EPIGENETIC PROGRAM

Multi-Targeting Epigenetic Program Overview

The multi-targeting epigenetic program is a result of two collaborations that will provide access to new epigenetic therapeutics for the Company’s oncology pipeline. In November 2015, Aptose entered into a definitive agreement with Moffitt Cancer Center for exclusive global rights to potent, multi-targeting, single-agent inhibitors for the treatment of hematologic and solid tumor cancers. The licensed molecules exhibit single-digit nanomolar potency against the Bromodomain and Extra-Terminal (BET) family members and synergistic, oncogenic kinases. Such multi-targeting BET-Kinase inhibitors are expected to be highly potent in inhibiting the proliferation and survival of certain hematologic and solid tumor cancers that depend on both BET functionality and aberrant kinase activity.

Aptose also entered into an exclusive drug discovery partnership with Laxai Avanti Life Sciences (LALS) to develop multiple epigenetic-based candidates, including the further optimization of compounds licensed from Moffitt Cancer Center. Under the partnership with LALS, Aptose will own global rights to all newly discovered candidates, including all generated intellectual property. Aptose expects a lead clinical candidate(s) to emerge by late 2016.

MA2-014 as a Representative Multi-Targeting BET-Kinase Inhibitor

At the 57th Annual American Society of Hematology (ASH) Meeting in December 2015, collaborators from Moffitt Cancer Center presented preclinical data for one of the representative multi-targeting BET-kinase inhibitors in the collaboration, MA2-014. MA2-014 was rationally designed to inhibit both the bromodomain 4 (BRD4) protein and the Janus kinase 2 (JAK2) for the potential treatment of various hematologic and solid tumor cancers, such as myeloproliferative neoplasms (MPNs). Clinically, JAK2 inhibitors alone have demonstrated the ability to improve symptoms of MPNs, but have not been shown to induce remission. Further, combining a bromodomain inhibitor and a JAK2 inhibitor has been shown to be more effective against MPN cells than JAK2 inhibition alone. This provided rationale to develop a single molecule with potent activity against both the bromodomain family proteins and JAK2.

MA2-014 exhibited similar anti-JAK2 activity as a known JAK2 inhibitor, TG101209, with an approximate ten-fold improvement in anti-BRD4 activity (Figure 1). MA2-014 also demonstrated a ten-fold improvement in inhibition of JAK2-V617F signaling over TG101209 and comparable activity to ruxolitinib. Ruxolitinib is the only FDA approved JAK inhibitor for MPNs. Furthermore, MA2-014 retained its potency against ruxolitinib-resistant cells. In long-term culture assays (Figure 2), JAK2-V617F driven MPN Uke1 cells do not experience resistance to MA2-014 as readily as they do to TG101209 or ruxolitinib. The approach of using single agents to target multiple epigenetic and signaling events may be useful for the treatment of various hematologic and solid tumor cancers with evolved resistance to kinase or BET inhibitors.

1
Figure 1:Relative In Vitro Enzymatic Potency of MA2-014 to Select Bromodomain and Janus Kinase Targets

Compounds were screened against the indicated BET domains (e.g. BRD4-1 indicates the first BET domain of BRD4) and kinases using AlphaScreen Assay for BETs and 32P-labelled kinase reactions (HotSpot Kinase Assay) for kinases by Reaction Biology Corporation.

2
Figure 2: MA2-014 Treatment of Uke1 Cells Exhibit Diminished Emergence of Resistance
Uke1 cells (1×105/mL) were cultured in DMSO, MA2-014, ruxolitinib, or TG101209, each at 50nM. Cells were counted every 2-3 days by trypan blue exclusion, and when cells demonstrated resistance (>=90% growth of DMSO control), the drug concentration was doubled. Data is presented as drug concentration over time, and therefore drug resistance is directly relative to the increasing slope of the data.

BET Inhibition

Members of the Bromodomain and Extra-Terminal (BET) domain proteins, which belong to a family of epigenetic readers, are fundamental for the binding and recognition of acetylated lysine (KAc) residues of histones. They have emerged as potential therapeutic targets in various hematologic and solid tumor cancers that are characterized by dysregulated epigenetic processes. BETs regulate many critical molecular and cellular processes, including transcription, chromatin remodeling, protein scaffolding, and signal transduction, and play key roles in cell growth and survival. Members of the BET protein family (BRD2, BRD3, BRD4, and BRDT) have been implicated in a number of disease pathways. For instance, translocation of BRD4 to the nuclear protein in testis (NUT) locus generates a fusion protein that results in c-Myc overexpression and leads to NUT midline carcinoma (NMC). Because direct inhibition of c-Myc by small molecules is not considered possible, the chemical inhibition of BETs holds potential as a novel therapeutic strategy to improve treatment for various c-Myc-driven cancers. The rationale to develop multi-targeting BET-kinase inhibitors is relevant as single-activity BET and oncogenic kinase inhibitors act synergistically in a variety of cancers, thereby prolonging therapeutic efficacy in the clinic. In addition, multi-targeting BET-kinase inhibitors may be a promising strategy to overcome tumor resistance as cancer cells are aptly able to avoid inhibition by a single-targeting drug.