TY - JOUR
T1 - Dendrimer-conjugated glutaminase inhibitor selectively targets microglial glutaminase in a mouse model of Rett syndrome
AU - Khoury, Elizabeth Smith
AU - Sharma, Anjali
AU - Ramireddy, Rajasekhar R.
AU - Thomas, Ajit G.
AU - Alt, Jesse
AU - Fowler, Amanda
AU - Rais, Rana
AU - Tsukamoto, Takashi
AU - Blue, Mary E.
AU - Slusher, Barbara
AU - Kannan, Sujatha
AU - Kannan, Rangaramanujam M.
N1 - Funding Information:
The authors would like to thank JHMI Bayview Immunology Flow Core (Dr. Mark Soloski and Raffaello Cimbro) and the JHMI Ross Flow Cytometry Core Facility (Xiaoling Zhang) for their assistance in fluorescence-activated cell sorting. The authors would also like to thank Dr. Wayne Mitzner for use of plethysmography equipment. This work was supported by the National Institutes of Health [Grants numbers R01 NS113140, U54 HD079123, R21NS10085] and the Hartwell Foundation, Memphis, TN.
Publisher Copyright:
© The author(s). This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/). See http://ivyspring.com/terms for full terms and conditions.
PY - 2020
Y1 - 2020
N2 - Background: Elevated glutamate production and release from glial cells is a common feature of many CNS disorders. Inhibitors of glutaminase (GLS), the enzyme responsible for converting glutamine to glutamate have been developed to target glutamate overproduction. However, many GLS inhibitors have poor aqueous solubility, are unable to cross the blood brain barrier, or demonstrate significant toxicity when given systemically, precluding translation. Enhanced aqueous solubility and systemic therapy targeted to activated glia may address this challenge. Here we examine the impact of microglial-targeted GLS inhibition in a mouse model of Rett syndrome (RTT), a developmental disorder with no viable therapies, manifesting profound central nervous system effects, in which elevated glutamatergic tone, upregulation of microglial GLS, oxidative stress and neuroimmune dysregulation are key features. Methods: To enable this, we conjugated a potent glutaminase inhibitor, N-(5-{2-[2-(5-amino-[1,3,4]thiadiazol-2-yl)-ethylsulfanyl]-ethyl}-[1,3,4]thiadiazol-2-yl)-2-phenyl-acetamide (JHU29) to a generation 4 hydroxyl PAMAM dendrimer (D-JHU29). We then examined the effect of D-JHU29 in organotypic slice culture on glutamate release. We also examined GLS activity in microglial and non-microglial cells, and neurobehavioral phenotype after systemic administration of D-JHU29 in a mouse model of RTT. Results: We report successful conjugation of JHU29 to dendrimer resulting in enhanced water solubility compared to free JHU29. D-JHU29 reduced the excessive glutamate release observed in tissue culture slices in a clinically relevant Mecp2-knockout (KO) RTT mouse. Microglia isolated from Mecp2-KO mice demonstrated upregulation of GLS activity that normalized to wild-type levels following systemic treatment with D-JHU29. Neurobehavioral assessments in D-JHU29 treated Mecp2-KO mice revealed selective improvements in mobility. Conclusion: These findings demonstrate that glutaminase inhibitors conjugated to dendrimers are a viable mechanism to selectively inhibit microglial GLS to reduce glutamate production and improve mobility in a mouse model of RTT, with broader implications for selectively targeting this pathway in other neurodegenerative disorders.
AB - Background: Elevated glutamate production and release from glial cells is a common feature of many CNS disorders. Inhibitors of glutaminase (GLS), the enzyme responsible for converting glutamine to glutamate have been developed to target glutamate overproduction. However, many GLS inhibitors have poor aqueous solubility, are unable to cross the blood brain barrier, or demonstrate significant toxicity when given systemically, precluding translation. Enhanced aqueous solubility and systemic therapy targeted to activated glia may address this challenge. Here we examine the impact of microglial-targeted GLS inhibition in a mouse model of Rett syndrome (RTT), a developmental disorder with no viable therapies, manifesting profound central nervous system effects, in which elevated glutamatergic tone, upregulation of microglial GLS, oxidative stress and neuroimmune dysregulation are key features. Methods: To enable this, we conjugated a potent glutaminase inhibitor, N-(5-{2-[2-(5-amino-[1,3,4]thiadiazol-2-yl)-ethylsulfanyl]-ethyl}-[1,3,4]thiadiazol-2-yl)-2-phenyl-acetamide (JHU29) to a generation 4 hydroxyl PAMAM dendrimer (D-JHU29). We then examined the effect of D-JHU29 in organotypic slice culture on glutamate release. We also examined GLS activity in microglial and non-microglial cells, and neurobehavioral phenotype after systemic administration of D-JHU29 in a mouse model of RTT. Results: We report successful conjugation of JHU29 to dendrimer resulting in enhanced water solubility compared to free JHU29. D-JHU29 reduced the excessive glutamate release observed in tissue culture slices in a clinically relevant Mecp2-knockout (KO) RTT mouse. Microglia isolated from Mecp2-KO mice demonstrated upregulation of GLS activity that normalized to wild-type levels following systemic treatment with D-JHU29. Neurobehavioral assessments in D-JHU29 treated Mecp2-KO mice revealed selective improvements in mobility. Conclusion: These findings demonstrate that glutaminase inhibitors conjugated to dendrimers are a viable mechanism to selectively inhibit microglial GLS to reduce glutamate production and improve mobility in a mouse model of RTT, with broader implications for selectively targeting this pathway in other neurodegenerative disorders.
KW - Glutaminase
KW - Microglia
KW - PAMAM dendrimer
KW - Rett syndrome
UR - http://www.scopus.com/inward/record.url?scp=85085909129&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85085909129&partnerID=8YFLogxK
U2 - 10.7150/thno.41714
DO - 10.7150/thno.41714
M3 - Article
C2 - 32483415
AN - SCOPUS:85085909129
SN - 1838-7640
VL - 10
SP - 5736
EP - 5748
JO - Theranostics
JF - Theranostics
IS - 13
ER -