CPX-351 exhibits hENT-independent uptake and can be potentiated by fludarabine in leukaemic cells lines and primary refractory AML

© 2018 Elsevier Ltd CPX-351, a liposomal formulation co-encapsulating cytarabine and daunorubicin (DNR) in a synergistic 5:1 M ratio, has shown favourable response in newly diagnosed elderly high-risk AML. This study assessed intracellular ara-CTP levels following in vitro exposure of human immortalised leukaemic cell lines and primary AML blasts to CPX-351, and investigated fludarabine potentiation of intracellular ara-CTP formation from CPX-351. Comparison of intracellular handling of CPX-351 to cytarabine in HL-60 cells indicated slower conversion to ara-CTP for CPX-351, but equivalent cytotoxicity to cytarabine and combined DNR/cytarabine (DA) at 48 h, mostly likely reflecting the need for intracellular liposome processing to release encapsulated drugs. Further assessment demonstrated cytotoxicity of CPX-351 to be superior to DA at 48 and 72 h in cytarabine-resistant THP-1 cells (p < 0.001), and this effect could not be inhibited upon blockade of human equilibrative nucleoside transporter (hENT) function with dipyridamole. Assessment of Flu-CPX in primary blasts from presentation AML patients (n = 5) demonstrated a more rapid and pronounced potentiation of ara-CTP from CPX-351 than in immortalised cell lines, with 4/5 patients showing significant increases in ara-CTP, notably for those that went on to fail induction and relapse treatment in vivo (n = 3). This suggests a favourable impact on patient outcome from Flu-CPX

The human equilibrative nucleoside transporter 1 mediates in vitro cytarabine sensitivity in childhood acute myeloid leukaemia

Cytarabine (ara-C) is the most effective agent for the treatment of acute myeloid leukaemia (AML). Aberrant expression of enzymes involved in the transport/metabolism of ara-C could explain drug resistance. We determined mRNA expression of these factors using quantitative-real-time-PCR in leukemic blasts from children diagnosed with de novo AML. Expression of the inactivating enzyme pyrimidine nucleotidase-I (PN-I) was 1.8-fold lower in FAB-M5 as compared to FAB-M1/2 (P=0.007). In vitro sensitivity to deoxynucleoside analogues was determined using the MTT-assay. Human equilibrative nucleoside transporter-1 (hENT1) mRNA expression and ara-C sensitivity were significantly correlated (rp=−0.46; P=0.001), with three-fold lower hENT1 mRNA levels in resistant patients (P=0.003). hENT1 mRNA expression also seemed to correlate inversely with the LC50 values of cladribine (rp=−0.30; P=0.04), decitabine (rp=−0.29; P=0.04) and gemcitabine (rp=−0.33; P=0.02). Deoxycytidine kinase (dCK) and cytidine deaminase (CDA) mRNA expression seemed to correlate with in vitro sensitivity to gemcitabine (rp=−0.31; P=0.03) and decitabine (rp=0.33; P=0.03), respectively. The dCK/PN-I ratio correlated inversely with LC50 values for gemcitabine (rp=−0.45, P=0.001) and the dCK/CDA ratio seemed to correlate with LC50 values for decitabine (rp=−0.29; 0.04). In conclusion, decreased expression of hENT1, which transports ara-C across the cell membrane, appears to be a major factor in ara-C resistance in childhood AML

How to design the cost-effectiveness appraisal process of new healthcare technologies to maximise population health : A conceptual framework

This paper presents a conceptual framework to analyse the design of the cost-effectiveness appraisal process of new healthcare technologies. The framework characterises the appraisal processes as a diagnostic test aimed at identifying cost-effective (true positive) and non-cost-effective (true negative) technologies. Using the framework, factors that influence the value of operating an appraisal process, in terms of net gain to population health, are identified. The framework is used to gain insight into current policy questions including (a) how rigorous the process should be, (b) who should have the burden of proof, and (c) how optimal design changes when allowing for appeals, price reductions, resubmissions, and re-evaluations. The paper demonstrates that there is no one optimal appraisal process and the process should be adapted over time and to the specific technology under assessment. Optimal design depends on country-specific features of (future) technologies, for example, effect, price, and size of the patient population, which might explain the difference in appraisal processes across countries. It is shown that burden of proof should be placed on the producers and that the impact of price reductions and patient access schemes on the producer's price setting should be considered when designing the appraisal process

Therapeutic potential of cladribine in combination with STAT3 inhibitor against multiple myeloma

<p>Abstract</p> <p>Background</p> <p>Cladribine or 2-chlorodeoxyadenosine (2-CDA) is a well-known purine nucleoside analog with particular activity against lymphoproliferative disorders, such as hairy cell leukemia (HCL). Its benefits in multiple myeloma (MM) remain unclear. Here we report the inhibitory effects of cladribine on MM cell lines (U266, RPMI8226, MM1.S), and its therapeutic potential in combination with a specific inhibitor of the signal transducer and activator of transcription 3 (STAT3).</p> <p>Methods</p> <p>MTS-based proliferation assays were used to determine cell viability in response to cladribine. Cell cycle progression was examined by flow cytometry analysis. Cells undergoing apoptosis were evaluated with Annexin V staining and a specific ELISA to quantitatively measure cytoplasmic histone-associated DNA fragments. Western blot analyses were performed to determine the protein expression levels and activation.</p> <p>Results</p> <p>Cladribine inhibited cell proliferation of MM cells in a dose-dependent manner, although the three MM cell lines exhibited a remarkably different responsiveness to cladribine. The IC50 of cladribine for U266, RPMI8226, or MM1.S cells was approximately 2.43, 0.75, or 0.18 μmol/L, respectively. Treatment with cladribine resulted in a significant G1 arrest in U266 and RPMI8226 cells, but only a minor increase in the G1 phase for MM1.S cells. Apoptosis assays with Annexin V-FITC/PI double staining indicated that cladribine induced apoptosis of U266 cells in a dose-dependent manner. Similar results were obtained with an apoptotic-ELISA showing that cladribine dramatically promoted MM1.S and RPMA8226 cells undergoing apoptosis. On the molecular level, cladribine induced PARP cleavage and activation of caspase-8 and caspase-3. Meanwhile, treatment with cladribine led to a remarkable reduction of the phosphorylated STAT3 (P-STAT3), but had little effect on STAT3 protein levels. The combinations of cladribine and a specific STAT3 inhibitor as compared to either agent alone significantly induced apoptosis in all three MM cell lines.</p> <p>Conclusions</p> <p>Cladribine exhibited inhibitory effects on MM cells <it>in vitro</it>. MM1.S is the only cell line showing significant response to the clinically achievable concentrations of cladribine-induced apoptosis and inactivation of STAT3. Our data suggest that MM patients with the features of MM1.S cells may particularly benefit from cladribine monotherapy, whereas cladribine in combination with STAT3 inhibitor exerts a broader therapeutic potential against MM.</p

Cladribine with cyclophosphamide and prednisone in the management of low-grade lymphoproliferative malignancies

The feasibility of combining cladribine with cyclophosphamide and prednisone in the management of indolent lymphoid malignancies was determined. Nineteen patients [nine chronic lymphocytic leukaemia (CLL), seven non-Hodgkin's lymphoma (NHL) and three macroglobulinaemia (M))] received cladribine 0.1 mg kg−1 per day as a subcutaneous bolus injection on days 1–3 (up to 5 injections) with intravenous cyclophosphamide 500 mg m−2 on day 1 and oral prednisone 40 mg m−2 on days 1–5 at 4-weekly intervals up to a maximum of six courses. A total of 80 courses were given. Overall response rate was 88%, with four patients achieving a complete clinical and haematological response and 12 achieving a partial response. Neutropenia WHO grade 4 in two patients and WHO grade 3 infection in one patient were the limiting toxicities on treatment. During the follow-up, WHO grade ≥3 haematological complications occurred in five patients and WHO grade ≥3 non-haematological complications in five patients. There were no treatment-related deaths. This study demonstrates the feasibility of the cladribine/cyclophosphamide/prednisone (CCP) combination that appears highly active and safe in the management of indolent lymphoid malignancies. © 1999 Cancer Research Campaig

The role of deoxycytidine-metabolizing enzymes in the cytotoxicity induced by 3′-amino-2′,3′-dideoxycytidine and cytosine arabinoside

The cellular metabolism of 3′-amino-2′,3′-dideoxycytidine (3′-NH 2 -dCyd), a cytotoxic agent previously reported to be a poor substrate for purified Cyd/dCyd deaminase (dCydD), was compared with that of cytosine arabinoside (ara-C) in cells that displayed dCydD activity (HeLa) and in cells that did not (L1210). Growth inhibition induced by 3′-NH 2 -dCyd was dependent on the levels of anabolic enzymes, particularly dCyd kinase (dCydK), whereas cytotoxicity induced by ara-C was dependent on the expression of both anabolic and catabolic enzyme activities. Competition kinetics using purified enzyme revealed that the binding affinity of ara-C to dCydK was 5-fold that of the amino analog. However, this binding advantage is apparently offset in cells that contain high levels of dCydD, since the K i values for this enzyme were 0.2 and 23 mm for ara-C and 3′-NH 2 -dCyd, respectively. This was reflected in the decrease in analog sensitivity observed between the two cell lines, whereby the concentrations of ara-C and 3′-NH 2 -dCyd required to inhibit growth by 50% were 200 and 7 times higher, respectively, in the dCydD-containing HeLa cells as compared with the dCydD-deficient L1210 cells. The metabolic stability and cytotoxicity of 3′-NH 2 -dCyd was independent of cell number. An unexpected finding was the extent to which the effectiveness of ara-C could be mitigated by the number of dCydD-containing cells. A completely cytotoxic concentration of ara-C was rendered nontoxic by a 10-fold increase in cell number. This observation was supported by an increase in I-β- d -arabinofuranosyluracil (ara-U) formation, a decrease in ara-C 5′-triphosphate (ara-CTP) accumulation, and a rise in cell viability with increasing cell number. These findings indicate that unlike ara-C, the effectiveness of 3′-NH 2 -dCyd is independent of the level of deaminase, which suggests its possible utility in situations in which high levels of deaminase are manifest.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/46922/1/280_2004_Article_BF00686406.pd