Purpose: To evaluate the preclinical pharmacokinetics and antitumor efficacy of a novel orally bioavailable poly(ADP-ribose) polymerase (PARP) inhibitor, ABT-888. Experimental Design: In vitro potency was determined in a PARP-1 and PARP-2 enzyme assay. In vivo efficacy was evaluated in syngeneic and xenograft models in combination with temozolomide, platinums, cyclophosphamide, and ionizing radiation. Results: ABT-888 is a potent inhibitor of both PARP-1 and PARP-2 with K i s of 5.2 and 2.9 nmol/L, respectively.The compound has good oral bioavailability and crosses the blood-brain barrier. ABT-888 strongly potentiated temozolomide in the B16F10 s.c. murine melanoma model. PARP inhibition dramatically increased the efficacy of temozolomide at ABT-888 doses as low as 3.1 mg/kg/d and a maximal efficacy achieved at 25 mg/kg/d. In the 9L orthotopic rat glioma model, temozolomide alone exhibited minimal efficacy, whereas ABT-888, when combined with temozolomide, significantly slowed tumor progression. In the MX-1breast xenograft model (BRCA1 deletion and BRCA2 mutation), ABT-888 potentiated cisplatin, carboplatin, and cyclophosphamide, causing regression of established tumors, whereas with comparable doses of cytotoxic agents alone, only modest tumor inhibition was exhibited. Finally, ABT-888 potentiated radiation (2 Gy/d  10) in an HCT-116 colon carcinoma model. In each model, ABT-888 did not display single-agent activity. Conclusions: ABT-888 is a potent inhibitor of PARP, has good oral bioavailability, can cross the blood-brain barrier, and potentiates temozolomide, platinums, cyclophosphamide, and radiation in syngeneic and xenograft tumor models. This broad spectrum of chemopotentiation and radiopotentiation makes this compound an attractive candidate for clinical evaluation.poly(ADP-ribose) polymerase (PARP)-1 is the founding member of a family of poly(ADP-ribosyl)ating proteins. All PARP family members are characterized by the ability to poly(ADP-ribosyl)ate protein substrates and all share a catalytic PARP homology domain (1). PARP-1 and the closely related PARP-2 are nuclear proteins and the only PARPs with DNA binding domains. These DNA binding domains localize PARP-1 and PARP-2 to the site of DNA damage serving as DNA damage sensors and signaling molecules for repair. The knockout of PARP-1 is sufficient to significantly impair DNA repair following damage via radiation (2) or cytotoxic (3) insult. The residual PARP-dependent repair activity (f10%) is due to PARP-2 (4, 5). These data imply that inhibition of only PARP-1 and PARP-2 will impair DNA repair following damage and that inhibition of other PARP family members is not required in the process. The functions of other PARP family members remain to be elucidated, but poly(ADP-ribosyl)ation has been implicated in many cellular processes, including differentiation, gene regulation, protein degradation, spindle maintenance, as well as replication and transcription (6).Higher expression of PARP in cancer compared with normal cells has been linked to...

Cells of the mononuclear phagocyte system are the predominant cell producing HIV-1 in most tissues including the central nervous system (CNS), spinal cord, lung and skin; infection is associated with dementia, neuropathy, pneumonitis, and dermatitis respectively. Different HIV-1 isolates vary markedly in their ability to infect mononuclear phagocytes productively. Here we describe molecular clones of a CNS-derived isolate, HIV-1(JR-FL), which can replicate efficiently in mononuclear phagocytes. Analysis by polymerase chain reaction of early events after infection indicates that the early phase of viral replication before reverse transcription determines tropism. Genetic mapping of the macrophage-tropic phenotype by construction of recombinant viruses indicates that mononuclear phagocyte infectivity can be determined by a 157-amino-acid region of the gp 120 glycoprotein of HIV-1(JR-FL). Significantly, this region is upstream from the previously defined CD4-binding domain. We propose that at least one determinant for mononuclear phagocyte tropism involves target cell interactions with regions of gp120 distinct from the CD4-binding domain.

Cloned cDNA and genomic sequences have been analyzed to deduce the amino acid sequence of phytochrome from etiolated Avena. Restriction endonuclease site polymorphism between clones indicates that at least four phytochrome genes are expressed in this tissue. Sequence analysis of two complete and one partial coding region shows approximately 98% homology at both the nucleotide and amino acid levels, with the majority of amino acid changes being conservative. High sequence homology is also found in the 5'-untranslated region but significant divergence occurs in the 3'-untranslated region. The phytochrome polypeptides are 1128 amino acid residues long corresponding to a molecular mass of 125 kdaltons. The known protein sequence at the chromophore attachment site occurs only once in the polypeptide, establishing that phytochrome has a single chromophore per monomer covalently linked to Cys-321. Computer analyses of the amino acid sequences have provided predictions regarding a number of structural features of the phytochrome molecule.

BackgroundVenetoclax (ABT-199), a first-in-class orally bioavailable BCL-2-selective inhibitor, was recently approved by the FDA for use in patients with 17p-deleted chronic lymphocytic leukemia who have received prior therapy. It is also being evaluated in numerous clinical trials for treating patients with various hematologic malignancies. As with any targeted cancer therapy, it is critically important to identify potential mechanisms of resistance, both for patient stratification and developing strategies to overcome resistance, either before it develops or as it emerges.MethodsIn order to gain a more comprehensive insight into the nature of venetoclax resistance mechanisms, we evaluated the changes in the BCL-2 family members at the genetic and expression levels in seven different venetoclax-resistant derived leukemia and lymphoma cell lines.ResultsGene and protein expression analyses identified a number of different alterations in the expression of pro- and anti-apoptotic BCL-2 family members. In the resistant derived cells, an increase in either or both the anti-apoptotic proteins BCL-XL or MCL-1, which are not targeted by venetoclax was observed, and either concomitant or exclusive with a decrease in one or more pro-apoptotic proteins. In addition, mutational analysis also revealed a mutation in the BH3 binding groove (F104L) that could potentially interfere with venetoclax-binding. Not all changes may be causally related to venetoclax resistance and may only be an epiphenomenon. For resistant cell lines showing elevations in BCL-XL or MCL-1, strong synergistic cell killing was observed when venetoclax was combined with either BCL-XL- or MCL-1-selective inhibitors, respectively. This highlights the importance of BCL-XL- and MCL-1 as causally contributing to venetoclax resistance.ConclusionsOverall our study identified numerous changes in multiple resistant lines; the changes were neither mutually exclusive nor universal across the cell lines tested, thus exemplifying the complexity and heterogeneity of potential resistance mechanisms. Identifying and evaluating their contribution has important implications for both patient selection and the rational development of strategies to overcome resistance.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-017-3383-5) contains supplementary material, which is available to authorized users.

Clinical applications of precision oncology require accurate tests that can distinguish true cancer specific mutations from errors introduced at each step of next-generation sequencing (NGS). To date, no bulk sequencing study has addressed the effects of cross-site reproducibility, nor the biological, technical and computational factors that influence variant identification. Here we report a systematic interrogation of somatic mutations in paired tumor-normal cell lines to identify factors affecting detection reproducibility and accuracy at six different centers. Using whole genome sequencing (WGS) and whole-exome sequencing (WES), we evaluated the reproducibility of different sample types with varying input amount and tumor purity, and multiple library construction protocols, followed by processing with nine bioinformatics pipelines. We found that read coverage and callers affected both WGS and WES reproducibility, but WES performance was influenced by insert fragment size, genomic copy content and the global imbalance score (GIV; G > T/C > A). Finally, taking into account library preparation protocol, tumor content, read coverage and bioinformatics processes concomitantly, we recommend actionable practices to improve the reproducibility and accuracy of NGS experiments for cancer mutation detection.