Subcutaneous Site-of-Absorption Study with the Monoclonal Antibody Tocilizumab in Minipigs: Administration Behind Ear Translates Best to Humans.

Minipigs have been proposed as animal model to study the subcutaneous (SC) absorption of monoclonal antibodies (mAb), because they are more translatable to humans than other species. However, the minipig SC tissue structure differs markedly depending on its location. This study explored different SC administration sites for mAb SC administration, to explore which site translates best to humans. The study assessed the SC absorption of tocilizumab (Actemra®) following administration at several injection sites: Inguinal area, flank, caudal to the ear, and interscapular area, in comparison with an IV group. After SC administration, tocilizumab absorption was most rapid from the inguinal administration site, and slowest after administration behind the ear, with absorption from the other sites in between. Tocilizumab bioavailability was 98.6, 88.3, 74.1, and 86.3% after administration in inguinal area, flank, behind the ear, and interscapular area, as determined by non-compartmental analysis. Fitting of a single first-order absorption rate constant by compartmental analysis was dissatisfactory. A combined fitting of all data was done assuming two different kinds of SC depots, one undergoing fast absorption, the other undergoing a slower absorption. The split between these absorption depots differed across administration sites, with absorption from "fast depot" in inguinal area > flank > interscapular area > behind the ear. Comparisons with clinical data show that tocilizumab PK after SC administration behind the ear translates best to humans, considering both bioavailability and rate of absorption. Whether this translation from minipigs to humans is prototypic for other mAb remains to be confirmed.

Special feature of mixed phosphotriester derivatives of cytarabine.

Despite the unquestionable therapeutic interest of bis(SATE) pronucleotides, a presystemic metabolism preventing the delivery of the prodrugs in target cancer cells or tumours may constitute a limitation to the in vivo development of such derivatives. In order to overcome these drawbacks several strategies have been envisaged and we report herein the application of the S-acyl-2-thioethyl (SATE) phenyl pronucleotide approach to the well-known cytotoxic nucleoside cytosine-1-beta-D-arabinofuranoside (cytarabine, araC). We describe modifications of the SATE moieties with the introduction of polar groups on the acyl residue, in order to study how these changes affect antitumoral activity and metabolic stability. Two different synthetic pathways were explored and lead to obtain the corresponding mixed derivatives in satisfactory yields. Cytotoxicity was studied in murine leukaemia cells L1210 as well as in cells derived from solid human tumours (Messa and MCF7). Biological evaluation of these compounds in cell culture experiments with nucleoside analogue-sensitive and resistant cell lines showed that the modified compounds were active at higher concentrations than unmodified cytarabine, yet were much able to partially reverse resistance due to deficient nucleoside transport or activation. These results can be correlated with an incomplete decomposition mechanism into the corresponding 5'-mononucleotide.

Preparation of 6,6,1',1',6',6'-hexadeutero sucrose.

The preparation of 6,6,1',1',6',6'-hexadeutero sucrose is reported. The synthesis is based on a triple oxidation of a protected sucrose 6,1',6'-triol to the corresponding 6,1',6'-tricarboxylic acid or ester, followed by reduction with lithium aluminium deuteride. This triple oxidation could be achieved either using cat. TEMPO-NaOCl (to the acid) or PDC-Ac(2)O-t-BuOH (to the t-butyl carboxylic ester).

Characterization of a gemcitabine-resistant murine leukemic cell line: reversion of in vitro resistance by a mononucleotide prodrug.

Resistance to cytotoxic nucleoside analogues is a major problem in cancer treatment. The cellular mechanisms involved in this phenomenon have been studied for several years, and some factors have been identified. Various strategies to overcome resistance have been suggested, but none has yet shown efficacy in vivo. We developed a gemcitabine-resistant cell line (L1210 10K) from the murine leukemic L1210 strain (L1210 wt) by continuous exposure to increasing concentrations of gemcitabine. L1210 10K is highly resistant to gemcitabine (14,833-fold), 1-beta-D-arabinofuranosylcytosine (ara-C; 2,100-fold), troxacitabine (>200-fold), and cladribine (160-fold) and slightly resistant to trimidox (7.22-fold), but does not display cross-resistance to fludarabine or nonnucleoside anticancer drugs. Deoxycytidine kinase mRNA was not detected by quantitative real-time reverse transcription-PCR in L1210 10K cells, whereas expression of thymidine kinase 1 and ribonucleotide reductase subunit R2 gene was moderately reduced. L1210 10K cells also demonstrated in vivo resistance to nucleoside analogues: gemcitabine- or ara-C-treated mice carrying L1210 10K had significantly shorter survival than gemcitabine- or ara-C-treated mice carrying L1210 wt (P < 0.05). UA911, a mononucleotide prodrug (pronucleotide) of ara-C was found to significantly sensitize L1210 10K cells in vitro. These results suggest that reduced deoxycytidine kinase expression is a mechanism of resistance to gemcitabine that is relevant in vivo and can be circumvented by a prodrug approach.