Oral Presentation ANZSCDB National Scientific Meeting 2019

Targeting the cytoskeletal regulator AFAP1L1 to control sarcoma migration and invasion. (62995)

Edward Bastow 1 2 3 , Jennifer Beaumont 2 , Sarmed Al-Samerria 2 , Kevin Li 2 , Sien R Tie 2 3 , David J McCarthy 2 3 , Carl Walkley 4 , Evan Ingley 1 2 3
  1. College of Science, Health, Engineering and Education, Murdoch University, Murdoch, WA, Australia
  2. Harry Perkins Institute of Medical Research and Centre for Medical Research, The University of Western Australia, Nedlands, WA, Australia
  3. Centre for Medical Research, The University of Western Australia, Crawley, WA, Australia
  4. Stem Cell Regulation Unit, St Vincent's Institute, Melbourne, VIC, Australia

Migration and invasion are controlled by invadopodia, delivering matrix-degrading enzymes to the invasion interface promoting cancer cell spread into healthy tissue. We have identified a novel pathway that regulates sarcoma cell migration and invasion in vitro via the molecule AFAP1L1 (Actin Filament Associated Protein-1 Like-1) (Oncogene 35:2098-111), the expression of which is strongly associated with malignant osteosarcoma, liposarcoma and leiomyosarcoma (p<0.0001). We now aim to test if we can regulate AFAP1L1-mediated sarcoma cell migration and invasion through expressing targetable fragments of AFAP1L1 (acting as specific activators/inhibitors) in sarcoma cells and undertaking in vitro and in vivo assays. We also aim to see if eliminating AFAP1L1 function in non-sarcoma cells would have any potential harmful effects.

We have generated sub-clonal lines of the human osteosarcoma cells U2-OS, MG-63 and 143B that stably express different regions of AFAP1L1 as eGFP fusions. The cells are then tested in vitro for proliferation, migration and invasion capacity in an IncuCyteZOOM instrument. Cells are then implanted in to both zebrafish embryos and immunocompromised mice to assay for in vivo migration, invasion and metastasis. We have generated AFAP1L1 knockout mice (using CRISPR/Cas9 technology) and characterized the effects on tissue morphology.

Cells expressing a C-terminal fragment of AFAP1L1 show significantly reduced, while those expressing an N-terminal fragment show significantly enhanced migration and invasion in vitro as well as in zebrafish and mouse in vivo assays. Cells expressing the C-terminal fragment secrete less while those expressing the N-terminal fragment secrete more matrix degrading enzymes. Knockout of AFAP1L1 had no major effects on viability, but the mice display effects on muscle and lung morphology.

Targeting the N- and C-terminal regions of AFAP1L1 has potential for disrupting osteosarcoma cell migration and invasion. Fully disrupting AFAP1L1 function in normal cells has no major deleterious effects but does show tissue specific alterations.