Periodic Reporting for period 1 - AlzheimersInAction (AlzheimersInAction: A multi-disciplinary approach to determine the cellular and molecular mechanisms underlying the inflammatory response to amyloid-β in Alzheimer’s disease.)
Reporting period: 2021-09-01 to 2023-08-31
In addition to devastation for the patients and their families, Alzheimer’s disease (AD) generates enormous costs for the wider European economy and is an urgent health priority. There are currently no drug treatments that can cure AD or other common forms of dementia, though recently there have been great progress with promising results in clinical trials with Lecanamab and Donanemab able to slow AD progression by 27% and 35% respectively. The absence of therapeutic interventions has come from a lack of understanding about the biological changes that cause most neurological diseases. It appears that we are now at a turning point in the fight against AD, and it is critical to fill in the gaps in our understanding of what biological changes are occurring during the early stages of AD in order to improve upon the development of treatments so we can further slow progression or delay the onset of AD.
To overcome the difficulties of co-morbidity and capturing processes in real time, we took a multi-disciplinary approach that combines and develops cutting-edge techniques in Drosophila cell biology, molecular biology, and time-lapse imaging, in parallel with exploitation of the powerful molecular genetics, versatile promoter systems and rapid development of flies. Our aim was to identify potential targets for therapeutic intervention that would slow or stop AD progression from the earliest stages. Throughout AlzheimersInAction, we developed an in-depth understanding of how an overproduction of amyloid-β (Aβ) causes downstream pathologies, and how activation of the immune cells during AD causes disease pathogenesis. Additionally, we undertook a large-scale compound screen which identified pathways that, when altered, could the overproduction of Aβ that occurs at the earliest stages of AD.
In conclusion, our findings from AlzheimersInAction have have identified compounds and pathways that can be targeted to slow or even stop the overproduction of Aβ that occurs in early AD. With recent clinical success using drugs that target Aβ at the earliest stages of AD, we believe that our findings can help to improve upon the effectiveness of such drugs and greatly improve the livelihood of Alzheimer's patients.
To overcome the difficulties of co-morbidity and capturing processes in real time, we took a multi-disciplinary approach that combines and develops cutting-edge techniques in Drosophila cell biology, molecular biology, and time-lapse imaging, in parallel with exploitation of the powerful molecular genetics, versatile promoter systems and rapid development of flies. Our aim was to identify potential targets for therapeutic intervention that would slow or stop AD progression from the earliest stages. Throughout AlzheimersInAction, we developed an in-depth understanding of how an overproduction of amyloid-β (Aβ) causes downstream pathologies, and how activation of the immune cells during AD causes disease pathogenesis. Additionally, we undertook a large-scale compound screen which identified pathways that, when altered, could the overproduction of Aβ that occurs at the earliest stages of AD.
In conclusion, our findings from AlzheimersInAction have have identified compounds and pathways that can be targeted to slow or even stop the overproduction of Aβ that occurs in early AD. With recent clinical success using drugs that target Aβ at the earliest stages of AD, we believe that our findings can help to improve upon the effectiveness of such drugs and greatly improve the livelihood of Alzheimer's patients.
WP1: To investigate different aspects of Aβ overproduction in AD patients and what causes neurodegeneration, we created several fly lines that recapitulated different aspects of this overproduction. Results: Length of Aβ was important in causing toxic effects and also secretion from neurons. This led to cognitive defects and reduced longevity. Subsequent immunostaining showed that plaque-like deposits of Aβ were not the cause of toxic effects.
WP2: We set out to determine if neuronal damage is localised to neurons that express Aβ or if non-secreting neighbouring neurons are also affected. We expressed toxic Aβ from subsets of neurons and assessed how expressing and non-expressing neighbouring neurons were affected. Results: Certain subsets of neurons are more susceptible to Aβ accumulation. Aβ was observed at distant neurons and, often these neurons had greater accumulation than some neurons that were actually producing and secreting the Aβ. These results support the observations that not all brain regions are affected equally in AD patients - some regions are more susceptible than others. It is unknown why different brain regions have varying levels of vulnerability. Now that we have replicated these findings in a model that is much easier and faster to genetically manipulate and image, we continue to aim to provide novel insight into this unknown.
WP3: We aimed to dissect the molecular mechanisms involved in these phenotypes. In addition to the plethora of genetic tools at our disposal, thanks to the genetic tractability of Drosophila, we also had the benefit of being able to feed the animals compounds as an alternative approach to investigate different pathways involved. Due to the timeframe of the project, we sped up our search of pathways by systematically worked through a large range of compounds by feeding them to the animals for several days and then assessing the effects on cognitive defects and longevity. We then developed RNAi fly lines and mutant lines for genes of interest identified through the screen. A list which grew longer as we gained novel insights from the compound screen. Results: Of particular interest to our research group, was the role of efferocytosis in our phenotype. We found that inhibition of this process could reduce the cognitive defects. The speed at which efferocytosis occurs has previously presented an immense challenge for researchers aiming to unravel this process, however, fortunately for us, over the past few years the fly embryo has emerged as a powerful model for studying this process in vivo. As such, we have started to develop a novel state-of-the-art understanding of the role that efferocytosis plays downstream of the overproduction of hAβ42 in the brain. Information which, as it stands, is close to peer-reviewed publication, but additionally, opens up a wealth of research for me to continue in my own lab when I secure the opportunity.
Overview
The project has found that the length of Aβ was important for causing toxic effects and also that it needed to be secreted from neurons. We also identified the importance of the Aβ being secreted, as there were no ill effects in our flies when the amyloid peptide was not secreted. We also discovered that certain types of neurons are more susceptible to Aβ accumulation, whether or not they produce this peptide themselves. Furthermore, we found that inhibition of efferocytosis could improve cognitive defects and have continued building a clear picture of how this process occurs using state-of-the-art equipment and techniques.
Exploitation and dissemination
Research was presented regularly at seminars in the university to a diverse audience [6 seminars], and internationally at the large neuroscience, Neuroscience 2022, and Drosophila, 62th ADRC, conferences [2 conferences]. We have established new collaborations both in the UK and the EU to assist in aspects of the project outside our field of expertise. Results are being compiled and developed for peer-review publication. No specific website has been developed for the project.
WP2: We set out to determine if neuronal damage is localised to neurons that express Aβ or if non-secreting neighbouring neurons are also affected. We expressed toxic Aβ from subsets of neurons and assessed how expressing and non-expressing neighbouring neurons were affected. Results: Certain subsets of neurons are more susceptible to Aβ accumulation. Aβ was observed at distant neurons and, often these neurons had greater accumulation than some neurons that were actually producing and secreting the Aβ. These results support the observations that not all brain regions are affected equally in AD patients - some regions are more susceptible than others. It is unknown why different brain regions have varying levels of vulnerability. Now that we have replicated these findings in a model that is much easier and faster to genetically manipulate and image, we continue to aim to provide novel insight into this unknown.
WP3: We aimed to dissect the molecular mechanisms involved in these phenotypes. In addition to the plethora of genetic tools at our disposal, thanks to the genetic tractability of Drosophila, we also had the benefit of being able to feed the animals compounds as an alternative approach to investigate different pathways involved. Due to the timeframe of the project, we sped up our search of pathways by systematically worked through a large range of compounds by feeding them to the animals for several days and then assessing the effects on cognitive defects and longevity. We then developed RNAi fly lines and mutant lines for genes of interest identified through the screen. A list which grew longer as we gained novel insights from the compound screen. Results: Of particular interest to our research group, was the role of efferocytosis in our phenotype. We found that inhibition of this process could reduce the cognitive defects. The speed at which efferocytosis occurs has previously presented an immense challenge for researchers aiming to unravel this process, however, fortunately for us, over the past few years the fly embryo has emerged as a powerful model for studying this process in vivo. As such, we have started to develop a novel state-of-the-art understanding of the role that efferocytosis plays downstream of the overproduction of hAβ42 in the brain. Information which, as it stands, is close to peer-reviewed publication, but additionally, opens up a wealth of research for me to continue in my own lab when I secure the opportunity.
Overview
The project has found that the length of Aβ was important for causing toxic effects and also that it needed to be secreted from neurons. We also identified the importance of the Aβ being secreted, as there were no ill effects in our flies when the amyloid peptide was not secreted. We also discovered that certain types of neurons are more susceptible to Aβ accumulation, whether or not they produce this peptide themselves. Furthermore, we found that inhibition of efferocytosis could improve cognitive defects and have continued building a clear picture of how this process occurs using state-of-the-art equipment and techniques.
Exploitation and dissemination
Research was presented regularly at seminars in the university to a diverse audience [6 seminars], and internationally at the large neuroscience, Neuroscience 2022, and Drosophila, 62th ADRC, conferences [2 conferences]. We have established new collaborations both in the UK and the EU to assist in aspects of the project outside our field of expertise. Results are being compiled and developed for peer-review publication. No specific website has been developed for the project.
Our discoveries, along with the tools we have developed, have allowed us to advance AD research and Drosophila research beyond state-of-the-art. For viewing the immune responses to Aβ, we have used developed cutting edge time-lapse imaging of the CNS in living, developing whole animals. To explore how Aβ overproduction affects the immune response, we used novel lines that utilised the pioneering CharON probe which allows visualisation of efferocytosis in real time. As such, we have started to develop a novel state-of-the-art understanding of the role that efferocytosis plays downstream of the overproduction of hAβ42 in the brain. Our direct comparison of secreted Aβ and non-secreted Aβ advances our understanding and development of future Aβ-based AD models. We have advanced the current understanding of Aβ depositon and accumulation, showing it is more complex than previously understood. The identification of a number of pathways that can be targeted to slow or even stop the overproduction of Aβ that occurs in early AD, will allow us to further progress AD therapeutics beyond state-of-the-art.