Intracellular cytarabine triphosphate in circulating blasts post-treatment predicts remission status in patients with acute myeloid leukemia.

Cytarabine remains the backbone of therapy in acute myeloid leukemia (AML). The ability to assess intracellular cytarabine triphosphate (ara-CTP) levels in patients receiving cytarabine represents a major goal in the prediction of treatment response. This study, conducted within a clinical setting, aimed to assess ara-CTP levels in circulating peripheral blasts from non-M3 AML patients receiving cytarabine at one of three dosing levels, using a novel biosensor assay. Results from the initial 72 hours post-commencement were correlated with day 28 remission status, with feasibility parameters concurrently assessed. Intracellular ara-CTP was detectable in ex vivo blasts post-treatment for standard-dose (SD) and high-dose (HD) patients (p < 0.05), and quantification revealed a 27-fold increase in intracellular steady-state concentration between the two dosing levels. For low-dose cytarabine, high rates of patient discharge and low intracellular concentrations limited analysis; however, assessment of intracellular ara-CTP concentration was achievable in a dwindling population of blasts for SD and HD treatment cohorts, with 4 hours post-treatment commencement potentially being most predictive of clinical response (r = -0.912, p = 0.0113). Concurrent assessment of peripheral leukemia-associated immunophenotype (LAIP)-positive cells revealed a decline in burden (0-72 hours), which correlated with remission status (p < 0.05). Unexpectedly high rates of night sampling led to challenges associated with sampling rates, but did not have an impact on patient compliance. Additional training of night staff improved feasibility substantially. Multiple peripheral sampling during the initial 72 hours of treatment is feasible in newly diagnosed patients, and ara-CTP is detectable over the initial 24 hours, facilitating prediction of chemosensitivity of leukemic blasts to cytarabine.

[A Case of Intrathecal Infusion of Methotrexate and Ara-C for a Patient with Meningitis Due to Recurrent Gastric Cancer].

A 69-year-old woman underwent total gastrectomy for advanced gastric cancer with pyloric stenosis. She had a good postoperative course and was discharged 2 weeks after surgery. She received adjuvant chemotherapy with S-1 after discharge. One month after the initiation of the adjuvant chemotherapy, she complained of wobbling and weakness of her limbs. She stopped intake of S-1, but the symptoms did not improve. She was admitted to the hospital, but she became unconscious and had headache and blurred vision. We conducted a cerebrospinal fluid examination and made a diagnosis of meningeal carcinomatosis. After we started intrathecal infusion of methotrexate and Ara-C, referring to case reports clinical symptoms, including unconsciousness, headache, and left upper limb paralysis, improved and the CEA level in cerebrospinal fluid decreased.

Pharmacogenetics of deoxycytidine kinase: identification and characterization of novel genetic variants.

Deoxycytidine kinase (DCK) is a rate-limiting enzyme in the activation of nucleoside analogs such as cytarabine (ara-C), gemcitabine, clofarabine, and others. The present study was undertaken to identify and to determine the functional consequences of genetic variants in DCK. We sequenced 1.5 kilobases of the DCK proximal promoter and all seven coding exons in International HapMap Project panels (n = 90 each) with European (Centre d' Etude du Polymorphisme Humain; CEPH) or African (Yoruba people in Ibadan, Nigeria; YRI) ancestry. Sixty-four genetic polymorphisms, including three nonsynonymous coding changes (I24V, A119G, and P122S) were identified. Compared with DCK-wild-type (WT) protein, the activity of the recombinant DCK24Val, DCK119Gly, and DCK122Ser proteins was 85 +/- 5, 66 +/- 3, and 43 +/- 4%, respectively. DCK119Gly and DCK122Ser mutants had lower Km (p < 0.01) and Vmax (p < 0.001) compared with DCK-WT protein. Lymphoblast cell lines from subjects heterozygous for the coding changes had significantly lower DCK activity compared with homozygous WT subjects. Ethnic differences were observed, with African ancestry subjects demonstrating significantly higher DCK mRNA expression compared with subjects with European ancestry. In both CEPH and YRI subjects, the C allele of a 3'-untranslated region single-nucleotide polymorphism (SNP) (35708 C>T) was significantly associated with lower DCK mRNA expression. This SNP was strongly linked with other intronic SNPs, forming a major haplotype block in both ethnic groups. In an exploratory analysis, the 35708C allele was also associated with lower blast ara-C-5'-triphosphate (ara-CTP) levels in acute myeloid leukemia patients receiving ara-C as continuous infusion. These results suggest that genetic variation in DCK influences its activity and expression and may predict the variability observed in intracellular levels of the ara-C active metabolite ara-CTP.

Polymeric nanogel formulations of nucleoside analogs.

Nanogels are colloidal microgel carriers that have been recently introduced as a prospective drug delivery system for nucleotide therapeutics. The crosslinked protonated polymer network of nanogels binds oppositely charged drug molecules, encapsulating them into submicron particles with a core-shell structure. The nanogel network also provides a suitable template for chemical engineering, surface modification and vectorisation. This review reveals recent attempts to develop novel drug formulations of nanogels with antiviral and antiproliferative nucleoside analogs in the active form of 5'-triphosphates, discusses structural approaches to the optimisation of nanogel properties, and discusses the development of targeted nanogel drug formulations for systemic administration. Notably, nanogels can improve the CNS penetration of nucleoside analogs that are otherwise restricted from passing across the blood-brain barrier. The latest findings reviewed here demonstrate an efficient intracellular release of nucleoside analogs, encouraging further applications of nanogel carriers for targeted drug delivery.

Development of an in vitro stage prior to neural transplantation: substrate adhesion of cell aggregates

Células fetales obtenidas del cuerpo estriado, corteza cerebral, septum y mesencéfalo ventral fueron sometidas a condiciones de cultivo en rotación durante varios períodos, con o sin el agregado de suero, o tratamiento con arabinósido de citosina (ARA-C). Luego de varios días en cultivo rotatorio, falta de suero en el medio de cultivo o tratamiento con ARA-C se observó reducción del número de agregados adherentes a cápsulas pretratadas con polilisina o Matrigel. Asimismo, luego de varios días en cultivo rotatorio se observó disminución de la adhesividad a una monocapa de células nerviosas fetales. Agregados de segunda generación resultantes de la tripsinización inicial y el ulterior reagregado durante 24 h de agregados previamente no adherentes, mostraron índices de adhesividad similares a los de primera generación. Estas observaciones sugieren que, en las presentes condiciones de cultivo, moléculas superficiales asociadas a membrana que median la adhesividad celular a sustrato sufren cambios reversibles con el tiempo. Puesto que la mayor parte de las condiciones de cultivo mencionadas afectan en forma primaria a las células de la glía, se sugiere también que este tipo celular sería básicamente responsable de los cambios observados en la adhesión a sustrato. Se considera que estos resultados tienen implicancia para el desarrollo de un estadio intermedio de cultivo "in vitro", previo al transplante de células nerviosas

Development of an in vitro stage prior to neural transplantation: substrate adhesion of cell aggregates

Células fetales obtenidas del cuerpo estriado, corteza cerebral, septum y mesencéfalo ventral fueron sometidas a condiciones de cultivo en rotación durante varios períodos, con o sin el agregado de suero, o tratamiento con arabinósido de citosina (ARA-C). Luego de varios días en cultivo rotatorio, falta de suero en el medio de cultivo o tratamiento con ARA-C se observó reducción del número de agregados adherentes a cápsulas pretratadas con polilisina o Matrigel. Asimismo, luego de varios días en cultivo rotatorio se observó disminución de la adhesividad a una monocapa de células nerviosas fetales. Agregados de segunda generación resultantes de la tripsinización inicial y el ulterior reagregado durante 24 h de agregados previamente no adherentes, mostraron índices de adhesividad similares a los de primera generación. Estas observaciones sugieren que, en las presentes condiciones de cultivo, moléculas superficiales asociadas a membrana que median la adhesividad celular a sustrato sufren cambios reversibles con el tiempo. Puesto que la mayor parte de las condiciones de cultivo mencionadas afectan en forma primaria a las células de la glía, se sugiere también que este tipo celular sería básicamente responsable de los cambios observados en la adhesión a sustrato. Se considera que estos resultados tienen implicancia para el desarrollo de un estadio intermedio de cultivo "in vitro", previo al transplante de células nerviosas (AU)