CELL REPLACEMENT THERAPY FOR PARKINSON’S DISEASE: The importance of neuronal subtype, cell source and connectivity for functional recovery

Abstract

Parkinson’s disease (PD) is a neurodegenerative disorder characterised by motor deficits such as slowness in movement, difficulty in initiating movement and tremor at rest. The cause of these motor symptoms is the selective loss of mesencephalic dopaminergic (mesDA) neurons, located in the substantia nigra (SN). These neurons project axons to the striatum where they release dopamine, a neurotransmitter that controls voluntary movement. Current drug treatments restore the lost dopamine, while initially efficacious, the beneficial effects wear off resulting in severe side effects. Thus, there is a clear requirement for alternative therapeutic options.

One such idea is cell replacement therapy (CRT). CRT aims to replace neurons that have degenerated in PD, with donor cells that have the potential to functionally re-integrate into the host circuitry. This involves transplantation of developing midbrain cells from aborted fetuses, (the part that form mesDA neurons), into the striatum of a PD patient. Clinical trials have demonstrated that CRT can provide long-lasting, significant clinical benefit. Although some patients do not respond as favourably. We still do not know what specific factors contribute to the success in transplantation i.e. what cells are responsible for motor recovery? Can the transplants reform damaged neuronal circuitry? Use of human fetal tissue raises several ethical issues, but are there alternative cell sources that can substitute effectively? The aim of this thesis was to understand how particular factors such as neuronal content, placement and cell source, affect functional outcome after transplantation into the rodent brain.

In paper №1, I detail the neurodegenerative and behavioural outcomes in a mouse lesion model of PD, which can be used as a platform for the development of novel therapeutic strategies. I also describe the development of a novel behavioural task that is predictive of mesDA neuron cell loss in mice. Previously, it was thought that transplanted neurons could not extend axons over long distances rendering transplantation into the SN a non-viable approach. In paper №2, I describe how mesDA neurons transplanted in the adult SN of a PD mouse model, extended axons across millimetres into the striatum, functionally reforming the nigrostriatal pathway. In paper №3, I also identify the specific mesDA population (A9) that is critical for functional recovery, with transplants that lack A9 neurons failing to improve motor recovery. A potentially pre-clinical aspect of this thesis is detailed in paper №4 where I describe a robust protocol for the generation of functional mesDA neurons from human embryonic stem cells that are functional in a rat model of PD. No evidence of tumour formation was observed in the transplanted animals, a major concern when utilising a pluripotent cell source.

Through understanding functional recovery in terms of neuronal subtype and connectivity, the work presented in this thesis aims to bring the prospect of CRT closer to the clinic, I also describe the generation of a very promising alternative cell source that could rival fetal tissue. Together this work contributes to making CRT a reality for the treatment of PD.

Abstract (Swedish)

Popular Abstract in Swedish

Patienter som drabbats av Parkinsons sjukdom (PD) saknar förmågan att medvetet kontrollera sina rörelser. Livslång medicinering krävs, vilket lindrar symptomen och leder till att rörelsemönstret förbättras, men med varierad effektivitet. Trots att de läkemedel (t ex L-DOPA) som finns tillgängliga idag har fantastisk effekt, minskar deras effektivitet i takt med att sjukdomen fortgår och ger istället en högre grad bieffekter. Förutom medicinering finns det idag få andra möjligheter till behandling av sjukdomen och efterfrågan på nya behandlingsformer är stor. Vi vet att det är de dopaminerga nervcellerna i mellanhjärnan (mesDA nervceller), vilka ansvarar för kontrollen av rörelser, som dör av då PD fortskrider. Kan det därför vara ett bra alternativ till behandling att försöka ersätta dessa celler?

Ett lovande och framgångsrikt tillvägagångssätt har visat sig vara att ta ut den del av hjärnan som ger upphov till mesDA nervceller från donerade aborterade foster, och sedan injicera dessa friska nervceller till patienter med PD. Resultat från dessa kliniska studier har visat att cellerna kan överleva i minst 14 år och att vissa patienter helt har kunnat sluta sin medicinering med L-DOPA. Dessvärre har inte alla patienter reagerat lika positivt på behandlingen och för vissa patienter har det inte blivit någon förbättring alls. Det finns också ett etiskt dilemma i och med användandet av celler från aborterade foster och om cellterapi ska kunna bli ett alternativ till behandling i stor skala måste man undersöka alternativa sätt att tillhandage dessa typer celler. Arbetet som presenteras i denna avhandling har till uppgift att ge förståelse för vilka faktorer som påverkar funktionaliteten av transplanterade mesDA nervceller i ett preklinisk sammanhang, d.v.s. var nervcellerna är placerade, vilka nervceller som är viktiga för återhämtning och vilka celler som bör användas.

Det typiska är att cellerna transplanteras in i striatum, vilket är det område där cellerna och dopaminet har funktionella effekter. Vi upptäckte i studier på möss att om mesDA celler transplanteras in i mellanhjärnan, där de i vanliga fall är placerade i en frisk hjärna kan cellerna innervera striatum flera millimeter bort, något som man tidigare trodde var omöjligt. Vi undersökte också vilka typer av mesDA nervceller som ger förbättring och återhämtning efter transplantation. Det visade sig att om mesDA nervceller som associeras med motorisk kontroll (A9) saknades förbättrades inte det motoriska mönstret i de djur som transplanterats. En sista fråga som behandlas i denna avhandling är huruvida man kan använda andra källor för att generera rätt typ av mesDA nervceller än fetal vävnad, så som humana stamceller. I vår studie kom vi fram till att det är möjligt att generera mesDA nervceller från humana stamceller med egenskaper förvånansvärt lika de i ett foster. De mesDA nervceller som kommer från stamcellerna kan överleva en transplantation, bildar inga tumörer, och bidrar till förbättrad funktion.

Arbetet i denna avhandling har bidragit till en ökad förståelse för hur transplantation av mesDA nervceller ger sin positiva effekt och vilka faktorer som bör tas hänsyn till om användandet av andra källor än fetal vävnad, så som stamceller, är önskvärt. Förhoppningen med detta arbete är att ta cellterapi som behandlingsform för Parkinsons sjukdom ett steg närmare kliniken.



Popular Abstract in English

Patients that suffer from Parkinson’s disease (PD) have lost the faculty to consciously control their movement. Lifelong medication is needed, which relieves symptoms, but often with different efficiencies. Although these drugs, namely L-DOPA, provide amazing clinical benefit: as the disease progresses and the drug continues to cause more side effects, the mobility of the patient decreases. Apart from medication, there are little clinical options available to patients, therefore there is a clear need for new therapies. We know that dopamine neurons of the midbrain (mesDA neurons) are responsible for controlling movement, and that these cells are lost during the progression of PD. Therefore, might it be feasible to try and replace these lost cells?

The most successful approach is called cell replacement therapy. This involves taking the region of the developing brain that will form mesDA neurons (from a fetus donated from elective abortions) and grafting these developing neurons into a patients brain. Initial clinical trials have proved that patients can survive with functioning grafts for at least 14 years and in some cases stop taking L-DOPA altogether. However this is the best-case scenario and not all patients respond to the same extent. Additionally, it is important for the widespread application of cell replacement therapy to consider alternative cell sources for large-scale application, given the ethical implications arising from the source of the tissue. The work presented in this thesis aims to understand what factors affect functionality of transplanted mesDA neurons in a preclinical setting; namely where the neurons are placed, what neurons are responsible for recovery and what cells are used.

In order to ensure a functional effect, cells are typically transplanted into their destination or target structure (the striatum). In studies in mice we have discovered that if mesDA cells are transplanted into their site of origin (the midbrain), they can make functional connections to the striatum which is several millimetres away. This phenomenon is something that was previously believed to be unachievable. The next question was to understand what type of mesDA neuron is responsible for mediating motor recovery after transplantation. Here we found that if the mesDA neurons associated with motor control (A9) are missing from transplants, a PD animal model does not recover motor function. The final question addressed in this thesis is: can use cell sources other than human fetal tissue (i.e. human stem cells), to generate a true mesDA neuron. We found that we can generate mesDA neurons from human embryonic stem cells that are remarkably similar to their fetal counterparts. The stem cell derived mesDA neurons can survive transplantation, not generate tumours, and provide functional benefit.

The work in this thesis has contributed to advancing our understanding of how transplants of mesDA neurons elicit their beneficial effects, and what factors we should consider when using alternative cell sources such as stem cells. It is hoped that this work brings us closer towards a cell replacement therapy for Parkinson’s disease.

• author: Grealish, Shane
• supervisor: Anders Björklund; Malin Parmar; Lachlan Thompson
• opponent: Prof. Macklis, Jeffrey, Harvard University
• organization: Neurobiology (research group); MultiPark: Multidisciplinary research focused on Parkinson´s disease
• publishing date: 2012
• type: Thesis
• publication status: published
• subject: Medical and Health Sciences; Basic Medicine; Neurosciences
• keywords: mouse behaviour, 6-hydroxydopamine, human embryonic stem cells, fetal tissue, connectivity, midbrain dopamine neuron subtype, cell transplantation, Parkinson’s disease
• in: Lund University Faculty of Medicine Doctoral Dissertation Series
• volume: 2012:4
• pages: 180 pages
• defense location: Segerfalksalen, Wallenberg Neuroscience Center
• defense date: 2012-01-27 13:00:00
• ISSN: 1652-8220
• ISBN: 978-91-86871-66-6
• language: English
• LU publication?: yes
• id: 74dbbb7c-8b19-4e17-bbb8-3613b7140800 (old id 2273302)
• date added to LUP: 2016-04-01 12:55:00
• date last changed: 2023-04-18 20:48:11