could potentially be targeted to restore differentiation arrest. These results show how mapping of clonal structure in cell populations at different stages of differentiation can reveal the response and evolution of clones during treatment response and relapse.

Randomized Phase III Study of Lenalidomide Versus Placebo in RBC Transfusion-Dependent Patients With Lower-Risk Non-del(5q) Myelodysplastic Syndromes and Ineligible for or Refractory to Erythropoiesis-Stimulating Agents.

PURPOSE: This international phase III, randomized, placebo-controlled, double-blind study assessed the efficacy and safety of lenalidomide in RBC transfusion-dependent patients with International Prognostic Scoring System lower-risk non-del(5q) myelodysplastic syndromes ineligible for or refractory to erythropoiesis-stimulating agents. PATIENTS AND METHODS: In total, 239 patients were randomly assigned (2:1) to treatment with lenalidomide (n = 160) or placebo (n = 79) once per day (on 28-day cycles). The primary end point was the rate of RBC transfusion independence (TI) ≥ 8 weeks. Secondary end points were RBC-TI ≥ 24 weeks, duration of RBC-TI, erythroid response, health-related quality of life (HRQoL), and safety. RESULTS: RBC-TI ≥ 8 weeks was achieved in 26.9% and 2.5% of patients in the lenalidomide and placebo groups, respectively (P < .001). Ninety percent of patients achieving RBC-TI responded within 16 weeks of treatment. Median duration of RBC-TI with lenalidomide was 30.9 weeks (95% CI, 20.7 to 59.1). Transfusion reduction of ≥ 4 units packed RBCs, on the basis of a 112-day assessment, was 21.8% in the lenalidomide group and 0% in the placebo group. Higher response rates were observed in patients with lower baseline endogenous erythropoietin ≤ 500 mU/mL (34.0% v 15.5% for > 500 mU/mL). At week 12, mean changes in HRQoL scores from baseline did not differ significantly between treatment groups, which suggests that lenalidomide did not adversely affect HRQoL. Achievement of RBC-TI ≥ 8 weeks was associated with significant improvements in HRQoL (P < .01). The most common treatment-emergent adverse events were neutropenia and thrombocytopenia. CONCLUSION: Lenalidomide yields sustained RBC-TI in 26.9% of RBC transfusion-dependent patients with lower-risk non-del(5q) myelodysplastic syndromes ineligible for or refractory to erythropoiesis-stimulating agents. Response to lenalidomide was associated with improved HRQoL. Treatment-emergent adverse event data were consistent with the known safety profile of lenalidomide.

A calcium- and calpain-dependent pathway determines the response to lenalidomide in myelodysplastic syndromes.

Despite the high response rates of individuals with myelodysplastic syndrome (MDS) with deletion of chromosome 5q (del(5q)) to treatment with lenalidomide (LEN) and the recent identification of cereblon (CRBN) as the molecular target of LEN, the cellular mechanism by which LEN eliminates MDS clones remains elusive. Here we performed an RNA interference screen to delineate gene regulatory networks that mediate LEN responsiveness in an MDS cell line, MDSL. We identified GPR68, which encodes a G-protein-coupled receptor that has been implicated in calcium metabolism, as the top candidate gene for modulating sensitivity to LEN. LEN induced GPR68 expression via IKAROS family zinc finger 1 (IKZF1), resulting in increased cytosolic calcium levels and activation of a calcium-dependent calpain, CAPN1, which were requisite steps for induction of apoptosis in MDS cells and in acute myeloid leukemia (AML) cells. In contrast, deletion of GPR68 or inhibition of calcium and calpain activation suppressed LEN-induced cytotoxicity. Moreover, expression of calpastatin (CAST), an endogenous CAPN1 inhibitor that is encoded by a gene (CAST) deleted in del(5q) MDS, correlated with LEN responsiveness in patients with del(5q) MDS. Depletion of CAST restored responsiveness of LEN-resistant non-del(5q) MDS cells and AML cells, providing an explanation for the superior responses of patients with del(5q) MDS to LEN treatment. Our study describes a cellular mechanism by which LEN, acting through CRBN and IKZF1, has cytotoxic effects in MDS and AML that depend on a calcium- and calpain-dependent pathway.

Pharmacokinetics and Pharmacodynamics with Extended Dosing of CC-486 in Patients with Hematologic Malignancies.

UNLABELLED: CC-486 (oral azacitidine) is an epigenetic modifier in development for patients with myelodysplastic syndromes and acute myeloid leukemia. In part 1 of this two-part study, a 7-day CC-486 dosing schedule showed clinical activity, was generally well tolerated, and reduced DNA methylation. Extending dosing of CC-486 beyond 7 days would increase duration of azacitidine exposure. We hypothesized that extended dosing would therefore provide more sustained epigenetic activity. Reported here are the pharmacokinetic (PK) and pharmacodynamic (PD) profiles of CC-486 extended dosing schedules in patients with myelodysplastic syndromes (MDS), chronic myelomonocytic leukemia (CMML) or acute myeloid leukemia (AML) from part 2 of this study. PK and/or PD data were available for 59 patients who were sequentially assigned to 1 of 4 extended CC-486 dosing schedules: 300mg once-daily or 200mg twice-daily for 14 or 21 days per 28-day cycle. Both 300mg once-daily schedules and the 200mg twice-daily 21-day schedule significantly (all P < .05) reduced global DNA methylation in whole blood at all measured time points (days 15, 22, and 28 of the treatment cycle), with sustained hypomethylation at cycle end compared with baseline. CC-486 exposures and reduced DNA methylation were significantly correlated. Patients who had a hematologic response had significantly greater methylation reductions than non-responding patients. These data demonstrate that extended dosing of CC-486 sustains epigenetic effects through the treatment cycle. TRIAL REGISTRATION: ClinicalTrials.gov NCT00528983.

Lenalidomide induces ubiquitination and degradation of CK1α in del(5q) MDS.

Lenalidomide is a highly effective treatment for myelodysplastic syndrome (MDS) with deletion of chromosome 5q (del(5q)). Here, we demonstrate that lenalidomide induces the ubiquitination of casein kinase 1A1 (CK1α) by the E3 ubiquitin ligase CUL4-RBX1-DDB1-CRBN (known as CRL4(CRBN)), resulting in CK1α degradation. CK1α is encoded by a gene within the common deleted region for del(5q) MDS and haploinsufficient expression sensitizes cells to lenalidomide therapy, providing a mechanistic basis for the therapeutic window of lenalidomide in del(5q) MDS. We found that mouse cells are resistant to lenalidomide but that changing a single amino acid in mouse Crbn to the corresponding human residue enables lenalidomide-dependent degradation of CK1α. We further demonstrate that minor side chain modifications in thalidomide and a novel analogue, CC-122, can modulate the spectrum of substrates targeted by CRL4(CRBN). These findings have implications for the clinical activity of lenalidomide and related compounds, and demonstrate the therapeutic potential of novel modulators of E3 ubiquitin ligases.

Discovery of bicyclic pyrazoles as class III histone deacetylase SIRT1 and SIRT2 inhibitors.

A series of bicyclic pyrazole carboxamides was synthesized and tested for inhibitory activity against the class III deacetylase sirtuin enzymes. Moderate to low micromolar inhibitory activities were obtained against SIRT1 and SIRT2. These bicyclic pyrazole compounds represent a new class of sirtuin inhibitors with a preference for SIRT1 over SIRT2.

p53 protein expression independently predicts outcome in patients with lower-risk myelodysplastic syndromes with del(5q).

Del(5q) myelodysplastic syndromes defined by the International Prognostic Scoring System as low- or intermediate-1-risk (lower-risk) are considered to have an indolent course; however, recent data have identified a subgroup of these patients with more aggressive disease and poorer outcomes. Using deep sequencing technology, we previously demonstrated that 18% of patients with lower-risk del(5q) myelodysplastic syndromes carry TP53 mutated subclones rendering them at higher risk of progression. In this study, bone marrow biopsies from 85 patients treated with lenalidomide in the MDS-004 clinical trial were retrospectively assessed for p53 expression by immunohistochemistry in association with outcome. Strong p53 expression in ≥ 1% of bone marrow progenitor cells, observed in 35% (30 of 85) of patients, was significantly associated with higher acute myeloid leukemia risk (P=0.0006), shorter overall survival (P=0.0175), and a lower cytogenetic response rate (P=0.009), but not with achievement or duration of 26-week transfusion independence response. In a multivariate analysis, p53-positive immunohistochemistry was the strongest independent predictor of transformation to acute myeloid leukemia (P=0.0035). Pyrosequencing analysis of laser-microdissected cells with strong p53 expression confirmed the TP53 mutation, whereas cells with moderate expression predominantly had wild-type p53. This study validates p53 immunohistochemistry as a strong and clinically useful predictive tool in patients with lower-risk del(5q) myelodysplastic syndromes. This study was based on data from the MDS 004 trial (clinicaltrials.gov identifier: NCT00179621).

Lenalidomide as a disease-modifying agent in patients with del(5q) myelodysplastic syndromes: linking mechanism of action to clinical outcomes.

Deletion of the long arm of chromosome 5, del(5q), is the most prevalent cytogenetic abnormality in patients with myelodysplastic syndromes (MDS). In isolation, it is traditionally associated with favorable prognosis compared with other subtypes of MDS. However, owing to the inherent heterogeneity of the disease, prognosis for patients with del(5q) MDS is highly variable depending on the presence of factors such as additional chromosomal abnormalities, >5 % blasts in the bone marrow (BM), or transfusion dependence. Over recent years, the immunomodulatory drug lenalidomide has demonstrated remarkable efficacy in patients with del(5q) MDS. Advances in the understanding of the pathogenesis of the disease have suggested that lenalidomide targets aberrant signaling pathways caused by haplosufficiency of specific genes in a commonly deleted region on chromosome 5 (e.g., SPARC, RPS14, Cdc25C, and PP2A). As a result, the agent specifically targets del(5q) clones while also promoting erythropoiesis and repopulation of the bone marrow in normal cells. This review discusses recent developments in the understanding of the mechanism of action of lenalidomide, and how this underlies favorable outcomes in patients with del(5q) MDS. In addition, we discuss how improved understanding of the mechanism of disease will facilitate clinicians' ability to predict/monitor response and identify patients at risk of relapse.