Jeff Bronstein, M.D., Ph.D
University of California, Los Angeles

Jeff Bronstein received his bachelor’s degree from the University of California, Berkeley and M.D. and Ph.D. from UCLA as a recipient of the Medical Scientist Training Program Award.   He completed a residency in Neurology and fellowship training in Movement Disorders at UCLA and at Queens Square in London.  Dr. Bronstein also completed a postdoctoral fellowship in molecular biology before being appointed an Assistant Professor of Neurology in 1994, Director of the Movement Disorders Program at UCLA in 1996. Professor of Neurology in 2006, and Professor of Molecular Toxicology in 2007.  His clinical interests include the management of Parkinson’s disease (PD) and other movement disorders, surgical treatment of PD, and developing new therapies for patients. Dr. Bronstein’s research interests include the study of the causes of PD (environmental and genetic) using cell and zebrafish models as well as population-based studies.  His research is supported by the NIH, Veteran Administration, and private foundations. Dr. Bronstein is the Principle Investigator of one of 6 National Parkinson’s Disease Centers (PADRECC) at the Veterans Administration Medical Center.

CLR01 and Parkinson’s disease

We have been studying the potential of CLR01 to slow or stop the progression of Parkinson’s disease (PD).   Since aggregation of α-synuclein appears to be central to the pathogenesis of PD, we have focused on CLR01 effects on α-synuclein toxicity. In collaboration with the Bitan group, we first studied tested CLR01 in an immortalized cell line (HEK) that conditionally expressed α-synuclein and found that it completely blocked α-synuclein toxicity. CLR01 also protected PC-12 cells against exogenously applied α-synuclein (Figure 1) (Prabhudesai et al., Neurotherapeutics, 2012).
Figure 1. CLR01 inhibits α-syn toxicity in cell culture. a)CLR01 inhibits endogenously expressed α-syn toxicity in HEK293 cells. α-Syn expression was induced by adding doxyclycline (Dox) in the absence or presence of CLR01. Cell numbers and cell death measured using propidium iodide (PI) were determined by flow cytometry. N = 12 per condition. *p < 0.0003, **p < 0.007. b) Differentiated PC-12 cells were treated with 20 μM α-syn incubated in the absence or presence of increasing concentrations of CLR01 for 48 h and cell viability was measured using the MTT assay. Inset: Viability of PC-12 cells treated similarly with α-syn in the presence or absence of 10-fold molar excess of CLR01 or CLR03. The data are an average of at least 3 independent experiments with 6 wells per condition.

CLR01 was then tested in two novel zebrafish models of Parkinson’s pathology.   In one of our models, human α-synuclein was overexpressed in neurons, which led to neuronal death and poor survival of the fish. CLR01 markedly attenuated α-synuclein toxicity (Figure 2) (Prabhudesai et al., Neurotherapeutics, 2012).

In a second model, we tested CLR01’s ability to protect against pesticide-induced toxicity (Lulla et al. Environ Health Perpect. 2016). Exposure to pesticides is known to be associated with an increased risk of developing PD and we recently found that chronic exposure to the fungicide Ziram is associated with a 3-fold increased risk of developing PD. Additional studies revealed that ziram and other pesticides in this class inhibit the ubiquitin proteasome system (UPS) resulting in increased α-synuclein in dopaminergic cells.  Zebrafish embryos exposed to low concentrations of ziram (50 nM) showed abnormal swimming and a significant loss of dopaminergic neurons.  This dopaminergic toxicity was markedly attenuated when we knocked down zebrafish synuclein with morpholinos suggesting that ziram-induced toxicity was at least partially synuclein-dependent. Treatment of the zebrafish with CLR01 also dramatically reduced ziram-induced dopaminergic neuron toxicity, showing that inhibiting the self-association of synuclein into toxic aggregates was sufficient to prevent its toxicity.

Taken together, these studies suggest that CLR01 is highly effective in protecting neurons in cells and zebrafish that model both genetic and environmental causes of PD.
Figure 2. CLR01 ameliorates α-syn neurotoxicity in ZF. a)ZF embryos were treated with CLR01 at 8 hpf and were monitored for abnormal appearance and survival. Bright-field and fluorescent overlay images were taken at 72 hpf (top). Green bars represent normal appearing embryos and red bars represent abnormal embryos (N = 132/condition). b) CLR01 prevents α-syn-induced apoptosis. ZF embryos expressing DsRed or α-syn were incubated in acridine orange 24 hpf and apoptotic cells were counted (N = 6 per condition). Ten μM CLR01 reduced α-syn-induced apoptosis to control levels (*p < 0.007 Syn-DsRed vs Syn-DsRed/CLr01 and DsRed control). Representative images on shown on the right.