Safety of Direct Administration of AAV2_CUhCLN2, a Candidate Treatment for the Central Nervous System Manifestations of Late Infantile Neuronal Ceroid Lipofuscinosis, to the Brain of Rats and Nonhuman Primates

Abstract

Late infantile neuronal ceroid lipofuscinosis (LINCL), a pediatric autosomal recessive neurodegenerative lysosomal storage disorder, results from mutations in the CLN2 gene and consequent deficiency in tripeptidyl-peptidase I (TPP-I) and progressive destruction of neurons. We have previously demonstrated that CNS gene transfer of AAV2(CU)hCLN2 (an AAV2-based vector expressing the human CLN2 cDNA) in rats and nonhuman primates mediates long-term TPP-I expression in the CNS neurons [Sondhi, D., Peterson, D.A., Giannaris, E.L., Sanders, C.T., Mendez, B.S., De, B., Rostkowski, A., Blancard, B., Bjugstad, K., Sladek, J.R., Redmond, D.E., Leopold, P.L., Kaminsky, S.M., Hackett, N.R., and Crystal, R.G. (2005). Gene Ther. 12, 1618-1632]. The present study tests the hypothesis that direct CNS administration of a clinical-grade AAV2(CU)hCLN2 vector to the CNS of rats and nonhuman primates at doses scalable to humans has a long-term safety profile acceptable for initiating clinical trials. Fischer 344 rats were injected bilaterally via the striatum with 2 x 10(10) particle units (PU) of AAV2(CU)hCLN2, using saline as a control. At 13, 26, and 52 weeks, vector and phosphate-buffered salineinjected rats were killed (n = 6 per time point), and blood, brain, and distant organs were assessed. There were no biologically significant differences between control and vector groups for complete blood count, serum chemistry, and neutralizing anti-AAV2 antibody levels. CNS administration of AAV2 CUhCLN2 did not result in any pathological changes in the brain that were attributable to the vector, although microscopic changes were observed along the track consistent with needle trauma. A total dose of 3.6 x 10(10) or 3.6 x 10(11) PU of AAV2(CU)hCLN2 was administered to the CNS of African Green monkeys at 12 locations, targeting the caudate nucleus, hippocampus, and overlying cortices. Monkeys (n = 3 at each dose) were killed 1, 13, 26, or 52 weeks after injection. Controls included sham-injected, saline-injected, and AAV2(CU)Null-injected (3.6 x 10(11) PU) monkeys. There were no biologically significant differences among vector-injected and control groups in any parameter of the general assessment, complete blood count, or serum chemistry assessed at multiple time points after vector administration. Importantly, no abnormal behavior was observed in any group in videotaped neurological assessment, where behaviors were quantified before administration and at multiple time points afterward. Histopathological examination of the CNS demonstrated that 1 week after administration, AAV2(CU)hCLN2 produced transient minor white matter edema with reactive glial cells in the corona radiata of the cerebrum along the injection track and in the surrounding white matter. This abnormality was not observed at 13, 26, or 52 weeks. Together with the long-term gene expression after gene transfer, these findings supported the initiation of clinical trials to assess the safety of AAV2(CU)hCLN2 administration to individuals with LINCL.

Survival advantage of neonatal CNS gene transfer for late infantile neuronal ceroid lipofuscinosis

“…Studies of experimental animal models and humans have shown that a significant challenge to providing effective gene therapy for the CNS manifestations of the lysosomal storage disorders is that most of these disorders diffusely affect the CNS, and thus effective therapy requires delivery of the product of the therapeutic vector throughout the brain (Beck, 2007;Sondhi et al, 2001). This challenge is further compounded by the physical limitations of administration of gene therapy vectors to the human CNS, including the number of burr holes that can be placed in the skull, the volume that can be administered per min, and the total time of anesthesia that can be used safely (Crystal et al, 2004;Hackett et al, 2005). Finally, corrective gene therapies for neurodegenerative diseases are not restorative but rather halt disease progression, making delivery prior to disease onset an important objective.…”

“…Finally, corrective gene therapies for neurodegenerative diseases are not restorative but rather halt disease progression, making delivery prior to disease onset an important objective. With these limitations in mind, three basic strategies have been employed to achieve therapeutic levels of gene expression throughout the brain: identifying gene transfer vectors that will diffuse through the CNS more effectively (Cearley and Wolfe2006;Deglon and Hantraye2005;Shevtsova et al, 2005;Sondhi et al, 2007), optimizing the method of delivery and volume and rate of infusion (Chen et al, 1999;Hackett et al, 2005;Hsich et al, 2002), and administration at an earlier time point, when the brain is smaller and immature (Bostick et al, 2007;Broekman et al, 2007;Daly et al, 1999;Griffey et al, 2006;Li and Daly2002;Passini et al, 2003;Passini and Wolfe2001;Rafi et al, 2005;Shen et al, 2001;Waddington et al, 2004). Studies from our laboratory and others have identified several serotypes of adeno-associated virus (AAV) vectors that are more effective than the first generation vectors used for CNS gene transfer (Broekman et al, 2006;Burger et al, 2005;Cearley and Wolfe2006;Harding et al, 2006;Sondhi et al, 2007;Taymans et al, 2007), the parameters for optimal administration of vectors to the CNS have been established (Chen et al, 1999;Hackett et al, 2005;Hsich et al, 2002) and several studies have shown that administration earlier in life results in an advantage over therapy in older animals (Bostick et al, 2007;Broekman et al, 2007;Daly et al, 1999;Griffey et al, 2006;Li and Daly2002;Passini et al, 2003;Passini and Wolfe2001...…”

Residual levels of tripeptidyl-peptidase I activity dramatically ameliorate disease in late-infantile neuronal ceroid lipofuscinosis

“…Initial studies examining the expression of TPPI by recombinant AAV and other viral vectors demonstrated high levels and widespread expression of the protein throughout the central nervous system (CNS) of healthy rodents [7,8]. More recently, AAVmediated gene therapy has been evaluated in a mouse model for LINCL.…”

When Ethics Constrains Clinical Research: Trial Design of Control Arms in “Greater Than Minimal Risk” Pediatric Trials

“…Preclinical and clinical studies have shown the feasibility and initial promise of using an adeno-associated virus (AAV) to transfer the human CLN2 cDNA to the brains of children with LINCL (Crystal et al, 2004;Hackett et al, 2005;Sondhi et al, 2005Sondhi et al, , 2007Passini et al, 2006;Worgall et al, 2008;Souwedaine et al, 2010). Preclinical studies with rodents showed that administration of the human CLN2 cDNA by means of an AAV2-based vector (AAV2 hCLN2) was associated with a decrease in the abnormal accumulation of storage material in CNS neurons (Hackett et al, 2005;Sondhi et al, 2005;Passini et al, 2006).…”

“…Preclinical studies with rodents showed that administration of the human CLN2 cDNA by means of an AAV2-based vector (AAV2 hCLN2) was associated with a decrease in the abnormal accumulation of storage material in CNS neurons (Hackett et al, 2005;Sondhi et al, 2005;Passini et al, 2006). This was followed by a clinical study in which the AAV2hCLN2 vector was administered directly to the CNS of 10 children with LINCL of severe or moderate severity in a neurosurgical procedure involving six burr holes, catheter insertion, and infusion of the vector in a total of 12 sites over several hours (Worgall et al, 2008;Souweidane et al, 2010).…”