Tracking of systemic primary glioblastoma cell lines in vivo (copy)
1 Purpose
Glioblastoma (GBM) show the highest incidence rates among primary brain tumors in adults with the most aggressive growth pattern. Despite current standard of treatment including resection, radiotherapy and adjuvant chemotherapy the prognosis is still dismal with a median survival of 14.6 months.
Although GBMs grow highly invasive, they rarely metastasize from the central nervous system. So far, the reason for this remains unknown. The previous assumption the blood-brain-barrier might function as a physical hurdle was questioned, when circulating tumor cells were found in blood samples of GBM patients without systemic metastases (Mourad, Farrell et al. 2005, Krol, Castro-Giner et al. 2018). Therefore, the current study aims to analyze the dissemination pattern and the underlying mechanisms of systemically injected GBM cells in vivo.
2 Distress
We have extensive experience regarding intracardiac injection and imaging of small animals and both procedures are causing mild harm to the animals. However, the growth of the tumors will cause moderate distress to the animals. The animals will be closely monitored and euthanized if the attached objective endpoint criteria are met.
3 Expected benefit
We hope to gain fundamental and novel insights into the spreading pattern of systemically injected GBM cells, which may contribute to a better understanding of glioma biology and lead to improved treatment strategies.
4 Number of animals, and what kind
155 nod/scid mice.
5 How to adhere to 3R
Replacement:
This project aims to analyze the dissemination pattern of systemically injected GBM cells in an organism. We therefore want to evaluate which organs will be invaded and to what extent, which cannot be investigated in current in vitro models.
Refinement:
The intracardiac injection technique has been refined within our group. Two trained people will be used, one to inject the tumor cells, and the other responsible for animal care. While the animals will be heated, the ultrasound-guided technique ensures the quickest and most accurate injections. Thus, the injections cause mild harm to the animals. During imaging, two trained persons will be present, one to prepare and monitor the animals, and the other to operate the imaging instrumentation. The animals will be heated during imaging. The imaging procedures do not cause any harm to the animals.
Regarding potential harm caused by tumor growth, animals will be monitored daily by trained people and weighed twice/week. As imaging will be performed regularly, we will be able to detect the tumor burden prior to any symptoms in animal behavior. Based on imaging and examination of the animals, we will record the animals’ well-being via our assessment system. Based on the specific criteria mentioned in the score-sheet, analgesia will be administered as early as necessary and animals will be sacrificed if the objective endpoints are met, prior to developing severe harm.
Reduction:
Using the ultrasound-guided injection technique, the success rate of tumor development is 100%, as our group was able to prove. Thus, we can cut down the number of animals in each experimental group (from 15 to 13 animals).
Glioblastoma (GBM) show the highest incidence rates among primary brain tumors in adults with the most aggressive growth pattern. Despite current standard of treatment including resection, radiotherapy and adjuvant chemotherapy the prognosis is still dismal with a median survival of 14.6 months.
Although GBMs grow highly invasive, they rarely metastasize from the central nervous system. So far, the reason for this remains unknown. The previous assumption the blood-brain-barrier might function as a physical hurdle was questioned, when circulating tumor cells were found in blood samples of GBM patients without systemic metastases (Mourad, Farrell et al. 2005, Krol, Castro-Giner et al. 2018). Therefore, the current study aims to analyze the dissemination pattern and the underlying mechanisms of systemically injected GBM cells in vivo.
2 Distress
We have extensive experience regarding intracardiac injection and imaging of small animals and both procedures are causing mild harm to the animals. However, the growth of the tumors will cause moderate distress to the animals. The animals will be closely monitored and euthanized if the attached objective endpoint criteria are met.
3 Expected benefit
We hope to gain fundamental and novel insights into the spreading pattern of systemically injected GBM cells, which may contribute to a better understanding of glioma biology and lead to improved treatment strategies.
4 Number of animals, and what kind
155 nod/scid mice.
5 How to adhere to 3R
Replacement:
This project aims to analyze the dissemination pattern of systemically injected GBM cells in an organism. We therefore want to evaluate which organs will be invaded and to what extent, which cannot be investigated in current in vitro models.
Refinement:
The intracardiac injection technique has been refined within our group. Two trained people will be used, one to inject the tumor cells, and the other responsible for animal care. While the animals will be heated, the ultrasound-guided technique ensures the quickest and most accurate injections. Thus, the injections cause mild harm to the animals. During imaging, two trained persons will be present, one to prepare and monitor the animals, and the other to operate the imaging instrumentation. The animals will be heated during imaging. The imaging procedures do not cause any harm to the animals.
Regarding potential harm caused by tumor growth, animals will be monitored daily by trained people and weighed twice/week. As imaging will be performed regularly, we will be able to detect the tumor burden prior to any symptoms in animal behavior. Based on imaging and examination of the animals, we will record the animals’ well-being via our assessment system. Based on the specific criteria mentioned in the score-sheet, analgesia will be administered as early as necessary and animals will be sacrificed if the objective endpoints are met, prior to developing severe harm.
Reduction:
Using the ultrasound-guided injection technique, the success rate of tumor development is 100%, as our group was able to prove. Thus, we can cut down the number of animals in each experimental group (from 15 to 13 animals).
Etterevaluering
Alle forsøk som er betydelig belastende skal etterevalueres av Mattilsynet
Begrunnelse for etterevalueringen
Forsøkets formål var å studere spredningsmønsteret til systemisk injiserte GBM-celler for å få grunnleggende kunnskap og ny innsikt. Dette kan på sikt bidra til en bedre forståelse av gliomcellenes biologi, og føre til forbedrede behandlingsstrategier. Forsøket ble vurdert som «betydelig belastende» grunnet injeksjoner i hjertet. Dyrene skal våkne fra anestesien, og en slik prosedyre krever særskilt tillatelse fra Mattilsynet, jf Forsøksdyrforskriften § 14, siste ledd. Derfor ble det satt krav om etterevaluering.
Søker angir at alle studier er gjennomført og fullført som planlagt. Forsøket vurderes av søkerne som vellykket, siden forsøket viser det forskjellige metastatiske potensialet til pasientavledede glioblastom-cellelinjer. Bildemetoder, inkludert bioluminescens, MR og PET-CT, ga de nødvendige data.
Forsøket var definert som en pilotstudie og antallet dyr var beregnet med power analyse. Totalt ble det benyttet 103 dyr. Dette er noen færre enn det ble søkt om og gitt tillatelse til. God teknisk utførelse ga gode data slik at færre dyr var nødvendig for å få tilstrekkelig med data.
Belastningsgrad er angitt av søker til å samsvare med det som ble oppgitt i søknaden. Dette vil si «moderat belastende». Dette samsvarer med vurderinger som Mattilsynet har gjort i tiden etter vedtaket om etterevaluering ble skrevet.
Scoreskjemaer ble benyttet, og fungerte tilfredsstillende, men ble utvidet/forbedret med tanke på nye forsøk. En parameter om redebygging ble lagt til. Det ble bekreftet at det er mulig å redusere frekvensen på billedtaking i videre studier. Dette vil redusere stress.
Søker angir at alle studier er gjennomført og fullført som planlagt. Forsøket vurderes av søkerne som vellykket, siden forsøket viser det forskjellige metastatiske potensialet til pasientavledede glioblastom-cellelinjer. Bildemetoder, inkludert bioluminescens, MR og PET-CT, ga de nødvendige data.
Forsøket var definert som en pilotstudie og antallet dyr var beregnet med power analyse. Totalt ble det benyttet 103 dyr. Dette er noen færre enn det ble søkt om og gitt tillatelse til. God teknisk utførelse ga gode data slik at færre dyr var nødvendig for å få tilstrekkelig med data.
Belastningsgrad er angitt av søker til å samsvare med det som ble oppgitt i søknaden. Dette vil si «moderat belastende». Dette samsvarer med vurderinger som Mattilsynet har gjort i tiden etter vedtaket om etterevaluering ble skrevet.
Scoreskjemaer ble benyttet, og fungerte tilfredsstillende, men ble utvidet/forbedret med tanke på nye forsøk. En parameter om redebygging ble lagt til. Det ble bekreftet at det er mulig å redusere frekvensen på billedtaking i videre studier. Dette vil redusere stress.