Development of small molecule RPN13 inhibitors for Treatment of Glioblastoma

Abstract. Glioblastoma (GBM) is an aggressive and deadly brain cancer with a one year median survival due to
the lack of effective targeted treatments. Our goal is to develop a small molecule inhibitor targeting RPN13
protein in the 19S proteasome complex as a new drug for GBM. RPN13 is a proteasome ubiquitin receptor that
recognizes ubiquitin-conjugated proteins that are thus tagged for degradation via the 20S proteasome enzymatic
complex. GBM is highly sensitive to proteasome inhibition due to its aggressive growth and accumulation of
misfolded polyubiquitinated proteins that is insufficiently managed despite increased proteasome levels and
enhanced proteasome activity (i.e. proteotoxic stress). Up Therapeutics judiciously designed drug-like RPN13
inhibitors to overcome limitations in the prototype RPN13 inhibitor RA190 identified at Johns Hopkins University.
We selected Up284 as lead compound based upon our preliminary studies showing drug-like characteristics,
specificity (binding to Cys88 of RPN13), and proteasome inhibition that produces a cytotoxic accumulation of
polyUb proteins, ER stress, and reactive oxygen species. Up284 treatment selectively triggers apoptosis in solid
tumor types including GBM cell lines and oncospheres. A suitable Up284 formulation was selected by measuring
aqueous solubility, stability and on-target activity in mice imaged using a proteasome-dependent in vivo reporter.
Up284 can be administered i.v. or orally, and it penetrates through the BBB. As a proof of concept for treatment
of GBM, Up284 demonstrated on-target activity in intracranial syngeneic GL261 tumor model by inhibiting
proteasome function in tumor tissues. Up284 also regressed tumors in syngeneic, xenograft and spontaneous
models of ovarian cancer with acceptable safety profile. Pilot toxicological data indicated Up284 is relatively well
tolerated with no evidence of behavioral abnormalities and normal CBC and blood chemistry panel. Given the
potential of Up284 as a new cancer drug and GBM as its lead indication, we propose:
Goal1a. Determine Up284 dose required in the brain to kill GBM oncospheres (1-3 months): Renewable
oncosphere formation in culture is a defining characteristic of certain brain tumor initiating cells. Up284 activity
against the growth of GBM oncospheres in the presence and absence of an in vitro BBB layer will be tested.
Goal1b. Up284 plasma and tissue distribution studies after IV vs Oral delivery in male andamp; female CD1 mice
(3-6 months): Milestones: Identify GBM therapeutic levels needed in the brain and how to deliver Up284 to
achieve them. Goal2. Efficacy of Up284 against the growth of orthotopic GBM oncospheres in male andamp;
female nude mice (6-12 months). We will test Up284 activity against an intracranial orthotopic transplantation
model using firefly luciferase-expressing Br23C oncospheres in nude female mice bearing intracranial Br23C
oncospheres or nude male mice bearing JHH-27 oncospheres, a temozolamide resistant model. Milestones:
Demonstrate that Up284 provides RECIST-defined complete response (CR) or partial response (PR) against
tumor growth using oncosphere-derived GBM models in mouse brain.Project Narrative. The objective of this STTR phase I project is to develop a small molecule
inhibitor targeting ubiquitin proteasome receptor RPN13 as a new targeted drug for treatment of
patients with the aggressive and rapidly fatal brain cancer glioblastoma.