An optimized protocol for the generation and monitoring of conditional orthotopic lung cancer in the KP mouse model using an adeno-associated virus vector compatible with biosafety level 1.

Deng, Haibin; Ge, Huixiang; Dubey, Christelle; Losmanova, Tereza; Medova, Michaela; Konstantinidou, Georgia; Mutlu, Seyran Mathilde; Birrer, Fabienne Esther; Brodie, Tess Melinda; Stroka, Deborah; Wang, Wenxiang; Peng, Ren-Wang; Dorn, Patrick; Marti, Thomas Michael (2023). An optimized protocol for the generation and monitoring of conditional orthotopic lung cancer in the KP mouse model using an adeno-associated virus vector compatible with biosafety level 1. Cancer immunology, immunotherapy : CII, 72(12), pp. 4457-4470. Springer 10.1007/s00262-023-03542-z

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BACKGROUND

The inducible Kras/p53 lung adenocarcinoma mouse model, which faithfully recapitulates human disease, is routinely initiated by the intratracheal instillation of a virus-based Cre recombinase delivery system. Handling virus-based delivery systems requires elevated biosafety levels, e.g., biosafety level 2 (BSL-2). However, in experimental animal research facilities, following exposure to viral vectors in a BSL-2 environment, rodents may not be reclassified to BSL-1 according to standard practice, preventing access to small animal micro-computed tomography (micro-CT) scanners that are typically housed in general access areas such as BSL-1 rooms. Therefore, our goal was to adapt the protocol so that the Cre-induced KP mouse model could be handled under BSL-1 conditions during the entire procedure.

RESULTS

The Kras-Lox-STOP-Lox-G12D/p53 flox/flox (KP)-based lung adenocarcinoma mouse model was activated by intratracheal instillation of either an adenoviral-based or a gutless, adeno-associated viral-based Cre delivery system. Tumor growth was monitored over time by micro-CT. We have successfully substituted the virus-based Cre delivery system with a commercially available, gutless, adeno-associated, Cre-expressing vector that allows the KP mouse model to be handled and imaged in a BSL-1 facility. By optimizing the anesthesia protocol and switching to a microscope-guided vector instillation procedure, productivity was increased and procedure-related complications were significantly reduced. In addition, repeated micro-CT analysis of individual animals allowed us to monitor tumor growth longitudinally, dramatically reducing the number of animals required per experiment. Finally, we documented the evolution of tumor volume for different doses, which revealed that individual tumor nodules induced by low-titer AAV-Cre transductions can be monitored over time by micro-CT.

CONCLUSION

Modifications to the anesthesia and instillation protocols increased the productivity of the original KP protocol. In addition, the switch to a gutless, adeno-associated, Cre-expressing vector allowed longitudinal monitoring of tumor growth under BSL-1 conditions, significantly reducing the number of animals required for an experiment, in line with the 3R principles.

Item Type:

Journal Article (Original Article)

Division/Institute:

09 Interdisciplinary Units > Microscopy Imaging Center (MIC)
04 Faculty of Medicine > Department of Haematology, Oncology, Infectious Diseases, Laboratory Medicine and Hospital Pharmacy (DOLS) > Clinic of Radiation Oncology
04 Faculty of Medicine > Department of Gastro-intestinal, Liver and Lung Disorders (DMLL) > Clinic of Thoracic Surgery
04 Faculty of Medicine > Department of Gastro-intestinal, Liver and Lung Disorders (DMLL) > Clinic of Pneumology
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > DBMR Forschung Mu35 > Forschungsgruppe Viszeralchirurgie
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > DBMR Forschung Mu35 > Forschungsgruppe Viszeralchirurgie

04 Faculty of Medicine > Pre-clinic Human Medicine > Institute of Pharmacology
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR)
04 Faculty of Medicine > Service Sector > Institute of Pathology > Clinical Pathology
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Forschungsbereich Mu50 > Forschungsgruppe Thoraxchirurgie
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > DBMR Forschung Mu35 > Forschungsgruppe Radio-Onkologie
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > DBMR Forschung Mu35 > Forschungsgruppe Radio-Onkologie

04 Faculty of Medicine > Service Sector > Institute of Pathology
04 Faculty of Medicine > Department of Gastro-intestinal, Liver and Lung Disorders (DMLL) > Clinic of Visceral Surgery and Medicine > Visceral Surgery
04 Faculty of Medicine > Pre-clinic Human Medicine > BioMedical Research (DBMR) > Forschungsbereich Mu50 > Forschungsgruppe Pneumologie (Erwachsene)

Graduate School:

Graduate School for Cellular and Biomedical Sciences (GCB)

UniBE Contributor:

Deng, Haibin, Ge, Huixiang, Dubey, Christelle, Losmanová, Tereza, Medova, Michaela, Konstantinidou, Georgia, Mutlu, Seyran Mathilde, Birrer, Fabienne Esther, Brodie, Tess Melinda, Stroka, Deborah, Peng, Ren-Wang, Dorn, Patrick, Marti, Thomas

Subjects:

600 Technology > 610 Medicine & health
500 Science > 570 Life sciences; biology

ISSN:

1432-0851

Publisher:

Springer

Language:

English

Submitter:

Pubmed Import

Date Deposited:

09 Oct 2023 12:46

Last Modified:

21 Jun 2024 10:17

Publisher DOI:

10.1007/s00262-023-03542-z

PubMed ID:

37796299

Uncontrolled Keywords:

AAV vector BSL-1 Cre-expressing vector Intratracheal instillation Lung adenocarcinoma Mouse model

BORIS DOI:

10.48350/186943

URI:

https://boris.unibe.ch/id/eprint/186943

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