Background Tigilanol Tiglate (TT) is a novel small molecule under development for local treatment of solid tumours via intratumoral (I.T.) injection. TT is a protein kinase C (PKC)/C1 domain activator that disrupts tumour vasculature and causes direct oncolysis of tumour cells. Together, these activities lead to haemorrhagic necrosis of injected tumours with enduring ablation of >70% of target tumours in both pre-clinical xenograft/syngeneic mouse models and cutaneous tumours presenting in the veterinary clinic.1–3 TT has completed a Phase I/IIa dose-escalation trial in humans (ACTRN12614000685617), with strong evidence of local anti-cancer efficacy and signs of abscopal effects in some patients.4 However, the underlying mechanism of action (MOA) of TT, together with its immunotherapeutic potential in oncology, is not fully understood.
Methods A combination of microscopy, immunofluorescence, immunoblotting, subcellular fractionation, intracellular ATP assays, LDH release assays and mixed lymphocyte reactions were used to probe the MOA of TT in vitro. TT-mediated damage associated molecular pattern (DAMP) release/externalization was assessed using luciferase (ATP), ELISA (HMGB1), flow cytometry and immunohistochemical (HMGB1, calreticulin) approaches. In vivo experimentation with TT utilized CT-26 and B16-F10 tumor bearing mice. Analysis of DAMP release and immune cell infiltration into TT treated human head and neck tumours (ACTRN12619001407189) was performed by immunohistochemistry.
Results Our data reveal that therapeutic concentrations of TT induce the death of cancer and endothelial cell lines via a pathway involving caspase activation and cleavage of the pore forming protein gasdermin E. TT promotes this mechanism of cell death by interacting with ER membranes, causing an ER stress response that results in loss of mitochondrial membrane potential, ATP depletion, organelle swelling and oncosis/pyroptosis. Treatment of cells with TT also led to the release of damage associated molecular patterns (DAMPs), indicative of an immunogenic cell death (ICD) pathway that also resulted in the generation of tumour-specific T cells in CT-26 tumor bearing mice. Whilst the induction of ICD is largely PKC-independent in vitro, PKC/C1 domain signaling appears necessary for efficacious tumour ablation in vivo. Consistent with our pre-clinical data, immunohistochemical analysis of TT-treated head and neck tumors found that drug stimulated DAMP release/externalisation and the recruitment of immune cells, principally CD8+ T cells, into remnant tumour mass.
Conclusions These data indicate that TT is an oncolytic small molecule with the potential to ablate target tumours and enhance immunotherapy combinations through promoting immune cell infiltration. TT is currently undergoing Phase II trials in head and neck cancer (NCT05234437) and soft tissue sarcoma (NCT05755113).
Boyle GM, D’Souza MMA, Pierce CJ, Adams RA, Cantor AS, Johns JP, Maslovskaya L, Gordon VA, Reddell PW, Parsons PG. Intra-Lesional Injection of the Novel PKC Activator EBC-46 Rapidly Ablates Tumors in Mouse Models. PLOS ONE 2014;9:e108887.
Cullen JK, Boyle GM, Yap PY, Elmlinger S, Simmons JL, Broit N, Johns J, Ferguson B, Maslovskaya LA, Savchenko AI, Mirzayans PM, Porzelle A, Bernhardt PV, Gordon VA, Reddell PW, Pagani A, Appendino G, Parsons PG, Williams CM. Activation of PKC supports the anticancer activity of tigilanol tiglate and related epoxytiglianes. Sci. Rep. 2021;11:207.
De Ridder TR, Campbell JE, Burke-Schwarz C, Clegg D, Elliot EL, Geller S, Kozak W, Pittenger ST, Pruitt JB, Riehl J, White J, Wiest ML, Johannes CM, Morton J, Jones PD, Schmidt PF, Gordon VA, Reddell PW. Randomized controlled clinical study evaluating the efficacy and safety of intratumoral treatment of canine mast cell tumors with tigilanol tiglate (EBC-46). J. Vet. Intern. Med. 2021;35:415–429.
Panizza BJ, de Souza P, Cooper A, Roohullah A, Karapetis CS, Lickliter JD. Phase I dose-escalation study to determine the safety, tolerability, preliminary efficacy and pharmacokinetics of an intratumoral injection of tigilanol tiglate (EBC-46). EBioMedicine. 2019;50:433–441.
Ethics Approval The study obtained ethics approval from the following commitees and boards: Metro South Human Research Ethics Committee, 199 Ipswich Road, Woolloongabba, QLD, 4102, Australia. Ethics approval number: HREC/2019/QMS/54004. Bellberry Limited, 123 Glen Osmond Road, Eastwood, SA 5063, Australia. Ethics approval number: 2019–10-846 (REGIS 2019/ETH13063). Tata Memorial Hospital - Institutional Review Board, IRB Office, Dr. E. Borges Marg, Parel, Mumbai - 400 012, India. Ethics approval number: IEC/1219/3370/001. Tata Medical Center - Institutional Review Board, 14 Major Arterial Road (EW), New Town, Rajarhat, Kolkata - 700 160, India. Ethics approval number: 2019/PHARMA/57/IRB39.
This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See http://creativecommons.org/licenses/by-nc/4.0/.
Statistics from Altmetric.com
If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.