Small molecule modulators of lncRNA NEAT1_2: A novel approach to enhancing the endogenous neuroprotective response in amyotrophic lateral sclerosis
Lead Research Organisation:
CARDIFF UNIVERSITY
Department Name: School of Biosciences
Abstract
Amyotrophic lateral sclerosis (ALS) is a type of motor neuron disease in which the nerve cells (motor neurons) that control the muscles degenerate and die. This causes in a decline in the ability of the patient to conduct everyday activities, with the progressive loss of function of the muscles responsible for breathing ultimately resulting in the patient dying of respiratory failure. Although in most cases the cause of ALS is unknown, in a small number of patients, the disease is caused by changes in the genes that encode a variety of different proteins. However, despite the different causes of ALS, a common feature in surviving motor neurons is the presence within the nucleus of structures called paraspeckles. These structures are thought to be a normal response to neuronal stress and are therefore considered to be neuroprotective. Consequently, it is hypothesised that increasing the number of paraspeckles should enhance the protection against the cellular stressors and thereby reduce the death of motor neurons in ALS.
A central component of the neuroprotective paraspeckle is NEAT1_2, an RNA that does not code for any protein (known as a long non-coding RNA or lncRNA). This molecule acts as a magnet which attracts proteins which stick to NEAT1_2 to form the mature paraspeckle. Our hypothesis is that if we can increase the amount of NEAT1_2 then the number of paraspeckles and resultant neuroprotective effects in ALS motor neurons will also be increased. RNA molecules are generally relatively shapeless which means it is difficult to identify compounds that can bind to them and alter their function. However, NEAT1_2 contains a highly structured region called the 'triple helix' which is amenable to the binding of small molecules. The binding of compounds to such structured regions is a very innovative approach to regulating the function of RNAs, and validation that this strategy can be successful has recently been demonstrated with another lncRNA, MALAT1.
In this project we aim to identify compounds that bind to the triple helix of NEAT1_2 with the aim of stabilising this lncRNA. This should increase the amount of NEAT1_2 in the nuclei of motor neurons which will in turn seed the formation of an increased number of neuroprotective paraspeckles in ALS and, potentially, other neurodegenerative disorders, thereby defining a new class of neuroprotective, disease-modifying drugs.
A central component of the neuroprotective paraspeckle is NEAT1_2, an RNA that does not code for any protein (known as a long non-coding RNA or lncRNA). This molecule acts as a magnet which attracts proteins which stick to NEAT1_2 to form the mature paraspeckle. Our hypothesis is that if we can increase the amount of NEAT1_2 then the number of paraspeckles and resultant neuroprotective effects in ALS motor neurons will also be increased. RNA molecules are generally relatively shapeless which means it is difficult to identify compounds that can bind to them and alter their function. However, NEAT1_2 contains a highly structured region called the 'triple helix' which is amenable to the binding of small molecules. The binding of compounds to such structured regions is a very innovative approach to regulating the function of RNAs, and validation that this strategy can be successful has recently been demonstrated with another lncRNA, MALAT1.
In this project we aim to identify compounds that bind to the triple helix of NEAT1_2 with the aim of stabilising this lncRNA. This should increase the amount of NEAT1_2 in the nuclei of motor neurons which will in turn seed the formation of an increased number of neuroprotective paraspeckles in ALS and, potentially, other neurodegenerative disorders, thereby defining a new class of neuroprotective, disease-modifying drugs.
Technical Summary
The unmet need
Amyotrophic lateral sclerosis (ALS), the most common form of motor neuron disease, is a fatal neuromuscular condition. The two drugs approved for ALS, riluzole and edaravone, extend patient survival by only a few months. Hence, novel disease-modifying therapies that are efficacious across different ALS subtypes are urgently needed.
Paraspeckles are a protective mechanism in ALS
ALS is aetiologically heterogeneous, with most cases being sporadic while the minority of cases that have a genetic cause can be divided into >25 genetic subtypes. However, an increase in the number of paraspeckles in the nuclei of motor neurons occurs irrespective of the cause of ALS. Paraspeckles are intranuclear membraneless ribonucleoprotein organelles (RNP granules) that have a protective function, and multiple structural proteins of paraspeckles are affected by ALS-causative mutations.
The lncRNA NEAT1_2 is a key component of the paraspeckle
Long noncoding RNA (lncRNA), NEAT1 (Nuclear Paraspeckle Assembly Transcript 1) exists in two isoforms, NEAT1_1 and NEAT1_2. The latter is the stress-induced isoform essential in the recruitment of RNA-binding proteins and the formation of paraspeckles (i.e., NEAT1_2 "seeds" the formation of paraspeckles).
Project hypothesis
Small molecules that stabilise NEAT1_2 will augment paraspeckle assembly within the motor neurons in ALS. This in turn will enhance their resilience and survival.
Project aim
To identify small molecules that interact with the triple helix (TH) structure at the 3' end of NEAT1_2. This TH is the NEAT1_2's stability element that is sufficiently complex to support small molecule binding.
Progress to date
This proposal describing a robust fluorescence displacement assay using NEAT1_2 TH RNA is based on work conducted by CoI Shelkovnikova when she worked with PI Atack and CoI Ward at the MDI. Secondary assays are established in CoI Shelkovnikova's new lab in Sheffield.
Amyotrophic lateral sclerosis (ALS), the most common form of motor neuron disease, is a fatal neuromuscular condition. The two drugs approved for ALS, riluzole and edaravone, extend patient survival by only a few months. Hence, novel disease-modifying therapies that are efficacious across different ALS subtypes are urgently needed.
Paraspeckles are a protective mechanism in ALS
ALS is aetiologically heterogeneous, with most cases being sporadic while the minority of cases that have a genetic cause can be divided into >25 genetic subtypes. However, an increase in the number of paraspeckles in the nuclei of motor neurons occurs irrespective of the cause of ALS. Paraspeckles are intranuclear membraneless ribonucleoprotein organelles (RNP granules) that have a protective function, and multiple structural proteins of paraspeckles are affected by ALS-causative mutations.
The lncRNA NEAT1_2 is a key component of the paraspeckle
Long noncoding RNA (lncRNA), NEAT1 (Nuclear Paraspeckle Assembly Transcript 1) exists in two isoforms, NEAT1_1 and NEAT1_2. The latter is the stress-induced isoform essential in the recruitment of RNA-binding proteins and the formation of paraspeckles (i.e., NEAT1_2 "seeds" the formation of paraspeckles).
Project hypothesis
Small molecules that stabilise NEAT1_2 will augment paraspeckle assembly within the motor neurons in ALS. This in turn will enhance their resilience and survival.
Project aim
To identify small molecules that interact with the triple helix (TH) structure at the 3' end of NEAT1_2. This TH is the NEAT1_2's stability element that is sufficiently complex to support small molecule binding.
Progress to date
This proposal describing a robust fluorescence displacement assay using NEAT1_2 TH RNA is based on work conducted by CoI Shelkovnikova when she worked with PI Atack and CoI Ward at the MDI. Secondary assays are established in CoI Shelkovnikova's new lab in Sheffield.
