Phase 2 study of low-dose clofarabine plus cytarabine for patients with higher-risk myelodysplastic syndrome who have relapsed or are refractory to hypomethylating agents.

BACKGROUND: The outcome of patients with higher-risk myelodysplastic syndromes (MDS) after hypomethylating agent (HMA) failure is poor. This study evaluated the safety and activity of a combination of low-dose clofarabine and cytarabine for these patients. METHODS: Seventy patients with higher-risk MDS who had no response, progressed, or relapsed after at least 4 cycles of HMA therapy were treated. RESULTS: The median age was 72 years. Thirty-nine percent of the patients had high-risk disease according to the International Prognostic Scoring System, and 50% of the patients had poor-risk cytogenetics. Twenty-three percent of the patients had therapy-related MDS. The median number of prior cycles of HMA was 6 (range, 4-45). The overall response rate was 44%. The 6-week mortality rate was 9%. Grade 3 and higher nonhematologic toxicities were rare, but infections occurred in 52% of the patients, and fever of unknown origin occurred in 33%. The median overall survival (OS) was 10 months (95% confidence interval, 1-37 months). Thirteen percent of the patients underwent allogeneic stem cell transplantation. The responding patients had a median OS of 22 months, whereas the nonresponding patients had a median OS of 4 months. A complex karyotype was associated with worse response rates and OS. CONCLUSIONS: The combination of low-dose clofarabine and cytarabine is clinically active in these patients with few treatment options. Cancer 2017;123:629-637. © 2016 American Cancer Society.

Unraveling Myelodysplastic Syndromes: Current Knowledge and Future Directions.

Myelodysplastic syndromes (MDS) affect more than 30,000 patients in the USA per year, most of whom are elderly, and these diseases are associated with dismal prognoses. The main features of MDS are ineffective hematopoiesis and aberrant myeloid differentiation. Furthermore, MDS are heterogeneous, both clinically and molecularly. This heterogeneity and the frequent occurrence of age-related comorbidities make the management of these diseases challenging. In fact, there have been no new drug approvals for MDS in the USA in the last 9 years, and few currently available investigational drugs are likely to be approved in the near future. Novel targeted treatment based on better understanding of the pathogenesis of MDS is needed to maximize patient outcomes. Here, we discuss new insights into diagnostic accuracy, prognostic assessment, pathogenic mechanisms, and effective treatments for MDS.

Combination of hyper-CVAD with ponatinib as first-line therapy for patients with Philadelphia chromosome-positive acute lymphoblastic leukaemia: a single-centre, phase 2 study.

BACKGROUND: Combination of chemotherapy with a tyrosine-kinase inhibitor is effective in the treatment of Philadelphia chromosome-positive acute lymphoblastic leukaemia. Ponatinib is a more potent BCR-ABL1 inhibitor than all other tyrosine-kinase inhibitors and selectively suppresses the resistant T315I clones. We examined the activity and safety of combining chemotherapy with ponatinib for patients with Philadelphia chromosome-positive acute lymphoblastic leukaemia in this continuing phase 2 trial. METHODS: In this single-centre, phase 2, single-arm trial, adult patients with previously untreated Philadelphia chromosome-positive acute lymphoblastic leukaemia were sequentially enrolled. Patients who had received fewer than two courses of previous chemotherapy with or without tyrosine-kinase inhibitors were also eligible. Patients had to be aged 18 years or older, have an Eastern Cooperative Oncology Group performance status of 2 or less, have normal cardiac function (defined by ejection fraction above 50%), and have adequate organ function (serum bilirubin ≤3·0 mg/dL and serum creatinine ≤3·0 mg/dL, unless higher concentrations were believed to be due to a tumour). Patients received eight cycles of hyperfractionated cyclophosphamide, vincristine, doxorubicin, and dexamethasone (hyper-CVAD) alternating with high-dose methotrexate and cytarabine every 21 days. Ponatinib 45 mg was given daily for the first 14 days of cycle 1 then continuously for the subsequent cycles. Patients in complete remission received maintenance with ponatinib 45 mg daily with vincristine and prednisone monthly for 2 years followed by ponatinib indefinitely. The primary endpoint for this study was event-free survival. The trial is registered at ClinicalTrials.gov, number NCT01424982. FINDINGS: 37 patients were enrolled and treated from Nov 1, 2011, to Sept 1, 2013. 2-year event-free survival rate was 81% (95% CI 64-90). Grade 3 or more toxic effects included infections during induction (20 [54%] patients), increased aspartate aminotransferase and alanine aminotransferase concentration (14 [38%] patients), thrombotic events (three [8%]), myocardial infarction (three [8%]), hypertension (six [16%]), skin rash (eight [22%]), and pancreatitis (six [16%] patients). Two patients died from from myocardial infarction potentially related to treatment; another patient also died from myocardial infarction related to sepsis. Two further patients died, one from bleeding and another from infection, both deemed unrelated to treatment. INTERPRETATION: The first results of this ongoing trial indicate that the combination of chemotherapy with ponatinib is effective in achieving early sustained remissions in patients with newly diagnosed Philadelphia chromosome-positive acute lymphoblastic leukaemia. New strategies, including dosing titration of ponatinib and optimised control of vascular risk factors, might further improve outcomes. FUNDING: ARIAD Pharmaceuticals Inc.

RUNX3 promoter hypermethylation is frequent in leukaemia cell lines and associated with acute myeloid leukaemia inv(16) subtype.

Correlative and functional studies support the involvement of the RUNX gene family in haematological malignancies. To elucidate the role of epigenetics in RUNX inactivation, we evaluated promoter DNA methylation of RUNX1, 2, and 3 in 23 leukaemia cell lines and samples from acute myeloid leukaemia (AML), acute lymphocytic leukaemia (ALL) and myelodysplatic syndromes (MDS) patients. RUNX1 and RUNX2 gene promoters were mostly unmethylated in cell lines and clinical samples. Hypermethylation of RUNX3 was frequent among cell lines (74%) and highly variable among patient samples, with clear association to cytogenetic status. High frequency of RUNX3 hypermethylation (85% of the 20 studied cases) was found in AML patients with inv(16)(p13.1q22) compared to other AML subtypes (31% of the other 49 cases). RUNX3 hypermethylation was also frequent in ALL (100% of the six cases) but low in MDS (21%). In support of a functional role, hypermethylation of RUNX3 was correlated with low levels of protein, and treatment of cell lines with the DNA demethylating agent, decitabine, resulted in mRNA re-expression. Furthermore, relapse-free survival of non-inv(16)(p13.1q22) AML patients without RUNX3 methylation was significantly better (P = 0·016) than that of methylated cases. These results suggest that RUNX3 silencing is an important event in inv(16)(p13.1q22) leukaemias.

Random survival forest model identifies novel biomarkers of event-free survival in high-risk pediatric acute lymphoblastic leukemia.

High-risk pediatric B-ALL patients experience 5-year negative event rates up to 25%. Although some biomarkers of relapse are utilized in the clinic, their ability to predict outcomes in high-risk patients is limited. Here, we propose a random survival forest (RSF) machine learning model utilizing interpretable genomic inputs to predict relapse/death in high-risk pediatric B-ALL patients. We utilized whole exome sequencing profiles from 156 patients in the TARGET-ALL study (with samples collected at presentation) further stratified into training and test cohorts (109 and 47 patients, respectively). To avoid overfitting and facilitate the interpretation of machine learning results, input genomic variables were engineered using a stepwise approach involving univariable Cox models to select variables directly associated with outcomes, genomic coordinate-based analysis to select mutational hotspots, and correlation analysis to eliminate feature co-linearity. Model training identified 7 genomic regions most predictive of relapse/death-free survival. The test cohort error rate was 12.47%, and a polygenic score based on the sum of the top 7 variables effectively stratified patients into two groups, with significant differences in time to relapse/death (log-rank P = 0.001, hazard ratio = 5.41). Our model outperformed other EFS modeling approaches including an RSF using gold-standard prognostic variables (error rate = 24.35%). Validation in 174 standard-risk patients and 3 patients who failed to respond to induction therapy confirmed that our RSF model and polygenic score were specific to high-risk disease. We propose that our feature selection/engineering approach can increase the clinical interpretability of RSF, and our polygenic score could be utilized for enhance clinical decision-making in high-risk B-ALL.

Investigating protein patterns in human leukemia cell line experiments: A Bayesian approach for extremely small sample sizes.

Human cancer cell line experiments are valuable for investigating drug sensitivity biomarkers. The number of biomarkers measured in these experiments is typically on the order of several thousand, whereas the number of samples is often limited to one or at most three replicates for each experimental condition. We have developed an innovative Bayesian approach that efficiently identifies clusters of proteins that exhibit similar patterns of expression. Motivated by the availability of ion mobility mass spectrometry data on cell line experiments in myelodysplastic syndrome and acute myeloid leukemia, our methodology can identify proteins that follow biologically meaningful trends of expression. Extensive simulation studies demonstrate good performance of the proposed method even in the presence of relatively small effects and sample sizes.

Calling Variants in the Clinic: Informed Variant Calling Decisions Based on Biological, Clinical, and Laboratory Variables.

Deep sequencing genomic analysis is becoming increasingly common in clinical research and practice, enabling accurate identification of diagnostic, prognostic, and predictive determinants. Variant calling, distinguishing between true mutations and experimental errors, is a central task of genomic analysis and often requires sophisticated statistical, computational, and/or heuristic techniques. Although variant callers seek to overcome noise inherent in biological experiments, variant calling can be significantly affected by outside factors including those used to prepare, store, and analyze samples. The goal of this review is to discuss known experimental features, such as sample preparation, library preparation, and sequencing, alongside diverse biological and clinical variables, and evaluate their effect on variant caller selection and optimization.

Preclinical activity of FF-10501-01, a novel inosine-5'-monophosphate dehydrogenase inhibitor, in acute myeloid leukemia.

BACKGROUND: FF-10501-01 is a selective inosine monophosphate dehydrogenase (IMPDH) inhibitor that has shown activity in cancer cell lines. We studied whether FF-10501-01 is effective in targeting a variety of hypomethylating agent (HMA)-sensitive and -resistant acute myelogenous leukemia (AML) cell lines. METHODS: We treated multiple cell lines (including HMA-resistant cells) with FF-10501-01 and analyzed proliferation, apoptosis, and cell cycle status. We also assessed HMA-FF-10501-01 combinations and the ability of extracellular guanosine to rescue cell proliferation in FF-10501-01-treated cells. We performed high-performance liquid chromatography (HPLC) to study guanine nucleotide levels in treated and untreated cells. Finally, we studied the effects of FF-10501-01 in fresh peripheral blood cells taken from AML patients. RESULTS: FF-10501-01 showed a strong dose-dependent effect on proliferation and induced apoptosis at approximately 30µM. The effects of FF-10501-01 treatment on cell cycle status were variable, with no statistically significant trends. Guanosine rescued proliferation in FF-10501-01-treated cells, and HPLC results showed significant decreases in phosphorylated guanosine levels in MOLM13 cells. FF-10501-01 effectively reduced proliferation at concentrations of 300µM and above in 3 primary AML samples. CONCLUSIONS: FF-10501-01 effectively induces AML cell death and reduces AML peripheral blood cell proliferation by targeting guanine nucleotide biosynthesis regardless of HMA resistance status.

PDE4 Differential Expression Is a Potential Prognostic Factor and Therapeutic Target in Patients With Myelodysplastic Syndrome and Chronic Myelomonocytic Leukemia.

BACKGROUND (OR PURPOSE): Inflammation has an essential role in the pathogenesis of myelodysplastic syndromes (MDS). Its expression is controlled by phosphodiesterase 4 (PDE4). Thus, PDE4 inhibitors might be useful therapeutic targets for MDS. PATIENTS (OR MATERIALS) AND METHODS: We evaluated the expression of each isoform of PDE4 (A, B, C, and D) using transcriptomic profiling and examined the potential impact on the outcome of patients with MDS in terms of survival and response to hypomethylating agents. Total RNA was extracted from CD34(+) bone marrow hematopoietic cells from healthy individuals (n = 10) and patients with MDS (n = 24) or chronic myelomonocytic leukemia (n = 19). RESULTS: The study cohort had a median follow-up period of 21.2 months (range, 0.2-68 months) and a median overall survival of 17.6 months (95% confidence interval, 9.6-25.6). The main finding of the present study was that PDE4 mean expression was generally higher in patients with MDS than in healthy individuals. Also, upregulated PDE4 expression seemed to have a possible negative effect on survival (P > .05). Moreover, lower, compared with higher, mean PDE4A and PDE4C expression is indicative of a response to a hypomethylating agent (0.09 and 0.03 vs. 0.54 and 0.49, respectively; P > .05). CONCLUSION: These results should be confirmed in a larger patient cohort. PDE4 expression could be an effective potential prognostic factor and therapeutic target for patients with MDS and chronic myelomonocytic leukemia. The role of PDE4 inhibitors should be explored in vitro against MDS cell lines and in preclinical mouse models of MDS.

Analysis of class I and II histone deacetylase gene expression in human leukemia.

Histone deacetylase (HDAC) inhibitors are well-characterized anti-leukemia agents and HDAC gene expression deregulation has been reported in various types of cancers. This study sought to characterize HDAC gene expression patterns in several types of leukemia. To do so, a systematic study was performed of the mRNA expression of all drug-targetable HDACs for which reagents were available. This was done by real-time PCR in 24 leukemia cell lines and 39 leukemia patients, which included AML, MDS and CLL patients, some of whom received HDAC inhibitor treatment. Among the samples analyzed, there was no discernible pattern in HDAC expression. HDAC expression was generally increased in CLL patients, except for HDAC2 and HDAC4. HDAC expression was also generally increased in VPA-treated MOLT4 cells. However, this increased expression was not seen in AML patients treated with vorinostat. In summary, increased HDAC expression was noted in CLL patients in general, but the HDAC expression patterns in myeloid malignancies appear to be heterogeneous, which implies that the role of HDACs in leukemia may be related to global expression or protein function rather than specific expression patterns.