544.6 - Emerging Microbiome Genetic Engineering Technology for Stable Levodopa Delivery in Parkinson’s Disease

Piyush Padhi (University of Georgia), Ahmed Abdalla (Iowa State University), Nick Backes (Iowa State University), Gary Zenitsky (University of Georgia), Huajun Jin (University of Georgia), Vellareddy Anantharam (University of Georgia), Arthi Kanthasamy (University of Georgia), Gregory Phillips (Iowa State University), Anumantha Kanthasamy (University of Georgia)

Dopamine replacement in the form of a levodopa/carbidopa tablet has been the gold-standard therapy for Parkinson’s disease (PD) for over six decades. Oral tablet dosing of L-DOPA 3-4 times/day is quite efficacious in controlling early-stage PD symptoms ). However, beyond its ~5-yr clinical honeymoon period, this treatment becomes increasingly associated with L-DOPA-induced dyskinesia (LID) and ‘ON-OFF’ motor complications resulting from the non-continuous, pulsatile delivery of L-DOPA to the brain. Addressing this challenge, we devised an innovative strategy to supplement the host’s microbiota with the human probiotic E.coli Nissle 1917 (EcN) genetically engineered to continuously produce L-DOPA (EcNLDOPA) with no significant side effects. By harnessing emerging synthetic biology and genetic engineering technologies, we successfully established four generations of EcNLDOPA(1-4) and characterized their ability to endogenously synthesize L-DOPA from tyrosine using the recombinant hpaB/C gene. The initially designed plasmid-based EcNLDOPA1,2 systems were further improved by developing the chromosome-integrated and regulatable systems EcN3-4L-DOPA using advanced Sce-ROPE genetic engineering. The steady-state levels of plasma and brain L-DOPA, dopamine and its metabolites, as well as gut colonization of EcNLDOPA, were assessed over time. Our results show that all EcNLDOPA strains effectively generate L-DOPA in a dose- and time-dependent manner in vitro, indicating that the biotherapeutic dosage can be personalized to each patient’s needs. Our mouse pharmacokinetic studies revealed that steady-state levels of plasma L-DOPA and brain dopamine can be achieved by oral administration of EcNLDOPA. Importantly, our pharmacodynamic efficacy studies of the L-DOPA response in the progressive neurodegenerative MitoPark mouse model of PD showed improved performance in motor, cognitive and mood-related tasks. Colonization studies show that our EcNLDOPA biotherapeutic requires only once or twice daily dosing. Our novel re-engineered, self-replicating EcNLDOPA microbial bio-therapeutics offers an entirely new L-DOPA treatment paradigm capable of ensuring long-term efficacy while minimizing side-effects

NIH R61NS112441



Genetically engineered Escherichia Nissle 1917 produces Levodopa (LDOPA) in a regulatable and consistent manner and capable of ameliorating Parkinson's disease (PD) motor and non-motor symptoms in PD animal model.