Aptamers Entirely Built from Therapeutic Nucleoside Analogues for Targeted Cancer Therapy.

Owing to the specific and high binding affinity of aptamers to their targets, aptamer-drug conjugates (ApDCs) have emerged as a promising drug delivery system for targeted cancer therapy. However, in a conventional ApDC, the aptamer segment usually just serves as a targeting moiety, and only a limited number of drug molecules are sequentially conjugated to the oligonucleotide, giving a relatively low drug loading capacity. To address this challenge, herein we employ four clinically approved nucleoside analogues, including clofarabine (Clo), ara-guanosine (AraG), gemcitabine (Ge), and floxuridine (FdU), to replace all natural nucleosides in aptamer sequences, generating a series of whole drug-constituted DNA-like oligomers that are termed drugtamers. Similar to their parent aptamers, the obtained drugtamers maintain the targeting capability and can specifically bind to the target receptors overexpressed on the cancer cell surface. With 100% drug loading ratio, active targeting capability, and enzyme-mediated release of active therapeutics, our drugtamers can strongly induce the apoptosis of cancer cells and inhibit the tumor progression, which enables a new potential for a better targeted cancer therapy.

Testicular disposition of clofarabine in rats is dependent on equilibrative nucleoside transporters.

Acute lymphoblastic leukemia (ALL) is the most common cancer in children and adolescents. Although the 5-year survival rate is high, some patients respond poorly to chemotherapy or have recurrence in locations such as the testis. The blood-testis barrier (BTB) can prevent complete eradication by limiting chemotherapeutic access and lead to testicular relapse unless a chemotherapeutic is a substrate of drug transporters present at this barrier. Equilibrative nucleoside transporter (ENT) 1 and ENT2 facilitate the movement of substrates across the BTB. Clofarabine is a nucleoside analog used to treat relapsed or refractory ALL. This study investigated the role of ENTs in the testicular disposition of clofarabine. Pharmacological inhibition of the ENTs by 6-nitrobenzylthioinosine (NBMPR) was used to determine ENT contribution to clofarabine transport in primary rat Sertoli cells, in human Sertoli cells, and across the rat BTB. The presence of NBMPR decreased clofarabine uptake by 40% in primary rat Sertoli cells (p = .0329) and by 53% in a human Sertoli cell line (p = .0899). Rats treated with 10 mg/kg intraperitoneal (IP) injection of the NBMPR prodrug, 6-nitrobenzylthioinosine 5'-monophosphate (NBMPR-P), or vehicle, followed by an intravenous (IV) bolus 10 mg/kg dose of clofarabine, showed a trend toward a lower testis concentration of clofarabine than vehicle (1.81 ± 0.59 vs. 2.65 ± 0.92 ng/mg tissue; p = .1160). This suggests that ENTs could be important for clofarabine disposition. Clofarabine may be capable of crossing the human BTB, and its potential use as a first-line treatment to avoid testicular relapse should be considered.

Population Pharmacokinetics of Clofarabine as Part of Pretransplantation Conditioning in Pediatric Subjects before Hematopoietic Cell Transplantation.

The primary objective of this work was to characterize the pharmacokinetics (PK) of systemic clofarabine (clo-fara) in pediatric allogeneic hematopoietic cell transplantation (HCT) recipients receiving either nucleoside monotherapy or a dual nucleoside analog preparative regimen. Fifty-one children (median age, 4.9 years; range, .25 to 14.9 years) undergoing allogeneic HCT for a variety of malignant and nonmalignant disorders underwent PK assessment. Plasma samples were collected over the 4 to 5 days of clo-fara treatment and quantified for clo-fara, using a validated liquid chromatography/tandem mass spectrometry assay. Nonlinear mixed-effects modeling was used to develop the population PK model, including identification of covariates that influenced drug disposition. In agreement with previously published models, a 2-compartment PK model with first-order elimination best described the PK of clo-fara. Final parameter estimates for clo-fara were consistent with previous reports and were as follows: clearance (CL), 23 L/h/15 kg; volume of the central compartment, 42 L/15 kg; volume of peripheral compartment, 47 L/15 kg; and intercompartmental CL, 9.8 L/h/15 kg. Unexplained variability was acceptable at 33%, and the additive residual error (reflective of the assay) was estimated to be 0.36 ng/mL. Patient-specific factors significantly impacting clo-fara CL included actual body weight and age. The covariate model was able to estimate clo-fara CL with good precision in children spanning a wide age range from infancy to early adulthood and demonstrates the need for variable dosing in children of different ages. For example, the dose required for a 6-month and 1-year old was approximately 43% and 17% lower, respectively, than the typical 40 mg/m2dose to achieve the median AUC0-24of 1.04 mg·h/L in the study population. Despite the known renal elimination of clo-fara, no significant clinical parameters for renal function were retained in the final model (P> .05). Coadministration of fludarabine with clo-fara did not alter the CL of clo-fara (P> .05). These results will help inform individualized dosing strategies for clo-fara to improve clinical outcomes and limit drug-related adverse events in children undergoing HCT.

Nucleoside Analogue-Based Supramolecular Nanodrugs Driven by Molecular Recognition for Synergistic Cancer Therapy.

The utilization of nanotechnology for the delivery of a wide range of anticancer drugs has the potential to reduce adverse effects of free drugs and improve the anticancer efficacy. However, carrier materials and/or chemical modifications associated with drug delivery make it difficult for nanodrugs to achieve clinical translation and final Food and Drug Administration (FDA) approvals. We have discovered a molecular recognition strategy to directly assemble two FDA-approved small-molecule hydrophobic and hydrophilic anticancer drugs into well-defined, stable nanostructures with high and quantitative drug loading. Molecular dynamics simulations demonstrate that purine nucleoside analogue clofarabine and folate analogue raltitrexed can self-assemble into stable nanoparticles through molecular recognition. In vitro studies exemplify how the clofarabine:raltitrexed nanoparticles could greatly improve synergistic combination effects by arresting more G1 phase of the cell cycle and reducing intracellular deoxynucleotide pools. More importantly, the nanodrugs increase the blood retention half-life of the free drugs, improve accumulation of drugs in tumor sites, and promote the synergistic tumor suppression property in vivo.

Clofarabine, high-dose cytarabine and liposomal daunorubicin in pediatric relapsed/refractory acute myeloid leukemia: a phase IB study.

Survival in children with relapsed/refractory acute myeloid leukemia is unsatisfactory. Treatment consists of one course of fludarabine, cytarabine and liposomal daunorubicin, followed by fludarabine and cytarabine and stem-cell transplantation. Study ITCC 020/I-BFM 2009-02 aimed to identify the recommended phase II dose of clofarabine replacing fludarabine in the abovementioned combination regimen (3+3 design). Escalating dose levels of clofarabine (20-40 mg/m2/day × 5 days) and liposomal daunorubicin (40-80 mg/m2/day) were administered with cytarabine (2 g/m2/day × 5 days). Liposomal DNR was given on day 1, 3 and 5 only. The cohort at the recommended phase II dose was expanded to make a preliminary assessment of anti-leukemic activity. Thirty-four children were enrolled: refractory 1st (n=11), early 1st (n=15), ≥2nd relapse (n=8). Dose level 3 (30 mg/m2clofarabine; 60 mg/m2liposomal daunorubicin) appeared to be safe only in patients without subclinical fungal infections. Infectious complications were dose-limiting. The recommended phase II dose was 40 mg/m2 clofarabine with 60 mg/m2 liposomal daunorubicin. Side-effects mainly consisted of infections. The overall response rate was 68% in 31 response evaluable patients, and 80% at the recommended phase II dose (n=10); 22 patients proceeded to stem cell transplantation. The 2-year probability of event-free survival (pEFS) was 26.5±7.6 and probability of survival (pOS) 32.4±8.0%. In the 21 responding patients, the 2-year pEFS was 42.9±10.8 and pOS 47.6±10.9%. Clofarabine exposure in plasma was not significantly different from that in single-agent studies. In conclusion, clofarabine was well tolerated and showed high response rates in relapsed/refractory pediatric acute myeloid leukemia. Patients with (sub) clinical fungal infections should be treated with caution. Clofarabine has been taken forward in the Berlin-Frankfurt-Münster study for newly diagnosed acute myeloid leukemia. The Study ITCC-020 was registered as EUDRA-CT 2009-009457-13; Dutch Trial Registry number 1880.