Parkinson's Disease Gene Therapy: Success by Design Meets Failure by Efficacy
Authors: Raymond T Bartus, Marc S Weinberg, R. Jude Samulski
Journal: Molecular Therapy (2013)

Over the past decade, nine gene therapy clinical trials for Parkinson's disease (PD) have been initiated and completed. Starting with considerable optimism at the initiation of each trial, none of the programs has yet borne sufficiently robust clinical efficacy or found a clear path toward regulatory approval. Despite the immediately disappointing nature of the efficacy outcomes in these trials, the clinical data garnered from the individual studies nonetheless represent tangible and significant progress for the gene therapy field. Collectively, the clinical trials demonstrate that we have overcome the major safety hurdles previously suppressing central nervous system (CNS) gene therapy, for none produced any evidence of untoward risk or harm after administration of various vector-delivery systems. More importantly, these studies also demonstrated controlled, highly persistent generation of biologically active proteins targeted to structures deep in the human brain. Therefore, a renewed, focused emphasis must be placed on advancing clinical efficacy by improving clinical trial design, patient selection and outcome measures, developing more predictive animal models to support clinical testing, carefully performing retrospective analyses, and most importantly moving forward—beyond our past limits. Over the past decade, nine gene therapy clinical trials for Parkinson's disease (PD) have been initiated and completed. Starting with considerable optimism at the initiation of each trial, none of the programs has yet borne sufficiently robust clinical efficacy or found a clear path toward regulatory approval. Despite the immediately disappointing nature of the efficacy outcomes in these trials, the clinical data garnered from the individual studies nonetheless represent tangible and significant progress for the gene therapy field. Collectively, the clinical trials demonstrate that we have overcome the major safety hurdles previously suppressing central nervous system (CNS) gene therapy, for none produced any evidence of untoward risk or harm after administration of various vector-delivery systems. More importantly, these studies also demonstrated controlled, highly persistent generation of biologically active proteins targeted to structures deep in the human brain. Therefore, a renewed, focused emphasis must be placed on advancing clinical efficacy by improving clinical trial design, patient selection and outcome measures, developing more predictive animal models to support clinical testing, carefully performing retrospective analyses, and most importantly moving forward—beyond our past limits. Parkinson's disease (PD) is a chronic, progressive neurodegenerative disease most widely recognized for the profound degeneration of mid-brain dopamine nigrostriatal neurons linked to serious motor symptoms.1Olanow CW Stern MB Sethi K The scientific and clinical basis for the treatment of Parkinson disease (2009).Neurology. 2009; 72: S1-136Crossref PubMed Scopus (664) Google Scholar However, PD is far more complex than commonly appreciated, with multiple etiologic variables and pathogenic pathways, complex pathologies, and a wide range of central nervous system (CNS) and non-CNS symptoms (Table 1).2Bonnet AM Jutras MF Czernecki V Corvol JC Vidailhet M Nonmotor symptoms in Parkinson's disease in 2012: relevant clinical aspects.Parkinsons Dis. 2012; 2012: 198316PubMed Google Scholar,3Braak H Del Tredici K Rüb U de Vos RA Jansen Steur EN Braak E Staging of brain pathology related to sporadic Parkinson's disease.Neurobiol Aging. 2003; 24: 197-211Abstract Full Text Full Text PDF PubMed Scopus (7140) Google Scholar Moreover, wide gaps in our understanding still exist at each disease level (i.e., etiology, pathogenesis, pathology, symptoms), and the cause-effect relationships between them remain especially obscure. Arguably, the most well-characterized relationship exists with regard to nigrostriatal degeneration linked to the key motor symptoms; currrent oral dopaminergic pharmaceuticals are effective in controlling these symptoms at early disease stages. However, the drugs' effectiveness decline with progressive pathology, leading to gradual incapacitation of patients by increased "off" time (i.e., periods of no symptomatic relief) and increasing side effects such as peak-dose dyskinesias.1Olanow CW Stern MB Sethi K The scientific and clinical basis for the treatment of Parkinson disease (2009).Neurology. 2009; 72: S1-136Crossref PubMed Scopus (664) Google Scholar Thus, adequate treatment of the nigrostriatal-mediated motor impairments continues to represent a significant unmet medical need, affecting over 4 million people worldwide.4Parkinson's Disease Foundation (2013). Statistics on Parkinson's. http://www.pdf.org/en/parkinson_statisticsGoogle Scholar Though a number of solutions have been conceived to improve the function of the degenerating dopaminergic system, translating these biopharmaceutical concepts to the clinic has been challenging due to obstacles associated with delivering macromolecules to the central nervous system in a persistent and targeted fashion.Table 1Complex etiology, pathogenesis, pathology, and symptomatology of Parkinson's diseaseEtiology Pathogenesis Pathology SymptomsGenetic mutationsaProteins encoded by mutated genes: (SNCAbα-synuclein;, PINK1cPTEN-induced putative kinase 1;, LRRK2dleucine-rich repeat kinase 2;, PRKNeparkin;, DJ1 -or PARK7fPark-7 (peptidase C56 protein family)., ATP13A2)Mitochrondrial dysfunction Oxidative damageLoss of function and gradual axonopathy/death of mid-brain dopamine neuronsLoss of voluntary motor function (bradykinesia, rigidity, tremors, and eventually postural instability)α-synuclein misfolding (& possible prion-like reactions)α-synuclein accumulation (Lewy bodies)ApoptosisTau phosphorylationEnvironmental factors (e.g.,age, pesticides, herbicides, head trauma, fertilizers)NeuroinflammationLoss of function and gradual death of neurons in several other PNS and CNS neuronal systemsCognitive deficits; psychiatric issues; olfactory deficits; REM sleep disturbances, autonomic dysfunction (e.g., urinary/GI complications)CNS, central nervous system; GI, gastrointestinal; PNS, peripheral nervous system; REM, rapid eye movement.a Proteins encoded by mutated genes:b α-synuclein;c PTEN-induced putative kinase 1;d leucine-rich repeat kinase 2;e parkin;f Park-7 (peptidase C56 protein family). Open table in a new tab CNS, central nervous system; GI, gastrointestinal; PNS, peripheral nervous system; REM, rapid eye movement. Progress achieved in the realm of gene therapy (GT) over the past decade has offered solutions to many of the delivery constraints,5Bartus RT Translating the therapeutic potential of neurotrophic factors to clinical 'proof of concept': a personal saga achieving a career-long quest.Neurobiol Dis. 2012; 48: 153-178Crossref PubMed Scopus (22) Google Scholar and several aspects of PD present it overtly as an ideal clinical indication to target using GT: (i) the well-defined, localizable, and targetable neuronal systems involved with major motor symptoms, (ii) the need for relatively small titer and volume of vector targeted to those sites, which avoids the systemic circulation of immunogenic materials, and (iii) the large and increasing demand for improved therapeutics with an aging population,4Parkinson's Disease Foundation (2013). Statistics on Parkinson's. http://www.pdf.org/en/parkinson_statisticsGoogle Scholar which in whole bolsters impact and financial support for research and development. Given this rationale, PD has, for better or worse, become a key exemplar for CNS GT. To date, the results of completed PD GT trials have supported the safety of GT targeting in the brain and many have further confirmed the successfully targeted expression of bioactive proteins in specific brain sites. However, none of the programs has yet produced sufficiently robust or reliable efficacy data to enable initiation of a pivotal phase 3 trial required for regulatory approval. We attempt to integrate the many successes and formidable challenges of GT treatment of PD, in an effort to seek the best path forward for PD and CNS GT as a whole. As compared with conventional small molecule drug testing, GT in the CNS is limited with respect to establishing initial dosing, quantifying targeting success, and accessing a comprehensive gauge of transgene production and localization, all of which provide the basis for iterative improvement with traditional drug development. While we expect that such limitations are attributable to a short-lasting gap between demand and current technological capability, the ramifications of such limitations include poor predictive power and a loss of continuity and progress between successive trials. Whereas continuing progress is being made in the visualization of viral vectors as they are delivered to the brain (e.g., the use of gadolinium-containing liposomes coupled with magnetic resonance imaging),6Szerlip NJ Walbridge S Yang L Morrison PF Degen JW Jarrell ST et al.Real-time imaging of convection-enhanced delivery of viruses and virus-sized particles.J Neurosurg. 2007; 107: 560-567Crossref PubMed Scopus (85) Google Scholar,7Fiandaca MS Varenika V Eberling J McKnight T Bringas J Pivirotto P et al.Real-time MR imaging of adeno-associated viral vector delivery to the primate brain.Neuroimage. 2009; 47: T27-T35Crossref PubMed Scopus (92) Google Scholar there is currently no direct means to determine the distribution or magnitude of transgene expression, since protein expression with CNS GT trials is both generated and produced entirely within the confines of the human brain. Therefore, it is not yet possible to confirm proper dose selection or monitor dosing levels, as would normally be done using standard pharmacokinetic (i.e., plasma level) methods. This severely limits the predictive and correlative strength of treatment endpoint measurements. One surrogate means by which transgene expression and function has been assessed is positron emission tomography (PET) imaging coupled with radiolabeled L-Dopa.8Bankiewicz KS Eberling JL Kohutnicka M Jagust W Pivirotto P Bringas J et al.Convection-enhanced delivery of AAV vector in parkinsonian monkeys; in vivo detection of gene expression and restoration of dopaminergic function using pro-drug approach.Exp Neurol. 2000; 164: 2-14Crossref PubMed Scopus (362) Google Scholar This approach has enabled the live imaging of the dopamine precursor's uptake, which when compared to pretreatment levels can be directly informative to aromatic L-amino acid decarboxylase (AADC) and tyrosine hydroxylase (TH) transgene production and function. One problem with this approach is the inadequacy of using such technology as a dosing criterion: a sufficient signal is not expected until weeks or months after vector is administered, making adjustments to dosing inefficient at best, even within the flexibility of a phase 1 trial. Due to the relative blindness of the researcher/clinician to targeting and dosing data, the process of translating and validating clinical results from prior animal data in GT programs has been characterized a "look-see" or trial and error approach. While this approach can still enable iterative improvement and therapeutic success, as demonstrated by the continued improvements to hemophilia GT treatment (discussed further below), it is not a paradigm easily applied to all drug development endeavors. Moreover, the repeated efforts can widen the so-called "valley of death" one confronts when attempting to move a program forward from initial, small open-label trials to the multi-center, double-blind, sham-surgery controlled trials required to establish efficacy. As we will point out in the next section, each of the PD approaches described below offer vivid examples of the look-see approach, often to the program's detriment. To date, five divergent GT approaches have been developed to treat the major motor symptoms of PD, all with the use of AAV or lentivirus vector platforms (Table 2). Each of the clinical approaches has focused on aspects of the basal ganglia, a group of subcortical neural sites located near the base of the forebrain that communicate intimately with the cerebral cortex and other brain areas. Most GT approaches directly target the terminals of the degenerating nigrostriatal neurons for gene delivery, while one approach indirectly attempts to resolve striatal neurochemical imbalance by increasing inhibitory control from the subthalamic nucleus.Table 2Summary of gene therapy clinical programs for Parkinson's diseaseTreatment (approach)Trial designYear beganSubject #Described as a Phase 1/2 trial, this open label (uncontrolled) study does not differ substantially from many dose-escalation Phase 1 safety studies that include secondary efficacy endpoints; thus, the distinction appears to be more a semantic preference than a reflection of a substantial difference in study design. dosedHighest total dose (vg)Target(S)Largest volume (pl)/siteSafety resultsEfficacy outcomesAAV2/GADPh1-uncontrolledaKaplitt et al.15.2003121 × 1012Subthal Nuc (unilat)50AcceptableAdvanced to Ph2Ph2-double-blindbLeWitt et al. 201116.200822/16*Twenty-two subjects were dosed but six were eliminated from efficacy analysis due to mistargeting of cannula.1 × 1012Subthal Nuc (Bilat)35AcceptableMixed results; program suspendedAAV2/AADCPh1-uncontrolledcChristine et al. 22.2004100.3 × 1012Putamen (Bilat)50AcceptableProgram suspended; revised Ph1 recently announcedAAV2/AADCPh1-uncontrolleddMuramatsu et al. 28.200760.3 × 1012Putamen (Bilat)50AcceptableNo further testing; revised Ph1 recently announced by USA groupAAV2/NRTNPh1-uncontrolledeMarks et al. 40.2005120.54 × 1012Putamen (Bilat)5(10)**Two 5 µl volumes infused via single needle tract ~4 mm apart.AcceptableAdvanced to Ph2Ph2A-double-blindfMarks et al. 38.2006380.54 × 1012Putamen (Bilat)5(10)**Two 5 µl volumes infused via single needle tract ~4 mm apart.AcceptableMixed results; revised Ph1 designedPh1-uncontrolledgBartus et al. 39.200962.4 × 1012Put + SN (Bilat)50AcceptableAdvanced to Ph2Ph2B-double-blindhBartus et al. 37. Palfi et al.91h2010242.4 × 1012Put + SN (Bilat)50AcceptableProgram suspendedLENTI/AADC-TH-CH1Ph1/2-uncontrolled#Described as a Phase 1/2 trial, this open label (uncontrolled) study does not differ substantially from many dose-escalation Phase 1 safety studies that include secondary efficacy endpoints; thus, the distinction appears to be more a semantic preference than a reflection of a substantial difference in study design.200815Lentivirus dosing is not comparable to that of AAV A five-fold dose range was tested involving 3 dose levels (1.9 × 107 transducing units (TU); 4.0 × 107 TU; 1.0 × 108 TU).Putamen (Bilat)AcceptableProgram suspended; additional work to optimize vector ongoingAAV2/GDNFPh1-uncontrolledkLonser90.k2013Ongoing0.7 × 1012Putamen (Bilat)150N/AN/ASynopsisTotal of seven phase 1 and three phase 2 trials2003-2013>139Tested up to 1 × 1012 vg AAVTargets have included subthalamic nucleus, putamen and SN50 pl (most common); 150 pl (largest)No safety issues or serious side effects notedEfficacy outcomes generally disappointingAAV, adeno-associated virus; SN, substantia nigra.a Kaplitt et al.15Kaplitt MG Feigin A Tang C Fitzsimons HL Mattis P Lawlor PA et al.Safety and tolerability of gene therapy with an adeno-associated virus (AAV) borne GAD gene for Parkinson's disease: an open label, phase I trial.Lancet. 2007; 369: 2097-2105Abstract Full Text Full Text PDF PubMed Scopus (838) Google Scholar.b LeWitt et al. 201116LeWitt PA Rezai AR Leehey MA Ojemann SG Flaherty AW Eskandar EN et al.AAV2-GAD gene therapy for advanced Parkinson's disease: a double-blind, sham-surgery controlled, randomised trial.Lancet Neurol. 2011; 10: 309-319Abstract Full Text Full Text PDF PubMed Scopus (469) Google Scholar.c Christine et al. 22Christine CW Starr PA Larson PS Eberling JL Jagust WJ Hawkins RA et al.Safety and tolerability of putaminal AADC gene therapy for Parkinson disease.Neurology. 2009; 73: 1662-1669Crossref PubMed Scopus (330) Google Scholar.d Muramatsu et al. 28Muramatsu S Fujimoto K Kato S Mizukami H Asari S Ikeguchi K et al.A phase I study of aromatic L-amino acid decarboxylase gene therapy for Parkinson's disease.Mol Ther. 2010; 18: 1731-1735Abstract Full Text Full Text PDF PubMed Scopus (258) Google Scholar.e Marks et al. 40Marks Jr, WJ Ostrem JL Verhagen L Starr PA Larson PS Bakay RA et al.Safety and tolerability of intraputaminal delivery of CERE-120 (adeno-associated virus serotype 2-neurturin) to patients with idiopathic Parkinson's disease: an open-label, phase I trial.Lancet Neurol. 2008; 7: 400-408Abstract Full Text Full Text PDF PubMed Scopus (477) Google Scholar.f Marks et al. 38Marks Jr, WJ Bartus RT Siffert J Davis CS Lozano A Boulis N et al.Gene delivery of AAV2-neurturin for Parkinson's disease: a double-blind, randomised, controlled trial.Lancet Neurol. 2010; 9: 1164-1172Abstract Full Text Full Text PDF PubMed Scopus (502) Google Scholar.g Bartus et al. 39R Bartus CERE-120 (AAV-neurturin) for the Treatment of Parkinson's Disease: Experience from 4 Clinical Trials and Human Autopsy Data. 2013 American Society of Gene and Cell Therapy 16th Annual Meeting: Salt Palace Convention Center in Salt Lake City, Utah, USAGoogle Scholar.h Bartus et al. 37Bartus R Baumann TL Siffert J Herzog CD Alterman R Boulis N et al.Safety/feasibility of targeting the substantia nigra with AAV2-neurturin in Parkinson's patients.Neurology. 2013; 80: 1698-1702Crossref PubMed Scopus (146) Google Scholar. Palfi et al.91Palfi S Gurruchaga JM Ralph GS Lepetit H Lavisse S Buttery PC et al.Long-term safety and tolerability of ProSavin, a lentiviral vector-based gene therapy of Parkinson's disease: a dose escalation, open-label, phase 1/2 trial.The Lancet. 2014; (http://dx.doi.org/10.1016/S0140-6736(13)61939-X)PubMed Google Scholark Lonser90Lonser RR A phase 1 open-label dose escalation safety study of convection-enhanced delivery (CED) of adeno-associated virus encoding glial cell line-derived neurotrophic factor (AAV2-GDNF) in subjects with advanced Parkinson's disease.Recombinant DNA Advisory Committee Protocol. 2009; : 0901-0962Google Scholar.* Twenty-two subjects were dosed but six were eliminated from efficacy analysis due to mistargeting of cannula.** Two 5 µl volumes infused via single needle tract ~4 mm apart.# Described as a Phase 1/2 trial, this open label (uncontrolled) study does not differ substantially from many dose-escalation Phase 1 safety studies that include secondary efficacy endpoints; thus, the distinction appears to be more a semantic preference than a reflection of a substantial difference in study design.## A five-fold dose range was tested involving 3 dose levels (1.9 × 107 transducing units (TU); 4.0 × 107 TU; 1.0 × 108 TU). Open table in a new tab AAV, adeno-associated virus; SN, substantia nigra. Three of the five PD GT approaches have attempted to ameliorate motor symptoms by altering the neurochemical conductivity of neurons mediating the motor behavior. This general strategy has involved delivering neurotransmitter-producing enzymes. In a first clinical study, AAV2/GAD (adeno-associated viral vector serotype 2/glutamic acid decarboxylase (GABA), a combination of GAD-65 and GAD-67) was delivered to the subthalamic nucleus9Luo J Kaplitt MG Fitzsimons HL Zuzga DS Liu Y Oshinsky ML et al.Subthalamic GAD gene therapy in a Parkinson's disease rat model.Science. 2002; 298: 425-429Crossref PubMed Scopus (287) Google Scholar,10Emborg ME Carbon M Holden JE During MJ Ma Y Tang C et al.Subthalamic glutamic acid decarboxylase gene therapy: changes in motor function and cortical metabolism.J Cereb Blood Flow Metab. 2007; 27: 501-509Crossref PubMed Scopus (110) Google Scholar (Figure 1b). GAD catalyzes the synthesis of gamma-aminobutyric acid, the major inhibitory neurotransmitter in the CNS, potentially providing lost inhibitory control in the basal ganglion motor system, thus restoring appropriate transynaptic balance. A second approach has attempted to improve the ability of the putamen to synthesize dopamine from exogenous L-Dopa (the immediate precursor for dopamine and the mainstay therapeutic for early-stage PD) by expressing the major L-DOPA-converting enzyme AADC in the putamen. Whereas one clinical investigation used AAV to deliver this enzyme,8Bankiewicz KS Eberling JL Kohutnicka M Jagust W Pivirotto P Bringas J et al.Convection-enhanced delivery of AAV vector in parkinsonian monkeys; in vivo detection of gene expression and restoration of dopaminergic function using pro-drug approach.Exp Neurol. 2000; 164: 2-14Crossref PubMed Scopus (362) Google Scholar a second study used a trisictronic (delivery of three transgenes from a single vector cassette) lenti-vector expressing AADC as well as TH and GTP-CH1 (guanosine 5′-triphosphate cyclohydrolase1), with the latter two enzymes also significantly contributing to dopamine's synthesis11Jarraya B Boulet S Ralph GS Jan C Bonvento G Azzouz M et al.Dopamine gene therapy for Parkinson's disease in a nonhuman primate without associated dyskinesia.Sci Transl Med. 2009; 1: 2ra4Crossref PubMed Scopus (152) Google Scholar (Figure 1c). The final two of five PD GT approaches have employed a neurotrophic factor delivery approach, which rather than directly attempting to modulate neuronal activity, instead confer "trophic" support to the dopaminergic and surrounding neuronal populations by inducing repair genes in activated cells12Aron L Klein R Repairing the parkinsonian brain with neurotrophic factors.Trends Neurosci. 2011; 34: 88-100Abstract Full Text Full Text PDF PubMed Scopus (94) Google Scholar,13Peterson AL Nutt JG Treatment of Parkinson's disease with trophic factors.Neurotherapeutics. 2008; 5: 270-280Crossref PubMed Scopus (132) Google Scholar (Figure 1c,e). This offers potential improvement in symptoms as well as delay in disease progression.5Bartus RT Translating the therapeutic potential of neurotrophic factors to clinical 'proof of concept': a personal saga achieving a career-long quest.Neurobiol Dis. 2012; 48: 153-178Crossref PubMed Scopus (22) Google Scholar,14Kells AP Eberling J Su X Pivirotto P Bringas J Hadaczek P et al.Regeneration of the MPTP-lesioned dopaminergic system after convection-enhanced delivery of AAV2-GDNF.J Neurosci. 2010; 30: 9567-9577Crossref PubMed Scopus (102) Google Scholar Both neurotrophic GT investigations to date have used AAV2 as the delivery vector; the studies diverge merely in the choice of which of two structural and functional analog proteins (neurturin (NRTN) or glia cell–derived neurotrophic factor (GDNF)) is packaged and expressed. Below we offer perspective on the outcomes of each of the PD GT clinical approaches to date. Following publication of nonhuman efficacy and safety data10Emborg ME Carbon M Holden JE During MJ Ma Y Tang C et al.Subthalamic glutamic acid decarboxylase gene therapy: changes in motor function and cortical metabolism.J Cereb Blood Flow Metab. 2007; 27: 501-509Crossref PubMed Scopus (110) Google Scholar and the successful completion of the first safety and tolerability trial for PD GT,15Kaplitt MG Feigin A Tang C Fitzsimons HL Mattis P Lawlor PA et al.Safety and tolerability of gene therapy with an adeno-associated virus (AAV) borne GAD gene for Parkinson's disease: an open label, phase I trial.Lancet. 2007; 369: 2097-2105Abstract Full Text Full Text PDF PubMed Scopus (838) Google Scholar a single controlled phase 2 trial with AAV/GAD was initiated. The trial involved 45 total subjects (roughly 50:50, treated:sham) and was completed without serious incident.16LeWitt PA Rezai AR Leehey MA Ojemann SG Flaherty AW Eskandar EN et al.AAV2-GAD gene therapy for advanced Parkinson's disease: a double-blind, sham-surgery controlled, randomised trial.Lancet Neurol. 2011; 10: 309-319Abstract Full Text Full Text PDF PubMed Scopus (469) Google Scholar Several interesting protocol innovations were employed, including prescreening all subjects with 18F-fluorodeoxyglucose PET to help assure a definitive diagnosis of PD. While this investigation is unique in that it achieved statistical significance on the primary efficacy endpoint (i.e., UPDRS motor-off), which represents a significant accomplishment in its own right, the clinical effect was described as "modest".17Coune PG Schneider BL Aebischer P Parkinson's disease: gene therapies.Cold Spring Harb Perspect Med. 2012; 2: a009431Crossref Scopus (80) Google Scholar Indeed, even the significant primary endpoint produced only a 3.4-point difference on the UPDRS, part 3 scale (clinician-scored motor evaluation), compared to sham control. Moreover, the slim statistical difference between groups (P = 0.04) likely positively influenced by the use of a reasonable, protocol-prescribed data analysis approach that would not meet the more rigorous "intent to treat" standards traditionally held by the US Food and Drug Administration (FDA). Specifically, five subjects, four of whom showed no benefit on the primary endpoint, were eliminated after the trial was completed because an magnetic resonance imaging-based review of the targeting identified their injections as being off-target.16LeWitt PA Rezai AR Leehey MA Ojemann SG Flaherty AW Eskandar EN et al.AAV2-GAD gene therapy for advanced Parkinson's disease: a double-blind, sham-surgery controlled, randomised trial.Lancet Neurol. 2011; 10: 309-319Abstract Full Text Full Text PDF PubMed Scopus (469) Google Scholar While two other efficacy measurements (both clinical impressions rating scales) also showed statistical significance (P < 0.05), these effects were also quite modest (i.e., < ½ point difference), while 21 other efficacy endpoints, including those commonly considered important for confirming clinical benefit (e.g., PDQ-39; multiple motor diary scores), showed no difference between groups. Thus, despite technically meeting the primary endpoint, the efficacy data from this trial were modest at best, which likely was responsible for the trial's sponsor (Neurologix, Fort Lee, NJ) discontinuing the AAV/GAD program. In retrospect, the AAV/GAD clinical trial highlighted several key early concerns for the PD GT field to consider: (i) whether modest nonhuman primate efficacy data (e.g., a 1-point improvement versus control on a 32-point behavioral scale) justified moving into clinical testing, (ii) the disappointingly modest efficacy data generated in the clinic, and (iii) the need for such high exclusion rates of treated subjects. The latter concern hints at potential challenges in performing consistent, accurate intracranial targeting—a concern that continues to be addressed and potentially resolved,18Su X Kells AP Huang EJ Lee HS Hadaczek P Beyer J et al.Safety evaluation of AAV2-GDNF gene transfer into the dopaminergic nigrostriatal pathway in aged and parkinsonian rhesus monkeys.Hum Gene Ther. 2009; 20: 1627-1640Crossref PubMed Scopus (99) Google Scholar albeit using real-time imaging instrumentation and methods not yet available in most stereotactic surgical operating rooms. Perhaps more importantly, even the hint of exclusionary data, regardless of the specific circumstances, can mar the perceived integrity of the results, and discount the apparent maturity of the field at large. Here, the modest efficacy data may be best ascribed to modest non-human primate findings. However, as we will examine, such modest clinical efficacy reported in this initial trial foreshadowed that of each subsequent clinical trial, even where preclinical studies showed more robust results. This clinical disappointment, therefore, can best be examined in the broader context of PD GT. Another early PD GT approach utilized AAV/AADC, which after showing preclinical efficacy in MPTP monkeys,8Bankiewicz KS Eberling JL Kohutnicka M Jagust W Pivirotto P Bringas J et al.Convection-enhanced delivery of AAV vector in parkinsonian monkeys; in vivo detection of gene expression and restoration of dopaminergic function using pro-drug approach.Exp Neurol. 2000; 164: 2-14Crossref PubMed Scopus (362) Google Scholar moved into clinical testing for 15 moderately advanced subjects in an open-label phase 1 clinical trial. The protocol for this trial was publicly discussed at the recombinant DNA advisory committee in late 2003,19G Pierce 2003 A phase 1 open-label safety study of intrastriatal infusion of adeno-associated virus encoding human aromatic L-amino acid decarboxylase (AAV-hAADC-2) in subjects with advanced parkinson's disease [AAV-hAADC-2-003]. NIH Recombinant DNA Advisory Committee-Appendix M.Google Scholar but subjects were not treated until 1 year later, during which time Genzyme, Boston, MA acquired the program from Avigen (Alameda, CA).202005 Genzyme Acquires Significant Portion of Avigen's Gene Therapy Technology. 21 December 2005 Press ReleaseGoogle Scholar First, it is worth noting the success of this trial, which like the preceding AAV/GAD study, included preliminary evidence for safety.21Mittermeyer G Christine CW Rosenbluth KH Baker SL Starr P Larson P et al.Long-term evaluation of a phase 1 study of AADC gene therapy for Parkinson's disease.Hum Gene Ther. 2012; 23: 377-381Crossref PubMed Scopus (200) Google Scholar,22Christine CW Starr PA Larson PS Eberling JL Jagust WJ Hawkins RA et al.Safety and tolerability of putaminal AADC gene therapy for Parkinson disease.Neurology. 2009; 73: 1662-1669Crossref PubMed Scopus (330) Google Scholar However, despite a reasonable scientific rationale and several animal studies demonstrating reasonably robust enhancement in nigrostriatal dopamine function with AAV2-AADC,8Bankiewicz KS Eberling JL Kohutnicka M Jagust W Pivirotto P Bringas J et al.Convection-enhanced deliver