Brain pharmacology of intrathecal antisense oligonucleotides revealed through multimodal imaging

Mazur, Curt; Powers, Berit; Zasadny, Kenneth R.; Sullivan, Jenna M.; Dimant, Hemi; Kamme, Fredrik; Hesterman, Jacob; Matson, John; Oestergaard, Michael; Seaman, Marc; Holt, Robert W.; Qutaish, Mohammed Q.; Polyak, Ildiko; Coelho, Richard V.; Gottumukkala, Vijay; Gaut, Carolynn M.; Berridge, Marc S.; Albargothy, Nazira J.; Kelly, Louise; Carare, Roxana O.; Hoppin, Jack; Kordasiewicz, Holly; Swayze, Eric E.; Verma, Ajay

doi:10.1172/jci.insight.129240

Cited by 62 publications

(71 citation statements)

References 44 publications

(99 reference statements)

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“…Studies with radiolabeled oligonucleotides and immunohistochemical studies with a phosphorothioate ASO–specific antibody have yielded insights into the ASO pharmacokinetics ( 8–10 ), but insights into ASO distribution in different cell types and subcellular compartments is typically lost. Fluorescence microscopy studies involving ASOs and siRNAs conjugated to fluorescent tags have made it possible to visualize ASOs ( 11–13 ), but those studies also have limitations. First, modifying NATs with fluorescent dyes (e.g.…”

Section: Introductionmentioning

confidence: 99%

High-resolution visualization and quantification of nucleic acid–based therapeutics in cells and tissues using Nanoscale secondary ion mass spectrometry (NanoSIMS)

Migawa

Chen

et al. 2020

Nucleic Acids Research

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Nucleic acid therapeutics (NATs) have proven useful in promoting the degradation of specific transcripts, modifying gene expression, and regulating mRNA splicing. In each situation, efficient delivery of nucleic acids to cells, tissues and intracellular compartments is crucial—both for optimizing efficacy and reducing side effects. Despite successes in NATs, our understanding of their cellular uptake and distribution in tissues is limited. Current methods have yielded insights into distribution of NATs within cells and tissues, but the sensitivity and resolution of these approaches are limited. Here, we show that nanoscale secondary ion mass spectrometry (NanoSIMS) imaging can be used to define the distribution of 5-bromo-2′-deoxythymidine (5-BrdT) modified antisense oligonucleotides (ASO) in cells and tissues with high sensitivity and spatial resolution. This approach makes it possible to define ASO uptake and distribution in different subcellular compartments and to quantify the impact of targeting ligands designed to promote ASO uptake by cells. Our studies showed that phosphorothioate ASOs are associated with filopodia and the inner nuclear membrane in cultured cells, and also revealed substantial cellular and subcellular heterogeneity of ASO uptake in mouse tissues. NanoSIMS imaging represents a significant advance in visualizing uptake and distribution of NATs; this approach will be useful in optimizing efficacy and delivery of NATs for treating human disease.

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Section: Introductionmentioning

confidence: 99%

High-resolution visualization and quantification of nucleic acid–based therapeutics in cells and tissues using Nanoscale secondary ion mass spectrometry (NanoSIMS)

Migawa

Chen

et al. 2020

Nucleic Acids Research

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“…Viral delivery of gene therapy achieves good CNS distribution but does not effectively target non-neuronal cells (46). Recently, we showed that following IT bolus injection in rats, ASO distributes rostrally in the CSF along the neuroaxis and rapidly adheres to pia and arterial walls, consistent with perivascular entry into the CNS (20). Over time, ASO accumulates in all major CNS cell types presumably by receptor mediated endocytosis (47).…”

Section: Csf Volume Scaling Across Species To Predict Aso Tissue Concmentioning

confidence: 97%

“…Antisense oligonucleotides (ASOs) are chemically modified single-stranded nucleic acids of [16][17][18][19][20][21][22][23][24][25] nucleotides in length that bind complementary RNA via Watson-Crick hybridization and can modulate the expression of genes by harnessing a variety of mechanisms (1). To reduce the expression of genes, ASOs can be designed to harness the RNase H1 mechanism (referred to herein as gapmers) (2).…”

Section: Introductionmentioning

confidence: 99%

“…In larger preclinical species such as rats and NHPs, we have previously demonstrated widespread distribution and activity of MOE-gapmer and uniform MOE ASOs in the CNS following central administration (8,(20)(21)(22)(23). Using a battery of live imaging techniques and postmortem analyses, MOEgapmer ASOs were shown to immediately distribute into the cranium following intrathecal (IT) injection in the rat and penetrate all major CNS structures by 6 h post-injection (20,23). Cellular uptake took place between 6-24 hours post-injection and stable regional and cellular distribution was observed from at least 1 to 14 days post-dosing.…”

Section: Introductionmentioning

confidence: 99%

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The atlas of RNase H antisense oligonucleotide distribution and activity in the CNS of rodents and non-human primates following central administration

Jafar‐Nejad

Powers

Soriano

et al. 2020

Preprint

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Antisense oligonucleotides (ASOs) have emerged as a new class of drugs to treat a wide range of diseases, including neurological indications. Spinraza, an ASO that modulates splicing of SMN2 RNA, has shown profound disease modifying effects in Spinal Muscular Atrophy (SMA) patients, energizing the field to develop ASOs for other neurological diseases. While SMA specifically affects spinal motor neurons, other neurological diseases affect different central nervous system (CNS) regions, neuronal, and non-neuronal cells. Therefore, it is critically important to characterize ASO distribution and activity in all major CNS structures and cell types to have a better understanding of which neurological diseases are amenable to ASO therapy. Here we present for the first time the atlas of ASO distribution and activity in the CNS of mice, rats, and non-human primates (NHP), species commonly used in preclinical therapeutic development. Following central administration of an ASO to rodents, we observe widespread distribution and robust activity throughout the CNS in neurons, oligodendrocytes, astrocytes, and microglia. This is also the case in NHP, despite a larger CNS volume and more complex neuroarchitecture. Our results demonstrate that ASO drugs are well suited for treating a wide range of neurological diseases for which no effective treatments are available.

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“…Brain regional analysis was done using a 3-dimensional mouse brain atlas available as the VivoQuant software plug-in. The 3D brain atlas is based on the Paxinos-Franklin atlas registered to a series of high-resolution magnetic resonance imaging with 100 µm near isotropic data and has been applied in other studies [15][16][17] . The brain was segmented into 14 regions within the skull of each animal using the CT as the reference for automatic registration.…”

Section: Data Analyses Of [ 18 F]fdg Uptakementioning

confidence: 99%

Assessment of Brain Glucose Metabolism Following Cardiac Arrest by [¹⁸F]FDG Positron Emission Tomography

Mitchell

Fang

Tsai

et al. 2020

Preprint

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AbstractBackgroundCardiac arrest (CA) patients who survived by cardiopulmonary resuscitation (CPR) can present different levels of neurological deficits ranging from minor cognitive impairments to persistent vegetative state and brain death. The pathophysiology of the resulting brain injury is poorly understood and whether changes in post-CA brain metabolism contribute to the injury are unknown. Here we utilized [18F]FDG-PET to study in vivo cerebral glucose metabolism 72 hours following CA in a murine cardiac arrest model.MethodsAnesthetized and ventilated adult C57BL/6 mice underwent 12-minute KCl-induced CA followed by CPR. Seventy-two hours following cardiac arrest, surviving mice were intraperitoneally injected with [18F]FDG (~186 μCi/200 μL) and imaged on Molecubes preclinical micro PET/CT imaging systems after a 30-minute awake uptake period. Brain [18F]FDG uptake was determined by the VivoQuant software on fused PET/CT images with the 3D brain atlas. Upon completion of PET imaging, remaining [18F]FDG radioactivity in the brain, heart, and liver was determined using a gamma counter.ResultsGlobal increases in brain [18F]FDG uptake in post-CA mice were observed compared to shams and controls. The median standardized uptake value (SUV) of [18F]FDG for CA animals was 1.79 vs. sham 1.25 (p<0.05) and control animals 0.78 (p<0.01). This increased uptake was consistent throughout the 60-minute imaging period and across all brain regions reaching statistical significance in the midbrain, pons, and medulla. Biodistribution analyses of various key organs yielded similar observations that the median [18F]FDG uptake for brain were 7.04%ID/g tissue for CA mice vs 5.537%ID/g tissue for sham animals, p<0.05).ConclusionsThis study has successfully applied [18F]FDG-PET/CT to measure changes in brain metabolism in a murine model of asystolic CA. Our results demonstrate increased [18F]FDG uptake in the brain 72 hours following CA, suggesting increased metabolic demand in the case of severe neurological injury. Further study is warranted to determine the etiology of these changes.

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Brain pharmacology of intrathecal antisense oligonucleotides revealed through multimodal imaging

Abstract: CMG were employees of Invicro, LLC when the studies were performed. JMS is currently an employee of Biogen Inc. NJA, LK, and ROC are employees of the University of Southampton. MB is an employee of 3D Imaging, and AV was an employee of Biogen Inc. at the time that this work was performed.

Cited by 62 publications

References 44 publications

High-resolution visualization and quantification of nucleic acid–based therapeutics in cells and tissues using Nanoscale secondary ion mass spectrometry (NanoSIMS)

High-resolution visualization and quantification of nucleic acid–based therapeutics in cells and tissues using Nanoscale secondary ion mass spectrometry (NanoSIMS)

The atlas of RNase H antisense oligonucleotide distribution and activity in the CNS of rodents and non-human primates following central administration

Assessment of Brain Glucose Metabolism Following Cardiac Arrest by [¹⁸F]FDG Positron Emission Tomography

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Brain pharmacology of intrathecal antisense oligonucleotides revealed through multimodal imaging

Abstract: CMG were employees of Invicro, LLC when the studies were performed. JMS is currently an employee of Biogen Inc. NJA, LK, and ROC are employees of the University of Southampton. MB is an employee of 3D Imaging, and AV was an employee of Biogen Inc. at the time that this work was performed.

Cited by 62 publications

References 44 publications

High-resolution visualization and quantification of nucleic acid–based therapeutics in cells and tissues using Nanoscale secondary ion mass spectrometry (NanoSIMS)

High-resolution visualization and quantification of nucleic acid–based therapeutics in cells and tissues using Nanoscale secondary ion mass spectrometry (NanoSIMS)

The atlas of RNase H antisense oligonucleotide distribution and activity in the CNS of rodents and non-human primates following central administration

Assessment of Brain Glucose Metabolism Following Cardiac Arrest by [18F]FDG Positron Emission Tomography

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About

Assessment of Brain Glucose Metabolism Following Cardiac Arrest by [¹⁸F]FDG Positron Emission Tomography