Poster Presentation 31st Lorne Cancer Conference 2019

βIII-Tubulin is a Brake for External Cell-Death Signalling in Pancreatic Cancer (#370)

George Sharbeen 1 , John Kokkinos 1 , Joshua McCarroll 2 3 , Cyrille Boyer 3 , Anouschka Akerman 1 , Janet Youkhana 1 , Tom Davis 4 , David Goldstein 1 , Phoebe Phillips 1 3
  1. Pancreatic Cancer Translational Research Group, Lowy Cancer Research Centre, UNSW Sydney, Sydney, NSW, Australia
  2. Children's Cancer Institute, Lowy Cancer Research Centre, UNSW Sydney, Sydney, NSW, Australia
  3. Australian Centre for NanoMedicine, UNSW Sydney, Sydney, NSW, Australia
  4. ARC Centre of Excellence in Convergent Bio-Nano Science and Technology, Monash Institute of Pharmaceutical Sciences, Monash University, Melbourne, VIC, Australia

Background: Pancreatic cancer (PC) requires new therapeutic strategies. We identified that inhibition of βIII-tubulin (shRNA) in PC cells decreased tumour growth and metastases in mice [1]. However, since there are no inhibitors of βIII-tubulin, we developed a nanoparticle to deliver βIII-tubulin siRNA to mouse pancreatic tumours [2].

Aims: (1) To assess the effect of therapeutic inhibition of βIII-tubulin using nanoparticle-siRNA on pancreatic tumour growth in mice. (2) To investigate the pro-apoptotic pathways controlled by βIII-tubulin in PC cells.

Methods: In vivo: Mice with orthotopic MiaPaCa-2 tumours (n=10/group) were treated (5-weeks post-implantation) with nanoparticle+βIII-tubulin-siRNA or control-siRNA for 3.5-weeks. Tumour volume was measured and cleaved caspase-8 assessed in tumours. In vitro: PC cells (MiaPaCa-2, PANC1) were treated ±βIII-tubulin-siRNA ±caspase-8 or 9 inhibitors and caspase-8/9 activities and apoptosis measured. Sensitivity to extrinsic-apoptosis inducers (TRAIL, TNFα, FasL) were tested (proliferation, apoptosis).

Results: Nanoparticle+βIII-tubulin-siRNA reduced PC tumour growth (103±17.5mm3; p<0.05) versus controls (194.9±31.2mm3). Silencing βIII-tubulin in PC cells significantly increased the activity of caspase-8 (MiaPaCa-2: 53±13% increase vs control, *p<0.05) and caspase-9 (45±9% increase vs control, *p<0.01). However, βIII-tubulin knockdown-induced apoptosis was only blocked by inhibiting caspase-8. Furthermore, knockdown of βIII-tubulin combined with TRAIL, TNFα or FasL increased apoptosis, and reduced cell proliferation. Inhibition of βIII-tubulin in vivo also increased caspase-8 cleavage in PC tumours.

Conclusions: Nanomedicine/siRNA inhibition of βIII-tubulin represents a potential new class of PC therapeutics. Our novel results demonstrated βIII-tubulin-siRNA in PC cells activates extrinsic-apoptosis and sensitises to inducers of extrinsic-apoptosis. βIII-tubulin inhibition thus represents a three-pronged attack on PC cells by changing the way cells respond to (1) a chemotherapeutic (TRAIL), (2) immune cytokines (FasL and TNFα) and (3) pancreatic stellate cell secretions (TNFα).

References:

[1] McCarroll, J.A., et al. Oncotarget, 2015. 6(4): p. 2235-49.

[2] Teo, J., et al. Biomacromolecules, 2016. 17(7): p. 2337-51.