Regulation of Polarized Protein Transport to Axons, Dendrites, and Sensory Cilia in Caeborhabditis Elegans Neurons

Neurons are highly polarized cells, capable of receiving, processing and transmitting information, with the help of their specialized domains, an axon and one or more dendrites. The molecular dissimilarities between these domains are critical for neuronal function, and are a result of asymmetric trafficking of a large number of cellular components including ion channels, neurotransmitter receptors, synaptic vesicles, and signaling proteins. Yet, despite the significance of asymmetric protein transport in neuronal polarity, the molecules and mechanisms that direct polarized transport to axons, dendrites and cilia in neurons are only partly understood. In this thesis, I describe my effort at understanding how neuronal proteins are asymmetrically localized. I pursued a genetic approach, employing the C. elegans nervous system as an in vivo model system for protein transport to axons, dendrites and cilia. Having established a system to visualize axon-dendrite compartmentalization in PVD mechanosensory neurons, I identified the microtubule-binding protein UNC- 33/CRMP and the ankyrin homolog UNC-44 as major regulators of polarized protein transport in C. elegans neurons. In both unc-33 and unc-44 mutants, axonal proteins are distributed randomly between axons and dendrites, and dendritic proteins are partly mislocalized to axons. In both mutants, the axonal kinesin UNC-104/KIF1A actively misdelivers axonal proteins to both axons and dendrites. An altered distribution of neuronal microtubules in unc-33 and unc-44 mutants suggests that a primary defect in microtubule organization underlies defective protein targeting. unc-44 is required for UNC-33 localization to axons, where its enrichment in a proximal axonal segment suggests analogies with the vertebrate axon initial segment. In parallel experiments, I analyzed odr-8 mutants, which were previously identified in screens for chemotaxis-defective mutants and shown to affect GPCR localization. odr-8 mutants fail to localize a subset of odorant receptors including the ODR-10 diacetyl receptor to sensory cilia. I found that odr-8 encodes the C. elegans homolog of mammalian UfSP2, which acts as a cysteine-protease specific for UFM1, a ubiquitin-like molecule. ODR-10::GFP is retained in the ER in odr-8 mutants, whereas cilia localization of ODR-10::GFP is enhanced in ufm-1 mutants. ufm-1 function is required for ER retention of ODR-10::GFP in odr-8 animals. Thus, ODR-8 and UFM-1 may act antagonistically to regulate ER exit and cilia localization of chemoreceptors such as ODR-10

Safety of long-term denosumab therapy: results from the open label extension phase of two phase 3 studies in patients with metastatic breast and prostate cancer

Purpose: Zoledronic acid (ZA) or denosumab treatment reduces skeletal-related events; however, the safety of prolonged therapy has not been adequately studied. Here, we describe safety results of extended denosumab therapy in patients with bone metastases from the open-label extension phase of two phase 3 trials. Methods: Patients with metastatic breast or prostate cancer received subcutaneous denosumab 120 mg Q4W or intravenous ZA 4 mg Q4W in a double-blinded fashion. Denosumab demonstrated superior efficacy in the blinded treatment phase; thus, patients were offered open-label denosumab for up to an additional 2 years. Results: Cumulative median (Q1, Q3) denosumab exposure was 19.1 (9.2, 32.2) months in the breast cancer trial (n = 1019) and 12.0 (5.6, 21.3) months in the prostate cancer trial (n = 942); 295 patients received denosumab for >3 years. No new safety signals were identified during the open-label phase, or among patients who switched from ZA to denosumab. During the blinded treatment phase, exposure-adjusted subject incidences of osteonecrosis of the jaw (ONJ) were 49 (1.9 %) and 31 (1.2 %) in the denosumab and ZA groups, respectively. In total, 32 (6.9 %) and 25 (5.5 %) new cases of ONJ (not adjusted for exposure) were reported for patients continuing and switching to denosumab, respectively. The incidences of hypocalcemia were 4.3 and 3.1 %, in patients continuing and switching to denosumab, respectively. Conclusion: These results describe the safety profile of denosumab after long-term exposure, or after switching to denosumab from ZA. No new safety signals were identified. Hypocalcemia rates were similar in the blinded treatment and open-label phases. ONJ rates increased with increasing exposure to antiresorptives, consistent with previous reports