Animal models - therapeutics Flashcards
Waterhouse et al. 1995
First demonstrated the importance of this regulatory process by creating CTLA4 knockout mice, which consequently had huge and uncontrollable proliferation and activation of T cells, leading to their death
Leach et al 1996
Showed CTLA4 to be a negative regulator of T cell activation, which earned a Nobel Prize. (worked with Jim Allison - Nobel Prize in Physiology or Medicine 2018)
They injected BALB/c mice with B7-1 transfected colon cancer cells.
These mice received an anti-CTLA4 antibody, an anti-CD28 antibody, or remained as untreated controls.
The anti-CTLA4 antibody led to the rejection of pre-established tumours – they regressed in size or were obliterated completely. In fact, 90% of the cancers in treated mice disappeared.
Sun et al 2020
used Zebrafish for forward-genetic screens aiming to identify mutations responsible for congenital scoliosis (CS), a complex genetic disorder in which there are vertebral malformations.
They performed an ENU mutagenesis screen and found a zebrafish line with CS-like vertebral malformations
Using CRISPR/Cas9 and positional cloning, they found that mutations in the dual serine/threonine and tyrosine kinase (dstyk) gene cause this phenotype
They showed that Dystk knockdown inhibits the formation of notochord vacuoles and lysosomes which causes these vertebral defects
Thus forward genetics in zebrafish revealed a gene likely responsible for CS, which previously had an unknown aetiology.
Yashamoto et al 2014
Performed a mosaic genetic screen of lethal mutations on the Drosophila X chromosome in attempt to identify genes responsible for rare genetic disorders
They identified 165 genes necessary for the development, maintenance and function of the nervous system. Simultaneously, they investigated rare variants in 1929 human exomes from families with unsolved Mendelian diseases
They then merged the fly and human datasets and identified the ANKLE2 gene (encoding Ankle2, a mitotic regulator) in a recessive disorder
They were then able to show that the ANKLE2 variant was responsible for a microcephaly phenotype, both in humans and in flies
Shah et al 2018
Showed that ANKLE2 protein interacts with a Zika protein called NS4A. By introducing this protein into Drosophila, it has been possible to show that this protein can also induce microencephaly → showed that NS4A inhibits ANKLE2 and thus contributes to ZIKV induced microcephaly
Budworth et al 2015
Created a novel mouse model in which the effects of inherited and somatic mutations could remarkably be separated.
OGG1 is a glycosylase involved in base excision repair that causes genetic instability at CAG repeats in mice with a truncated mHTT fragment, and OGG1 only initiates somatic expansion
In this study, the authors crossed Hdh heterozygous knock-in mice (which have disease-length CAG repeats in the mHTT locus) with OGG1(+/-) heterozygous knockout mice
The cross produced 9 genotypes that expressed combinations of the wild-type and expanded full-length mutant HD allele, and normal, lower, or absent OGG1. This enabled them to study the effect of each genotype on HD development independently.
They found that OGG1(-/-) mice had delayed HD onset by 7-10 months compared to OGG(+/+) mice, although they both inherit a similar disease-length HD allele
Moreover, a pharmacological agent (XJB-5-131) that decreases DNA substrates for OGG1 inhibited the lengthening of the repeat tracks and decreased instability.
Moreover, the authors proposed that the onset of toxicity would be the sum of the inherited and somatic expansions
Therefore, blocking somatic expansion is beneficial, so pharmacological approaches to delay disease progression could be possible.
Chen et al 2013
- Chen et al. (2013) explored this PFC hypoactivity in rats with compulsive cocaine-seeking behaviour using invasive shocks and subsequently optogenetics
- In their study, rats underwent extensive cocaine self-administration training (where they received cocaine by pressing a lever)
- After 8 weeks of self-administration, the rats receive foot shocks paired with the lever pressing and cocaine administration.
- One group stopped pressing the lever when they were shocked (these were defined as the shock-sensitive group), and the other group kept pressing lever, not caring that they were shocked (the shock-resistant group). In the latter group, drug-seeking behaviour therefore persisted in the face of harm, which is relatable to the human addiction phenotype
- When the authors performed whole-cell recordings in the prelimbic cortex, rats in the shock-resistant group needed more current to induce an action potential
- It was therefore speculated that if these PFC neurons could be artificially boosted, we could potentially rescue the phenotype of addiction
- They then explored this in an optogenetic model, whereby AAV expressing ChR2 and YFP was injected into the prelimbic cortex of shock-resistant rats, and photoactivation of the ChR2 neurons attenuated their foot-shock resistant cocaine-seeking behaviour
- This indicated that targeted stimulation of the PFC by deep brain stimulation could serve as a promising therapy for treating compulsive drug use
Terraneo et al 2016
conducted a pilot study where they applied brain stimulation in this manner using non-invasive TMS (transcranial magnetic stimulation) in attempt to reduce cocaine consumption, and their trial yielded promising results!
o They performed repetitive TMS (rTMS) in the dorsolateral PFC (DLPFC) in humans to see if this could reduce their cocaine usage
o 32 cocaine addicts were randomly assigned to a group receiving rTMS on the left DLPFC, or a control group (receiving pharmacological agents) for 29 days
o This was followed by a 63-day follow up, where everyone was offered rTMS
o No significant adverse effects were reported, a significantly higher number of cocaine-free urine tests were achieved in the rTMS group compared to the control group, and the rTMS group had significantly lower craving for cocaine
• This study supports the potential therapeutic use of TMS in cocaine addicts + warrants larger clinical trials
• Moreover, it shows how optogenetics in animal models can be used to reveal fundamental neuronal circuitry, which can then open the doors for therapeutic intervention
Wang et al (2017)
reported that YL001, a new drug that inhibits EG5 (a mitosis-specific microtubule-dependent motor protein), inhibits tumour growth in a variety of cell lines by 60% and when they tested this in a xenograft mouse model, it prolonged median survival time by over 50%.
Led to 7th April 2020 clinical trial
Ridges et al 2012
Used zebrafish to screen small compounds that may inhibit T cell maturation in embryonic development, as a potential anti-cancer therapy.
They tested over 25,000 molecules and identified Lenaldeker (LDK), a compound that eliminated mature T cells in zebrafish developments, and induced long-term remission in zebrafish with c-Myc-induced T-cell acute lymphoblastic leukaemia (T-ALL)
This was confirmed in mice to inhibit the growth of human T-ALL xenografts
McKernan et al 2000
Generated transgenic mice with a single amino acid substitution in the alpha 1 subunit, and found that sedation was not induced by diazepam in these mice, but the anxiolytic effects remain
There was however a caveat of this study, as the authors measured sedation by motor activity, but perhaps there was some kind of motor impairment instead of sedative effects.
They also administered a partial agonist of the alpha 2, 3 and 5 subunits to normal mice, with an antagonist at alpha 1, and found that anxiolysis was retained, but motor activity was not reduced
Seok et al 2013
- Seok et al. (2013) conducted a large-scale study that highlights how poorly mouse models of inflammation reflect the human condition
- This involved analysing the gene expression profiles of leukocytes in mouse models and human subjects that were elicited by inflammatory challenges (including trauma, sepsis and endotoxemia).
- The authors found that the expression profiles amongst humans across these diverse conditions were similar, suggesting a common pathway in humans independent of the initiating stress
- However, the expression profile in the corresponding mice models differed significantly from each other, and from the human data
- In fact, the correlations of the gene changes between humans and mice was equivalent to what would be expected by random chance
- This may explain why so many of the clinical trials for these disorders fail, and emphasises the need for more human-representative models, such as the DRAG mouse model
- There were however caveats to this work, for example they did only investigate one strain of mice, different microbiotal compositions in species could have influenced the profiles, and humans received medical care that wasn’t accounted for
Schenk et al 1999
o Schenk et al (1999) found that immunising the PDAPP transgenic mouse (which overexpresses mutant human APP hence develops the hallmarks of AD) with Aβ protein fragments prevented Aβ plaque formation and hence the development of AD in the young mice.
o The study also importantly showed that immunisation slowed progression and reduced extent of the disease in older mice, meaning that this method could potentially provide a treatment rather than just a prophylactic measure
Elan Corporation trial (2000)
disaster. 6% of patients got meningeal encephalitis. Trial abandoned in 2002
Oliver et al. (2009)
analysed the effects of variable pre-natal stress on the Bdr schizophrenia mouse model, and found that the genetic mutation together with the aversive pre-natal environment produced behaviour more realistic of schizophrenia
