Synergistic and targeted therapy with a procaspase-3 activator and temozolomide extends survival in glioma rodent models and is feasible for the treatment of canine malignant glioma patients

Abstract

PurposeGlioblastoma is a deadly brain cancer with a median survival time of ∼15 months. Ionizing radiation plus the DNA alkylator temozolomide (TMZ) is the current standard therapy. PAC-1, a procaspase-3 activating small molecule, is blood-brain barrier penetrant and has previously demonstrated ability to synergize with diverse pro-apoptotic chemotherapeutics. We studied if PAC-1 could enhance the activity of TMZ, and whether addition of PAC-1 to standard treatment would be feasible in spontaneous canine malignant gliomas.Experimental DesignUsing cell lines and online gene expression data, we identified procaspase-3 as a potential molecular target for most glioblastomas. We investigated PAC-1 as a single agent and in combination with TMZ against glioma cells in culture and in orthotopic rodent models of glioma. Three dogs with spontaneous gliomas were treated with an analogous human glioblastoma treatment protocol, with concurrent PAC-1.ResultsProcaspase-3 is expressed in gliomas, with higher gene expression correlating with increased tumor grade and decreased prognosis. PAC-1 is cytotoxic to glioma cells in culture and active in orthotopic rodent glioma models. PAC-1 added to TMZ treatments in cell culture increases apoptotic death, and the combination significantly increases survival in orthotopic glioma models. Addition of PAC-1 to TMZ and radiation was well-tolerated in 3 out of 3 pet dogs with spontaneous glioma, and partial to complete tumor reductions were observed.ConclusionsProcaspase-3 is a clinically relevant target for treatment of glioblastoma. Synergistic activity of PAC-1/TMZ in rodent models and the demonstration of feasibility of the combined regime in canine patients suggest potential for PAC-1 in the treatment of glioblastoma.

Immunohistochemical Characterization of Procaspase-3 Overexpression as a Druggable Target With PAC-1, a Procaspase-3 Activator, in Canine and Human Brain Cancers

“…Despite low expressions of PC-3 identified in normal brain tissues, the observation for histopathologic off-target effects in brain tissues have not been observed in preclinical toxicity studies with healthy rodents receiving orally administered PAC-1 at clinically relevant dosages. More importantly and translationally relevant, PAC-1 has been well-tolerated clinically and neurologically in dogs with spontaneous cancers including gliomas, as well as in humans diagnosed with diverse tumor histologies (34, 47, 48).…”

“…PAC-1 is a blood-brain barrier (BBB) penetrant, small molecule, pro-apoptotic activator of procaspase-3 (PC-3), that possesses favorable pharmacokinetics, tolerability, and synergistic activities when combined with conventional treatment modalities in animal models of glioma, including naturally-occurring brain cancer in pet dogs (34). Mechanistically, PAC-1 activates PC-3 in vitro and in cancer cells through the chelation of inhibitory zinc (35, 36), and based upon PAC-1's binding affinity for zinc ( K d ~40 nM), selective chelation of labile zinc from PC-3 is achieved in the absence of disrupting the function of proteins containing essential zinc ions (37).…”

“…Importantly, cellular sensitivity to apoptosis induction with PAC-1 is associated with the resting cellular PC-3 concentration, and given many malignantly transformed cells have elevated PC-3 levels, PAC-1 therapy allows selective induction of apoptosis of cancerous cells (38). While PAC-1 has demonstrated promising preclinical activity in murine and canine models of glioma (34), whether PC-3 is robustly overexpressed and a conserved therapeutic target in naturally-occurring human and canine brain cancer malignancies has not been systematically evaluated. To address this gap in current knowledge, we sought to investigate PAC-1's broader applicability for the treatment of various brain cancer malignancies, as well as to justify the inclusion of pet dogs as a comparative tumor model for PC-3 activating strategies, we performed large-scale validation of PC-3 as a druggable target across 651 human and canine brain tumor samples, evaluated the prognostic significance of PC-3 expressions in human glial tumors, and tested sensitivity of immortalized glioma cell lines to PAC-1 under biologically achievable conditions.…”

Companion canines: an under-utilised model to aid in translating anti-metastatics to the clinic

“…However, it also may potentiate other therapies. A PAC-1/doxorubicin combination treatment lead to a biologic response in 3/6 dogs with metastatic OSA, and 4/4 dogs with lymphoma [26], whilst a PAC-1/ temozolomide (TMZ) combination achieved biological responses in 3/3 dogs with glioma [27]. Although there are trials at different Institutes within the USA that are currently recruiting canines with lung metastatic OSA to further assess the effectiveness of the PAC-1/doxorubicin combination, Phase I clinical trials using the PAC-1/TMZ combination in humans with high grade gliomas (glioblastoma multiforme or anaplastic astrocytoma after progression following standard first line therapy) have already begun (ClinicalTrials.…”

Of Mice, Dogs, Pigs, and Men: Choosing the Appropriate Model for Immuno-Oncology Research

“…In addition to mifamurtide, other investigational agents that included pet dogs with cancer in the pathway towards investigational new drug designation and human Phase I clinical trials include GS-9219, KPT-335, and PAC-1. 107,[201][202][203][204][205] Given the immune competency of pet dogs with cancer, and underscoring the unique and valuable potential of large animal models in cancer research, the NCI recently launched a request for proposals to support canine clinical studies evaluating the feasibility and activity of immunotherapeutic agents and novel drug combinations such as immune modulators, molecular targeted agents, chemotherapy, and/or radiation. 206 Clinical studies will be accompanied by laboratory correlative studies that seek to describe, characterize, and understand the cellular and molecular mechanisms that determine the antitumor response (or lack of response) in dogs with spontaneous tumors.…”