Poly(ADP-ribose) polymerases have shown true promise in early clinical studies due to reported activity in BRCA-associated cancers. PARP inhibitors may represent a potentially important new class of chemotherapeutic agents directed at targeting cancers with defective DNA-damage repair. In order to widen the prospective patient population that would benefit from PARP inhibitors, predictive biomarkers based on a clear understanding of the mechanism of action are required. In addition, a more sophisticated understanding of the toxicity profile is required if PARP inhibitors are to be employed in the curative, rather than the palliative, setting. PARP inhibitors have successfully moved into clinical practice in the past few years, with approval granted from the Food and Drug Administration (FDA) and European Medicines Agency (EMA) within the past two years. The United States FDA approval of olaparib applies to fourth-line treatment in germline BRCA-mutant ovarian cancer, and European EMA approval of olaparib for maintenance therapy in both germline and somatic BRCA-mutant platinum-sensitive ovarian cancer. This review covers the current understanding of PARP, its inhibition, and the basis of the excitement surrounding these new agents. It also evaluates future approaches and directions required to achieve full understanding of the intricate interplay of these agents at the cellular level.

This report defines the identity of a calcium-regulated membrane guanylate cyclase transduction system in the cilia of olfactory sensory neurons, which is the site of odorant transduction. The membrane fraction of the neuroepithelial layer of the rat exhibited Ca(2+)-dependent guanylate cyclase activity, which was eliminated by the addition of EGTA. This indicated that the cyclase did not represent a rod outer segment guanylate cyclase (ROS-GC), which is inhibited by free Ca(2+). This interpretation was supported by studies with the Ca(2+) binding proteins, GCAPs (guanylate cyclase activating proteins), which stimulate photoreceptor ROS-GC in the absence of Ca(2+). They did not stimulate the olfactory neuroepithelial membrane guanylate cyclase. The olfactory neuroepithelium contained a Ca(2+) binding protein, neurocalcin, which stimulated the cyclase in a Ca(2+)-dependent fashion. The cyclase was cloned from the neuroepithelium and was found to be identical in structure to that of the previously cloned cyclase termed GC-D. The cyclase was expressed in a heterologous cell system, and was reconstituted with its Ca(2+)-dependent activity in the presence of recombinant neurocalcin. The reconstituted cyclase mimicked the native enzyme. Immunocytochemical studies showed that the guanylate cyclase coexists with neurocalcin in the apical region of the cilia. Deletion analysis showed that the neurocalcin-regulated domain resides at the C-terminal region of the cyclase. The findings establish the biochemical, molecular, and functional identity of a novel Ca(2+)-dependent membrane guanylate cyclase transduction system in the cilia of the olfactory epithelium, suggesting a mechanism of the olfactory neuroepithelial guanylate cyclase regulation fundamentally distinct from the phototransduction-linked ROS-GC.

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Rod outer segment membrane guanylate cyclase1 (ROS-GC1) is the original member of the membrane guanylate cyclase subfamily whose distinctive feature is that it transduces diverse intracellularly generated Ca 2+ signals in the sensory neurons. In the vertebrate retinal neurons, ROS-GC1 is pivotal for the operations of phototransduction and, most likely, of the synaptic activity. The phototransduction-and the synapse-linked domains are separate, and they are located in the intracellular region of ROS-GC1. These domains sense Ca 2+ signals via Ca 2+ -binding proteins. These proteins are ROS-GC activating proteins, GCAPs. GCAPs control ROS-GC1 activity through two opposing regulatory modes. In one mode, at nanomolar concentrations of Ca 2+ , the GCAPs activate the cyclase and as the Ca 2+ concentrations rise, the cyclase is progressively inhibited. This mode operates in phototransduction via two GCAPs: 1 and 2. The second mode occurs at micromolar concentrations of Ca 2+ via S100 . Here, the rise of Ca 2+ concentrations progressively stimulates the enzyme. This mode is linked with the retinal synaptic activity. In both modes, the final step in Ca 2+ signal transduction involves ROS-GC dimerization, which causes the cyclase activation. The identity of the dimerization domain is not known. A heterozygous, triple mutation -E786D, R787C, T788M-in ROS-GC1 has been connected with autosomal cone-rod dystrophy in a British family. The present study shows the biochemical consequences of this mutation on the phototransduction-and the synapse-linked components of the cyclase. (1) It severely damages the intrinsic cyclase activity. (2) It significantly raises the GCAP1-and GCAP2-dependent maximal velocity of the cyclase, but this compensation, however, is not sufficient to override the basal cyclase activity. (3) It converts the cyclase into a form that only marginally responds to S100 . The mutant produces insufficient amounts of the cyclic GMP needed to drive the machinery of phototransduction and of the retinal synapse at an optimum level. The underlying cause of the breakdown of both types of machinery is that, in contrast to the native ROS-GC1, the mutant cyclase is unable to change from its monomeric to the dimeric form, the form required for the functional integrity of the enzyme. The study defines the CORD in molecular terms, at a most basic level identifies a region that is critical in its dimer formation, and, thus, discloses a single unifying mechanistic theme underlying the complex pathology of the disease.Phototransduction is a biochemical process by which the vertebrate rods and cones generate electrical signals in response to captured photons (reviewed in refs 1 and 2). This process occurs in the rod (or cone) outer segments (ROS). 1The photon induces a decline in the level of cyclic GMP and closure of the cyclic GMP-gated cation channels. This results in a drop of intracellular Ca 2+ from ∼500 nM in darkadapted photoreceptor cell to below 100 nM after illumination. This drop activates ROS guanylate cyc...

BackgroundSecretion of human chorionic gonadotropin, especially its beta subunit by malignant trophoblastic tumors and varieties of tumors of different origin is now well documented; however the role of hCG in tumorogenesis is still unknown.ResultsThis study documents the molecular presence of human chorionic gonadotropin beta subunit in uterine cervix cancer tissues and investigates a novel technique to reduce hCGβ levels based on expression of a modified U1 snRNA as a method to study the hormone's role in biology of human cervical cancer cells cultured in vitro. The property of U1 snRNA to block the accumulation of specific RNA transcript when it binds to its donor sequence within the 3' terminal exon was used. The first 10 nucleotides of the human U1 snRNA gene, which normally binds to the 5'ss in pre-mRNA were replaced by a sequence complementary to a 10-nt segment in the terminal exon of the hCGβ mRNA. Three different 5' end-mutated U1 snRNA expression plasmids were tested, each targeting a different sequence in the hCGβ mRNA, and we found each one blocked the expression of hCGβ in HeLa cells, a cervix carcinoma cell line, as shown by immunohistochemistry and qRT-PCR. Reduction of hCGβ levels resulted in a significantly increased apoptosis rate with almost 90% of cells transfected with modified anti-hCGβ U1 snRNAs showing morphological changes characteristic of the apoptotic process.ConclusionThese data suggest that human chorionic gonadotropin beta subunit may act as a tumor growth-stimulating factor.