Relapse is the leading cause of mortality in children with acute lymphoblastic leukemia (ALL). Among chemotherapeutics, thiopurines are key drugs in the backbone of ALL combination therapy. Using whole-exome sequencing, we identified relapse-specific mutations in phosphoribosyl pyrophosphate synthetase 1 (PRPS1), a rate-limiting purine biosynthesis enzyme, in 24/358 (6.7%) relapse B-ALL cases. All individuals who harbored PRPS1 mutations relapsed early on-treatment, and mutated ALL clones expanded exponentially prior to clinical relapse. Our functional analyses of PRPS1 mutants uncovered a new chemotherapy resistance mechanism involving reduced feedback inhibition of de novo purine biosynthesis and competitive inhibition of thiopurine activation. Notably, the de novo purine synthesis inhibitor lometrexol can effectively abrogate PRPS1 mutant-driven drug resistance. Overall these results highlight the importance of constitutive activation of de novo purine pathway in thiopurine resistance, and offer therapeutic strategies for the treatment of relapsed and resistant ALL.

The in vivo(212)Pb/(212)Bi generator is promising for application in targeted alpha therapy (TAT) of cancer. One main limitation of its therapeutic application is due to potential release of (212)Bi from the radioconjugate upon radioactive decay of the mother nuclide (212)Pb, potentially leading to irradiation of healthy tissue. The objective of the present work is to assess whether the chelate CHX-A''-DTPA (N-(2-aminoethyl)-trans-1,2-diaminocyclohexane-N,N',N''-pentaacetic acid) bound to a biological carrier molecule may be able to re-complex released (212)Bi under in vivo conditions to limit its translocation from the target site. CHX-A''-DTPA was bound to bovine gamma globulin (BGG) to mimic a model conjugate and the stability of the Bi-CHX-A''-DTPA-BGG conjugate was studied in blood serum by ultrafiltration. TRLFS experiments using Cm(III) as a fluorescent probe demonstrated that linking CHX-A''-DTPA to BGG does not affect the coordination properties of the ligand. Furthermore, comparable stability constants were observed between Bi(III) and free CHX-A''-DTPA, BGG-bound CHX-A''-DTPA and DTPA. The complexation constants determined between Bi(III) and the chelate molecules are sufficiently high to allow ultra trace amounts of the ligand to efficiently compete with serum transferrin controlling Bi(III) speciation in blood plasma conditions. Nevertheless, CHX-A''-DTPA is not able to complex Bi(III) generated in blood serum because of the strong competition between Bi(III) and Fe(II) for the ligand. In other words, CHX-A''-DTPA is not "selective" enough to limit Bi(iii) release in the body when applying the (212)Pb/(212)Bi in vivo generator.

Natural abundance deuterium 2D NMR spectroscopy in weakly ordering, polypeptide chiral liquid crystals is a powerful technique that enables determination of enantiotopic isotopic ratios ((2)H/(1)H)( i ) at the methylene groups of long-chain fatty acids. This technique has been used to study the bioconversion of linoleic acid to vernoleic acid with the objective of establishing the in-vivo site-specific fractionation of (2)H associated with this process. The fractionation pattern was investigated in Euphorbia lagascae and Vernonia galamensis, plants that use different enzyme systems to perform the Δ(12)-epoxidation: a cytochrome P450 monooxygenase in the former and a di-iron dioxygenase in the latter. The specific interest in this study was to ascertain whether different ((2)H/(1)H)( i ) isotopic ratios in substrate and product might reflect distinct features of the nature of the reaction centre. However, both the linoleate (substrate) samples and both vernoleate (product) samples isolated from the seed oils of the two plants had remarkably similar (2)H isotope profiles, with selection against (2)H in the positions around the Δ(12)-epoxidation site. This is interpreted as indicating that, despite differences in the form in which the activated Fe is presented and in the architecture of the active site, the ((2)H/(1)H)( i ) isotopic pattern is determined by features common to the reaction. It is suggested that the effects acting as the overall determinants of the final ((2)H/(1)H)( i ) distribution in the product are the encumbrance of the active site pocket and constraints to conformational readjustment during the linoleate to vernoleate transformation.

Indium-DTPA-tagged liposomes were studied in the present work as carriers of in vivo 212Pb/ 212Bi generator to be used in targeted alpha therapy. The liposomal uptake of 212Pb, into preformed liposomes, was investigated using different lipophilic chelate (DCP, 2,3-dimercapto-1-propanol (BAL), sodium acetate, and A23187), as a function of various parameters (temperature, concentrations of lipids, Pb, DTPA, ...) with 212Pb as a tracer. Different formulations of liposomes were tested to evaluate the radiolabeling efficiency. No complexing agent was necessary for the passage of Pb2+ through the membrane. It occurs naturally via a partial permeability of the lipid bilayer, which increases with the temperature. A complexing agent (DTPA) appears necessary to concentrate Pb in the internal compartment of the liposomes. Conditions were found (T=65 ºC, internal DTPA concentration of 0.025 M, pH 7.4, ...) yielding a high and rapid uptake of 212Pb in liposomes. The protocol established provides a novel method for the efficient entrapment of about 2–3 Pb atoms per liposome with a yield of 75% in conditions relevant for nuclear medicine.

The China Bioanalysis Forum (CBF), a volunteer and science focused group, was recently formed. The formation of CBF is a response to the urgent needs of updating current Chinese bioanalytical guidelines, the harmonization of Chinese and international guidelines, and establishing a platform for scientific exchange within the Chinese bioanalytical community. The mission, organization and the future plan of CBF are discussed in this short communication.

The interaction between thorium and human serum components was studied using difference ultraviolet spectroscopy (DUS), ultrafiltration and high-pressure-anion exchange chromatography (HPAEC) with external inductively conducted plasma mass spectrometry (ICP-MS) analysis. Experimental data are compared with modelling results based on the law of mass action. Human serum transferrin (HSTf) interacts strongly with Th(IV), forming a ternary complex including two synergistic carbonate anions. This complex governs Th(IV) speciation under blood serum conditions. Considering the generally used Langmuir-type model, values of 1033.5 and 1032.5 were obtained for strong and weak sites, respectively. We showed that trace amounts of diethylene triamine pentaacetic acid (DTPA) cannot complex Th(IV) in the blood serum at equilibrium. Unexpectedly this effect is not related to the competition with HSTf but is due to the strong competition with major divalent metal ions for DTPA. However, Th-DTPA complex was shown to be stable for a few hours when it is formed before addition in the biological medium; this is related to the high kinetic stability of the complex. This makes DTPA a potential chelating agent for synthesis of 226Th-labeled biomolecules for application in targeted alpha therapy

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Previously we reported the establishment of a pair of primary HCC cell lines from LIX-004, a patient-derived HCC xenograft model. LIXC004-NA was developed from an untreated PDX tumor; LIXC004-SR was generated from a PDX tumor progressed despite long term in vivo treatment with sorafenib. The two cell lines displayed similar response to sorafenib treatment in vitro. However, LIXC004-NA is sensitive and LIXC004-SR is resistant to sorafenib treatment in vivo. IHC and in vitro functional studies revealed activation of alternative angiogenic pathways as one of the potential reasons for the sorafenib resistance of LIXC004-SR. To fully explore different mechanisms/pathways, we performed a) micro-array analyses; b) whole exome sequencing; c) RNA-seq analyses of the two cell lines; d) metabolomic analyses of the cell lines in vitro culture in the presence and absence of different therapeutics; e) RNAseq analyses of the tumors derived from these two cell lines. The combination of different types of -omic analyses is our systematic approach to address the potential mechanisms of drug resistance without bias. The two cell lines shared a lot of the genetic mutation and expression profile for most genes. Differentially mutated or expressed genes were also identified at basal level. More changes were observed with the treatment of different therapeutics as well as analyses of the xenograft derived from the cell lines. Pathway analyses are performed to explore the underlying mechanisms for the development of drug resistance. Citation Format: Gang Hu, Alicia Du, Yong Huang, Kunyan Liu, Fubo Xie, Xuzhen Tang, Xueyan Yang, Qi Gu, Yixin Zhang, Weikang Tao, Yingjia Zhang, Wei Tang, He Zhou. Multiple -omic analyses of a pair of primary HCC tumor cell lines with different drug response revealed the mechanisms of drug resistance. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4425. doi:10.1158/1538-7445.AM2015-4425