Cancer modeling by Transgene Electroporation in Adult Zebrafish (TEAZ)

Abstract

Transgenic animals are invaluable for modeling cancer genomics, but often require complex crosses of multiple germline alleles to obtain the desired combinations.Zebrafish models have advantages in that transgenes can be rapidly tested by mosaic expression, but these typically lack spatial and temporal control of tumor onset, which limits their utility for the study of tumor progression and metastasis. To overcome these limitations, we have developed a method called Transgene Electroporation in Adult Zebrafish (TEAZ). TEAZ can deliver DNA constructs with promoter elements of interest 2 to drive fluorophores, oncogenes, or CRISPR-Cas9-based mutagenic cassettes in specific cell types. Using TEAZ, we created a highly aggressive melanoma model by expression of BRAF V600E in spatially constrained melanocytes in the context of p53 deficiency and Cas9-mediated inactivation of Rb1. Unlike prior models that take ~4 months to develop, we found that TEAZ leads to tumor onset in ~7 weeks and these develop in fully immunocompetent animals. As the resulting tumors initiated at highly defined locations, we could track their progression via fluorescence and documented deep invasion into tissues and metastatic deposits. TEAZ can be deployed to other tissues and cell types such as the heart with the use of suitable transgenic promoters.The versatility of TEAZ makes it widely accessible for rapid modeling of somatic gene alterations and cancer progression at a scale not achievable in other in vivo systems.

Precise base editing for thein vivostudy of developmental signaling and human pathologies in zebrafish

“…In the future, this will allow to rapidly model and study in vivo combinations of endogenous mutations occurring in specific cancer patients or in genetic disorders caused by somatic mosaicism. Our approach could thus be applied in zebrafish for the precise modelling of complex combinations of cancer-causing mutations in adult animal models as currently possible by transgenic overexpression or somatic gene inactivation (Callahan et al, 2018).…”

Deciphering Melanoma Cell States and Plasticity with Zebrafish Models

“…Live imaging of adult fish was also critical to capture a subpopulation of melanoma cells at the site of residual disease in situ, providing some of the first images of persister cells within an endogenous MRD site (Travnickova et al, 2019) (Figure 2b). These studies illustrate one of the particular strengths of the (Ceol et al, 2011), editing (Ablain et al, 2018), or electroporation (Transgene electroporation in adult zebrafish; TEAZ) (Callahan et al, 2018) allow to spatially and temporally control gene expression.…”

“…For example, using exogenous Endothelin 3 (EDN3) peptide and edn3b mutants, Kim and colleagues showed that EDN3 microenvironment cues control phenotypic switching toward a proliferative and differentiated state in melanoma (Kim et al, 2017). Zhang and colleagues demonstrated that cancer-associated adipocytes directly transfer lipids to zebrafish melanomas to promote growth (Zhang et al, 2018).…”

Genetic Engineering of Zebrafish in Cancer Research

“…Oncogene expression through transgenesis via the injection of transgenes into one-cell stage embryos implies that these oncogenes will be expressed from early embryogenesis, with potentially deleterious effects during development. To bypass this possible limitation, Richard White and colleagues (Memorial Sloan Kettering Cancer Center, New York City, NY, USA) implemented a method called TEAZ (Transgene Electroporation in Adult Zebrafish) to deliver transgenes driving oncogene as well as CRISPR/Cas9 components expression, directly into somatic tissues of adult zebrafish using electroporation [239]. One interesting outcome of this approach is that it is possible to track over the time the spreading of the tumor from its origin, at the electroporation point.…”