Acute lymphoblastic leukemia (ALL) is the most frequent pediatric cancer and, despite significant treatment advances, relapsed ALL remains the second leading cause of childhood death. In T-cell ALL, more than 10% of the patients show poor response to frontline therapy and about a third of the patients in remission develop recurrence of their leukemia. Importantly, effective, curative therapies are lacking for children with relapse or refractory disease. Our strategy for T-ALL was to develop agents that target master molecular regulators controlling the activity of multiple complementary, non-recurrent signaling pathways, integrating oncogenic signals and microenvironment cues.
Ref-1/APE1 (Ref-1) is a multi-function protein that exerts redox control of multiple transcription factors (TFs), regulating their DNA binding and downstream transcriptional programs. These include TFs playing important roles in T-cell ALL, namely NF-κB, AP-1 and STAT3, the latter of which we have shown recently to be required for T-cell leukemogenesis. We showed that Ref-1 is expressed by leukemia T-cells in the malignant bone marrow (BM), and that its expression is increased in drug-resistant T-ALL cells. Analyses of multiple leukemia transcriptome databases showed significant increased expression of Ref-1 in T-cell ALL specimens, as well as of other genes in the Ref-1/SET molecular axis. Molecular and functional studies showed that disruption of Ref-1 redox function markedly inhibits leukemia T-cell survival and proliferation, triggering molecular changes promoting cell apoptosis.
We identified three new small molecule chemical entities - APX2007, APX2009 and APX2032, that significantly inhibit the reduction of Ref-1 and the DNA binding of Ref-1-regulated TFs as assessed by EMSA. These compounds (cpds) are significantly more potent than a previously identified Ref-1 redox antagonist, E3330. Functional studies showed that all three APX cpds markedly inhibit leukemia cell survival. Potent inhibition of tumor cell viability was seen in primary cells from ALL patients, relapsed T-ALL, and cells from a murine model of Notch-induced leukemia. Blockade of Ref-1 redox triggers significant leukemia cell apoptosis, and correlates with down-regulation of survival genes regulated by the Ref-1 ‘targets’ STAT3 and NF-kB. Blockade of Ref-1 redox by the APX cpds markedly inhibits the viability of drug- resistant T-ALL cells, with an antitumor efficacy comparable to chemotherapy-sensitive leukemia cells. This is significant since glucocorticoid-resistance is predictive of ALL relapse, and glucocorticoid-resistant leukemia T-cells show reduced sensitivity to inhibitors of other leukemia-associated signaling pathways (as PI3K/Akt, mTOR).
Preliminary safety studies in mice using a clinical formulation demonstrated the systemic administration of the APX cpds do not result in acute adverse reactions or significant hematological toxicities. Importantly, studies in a xenograft model of glucocorticoid-, doxorubicin-resistant human T-ALL showed that treatment with the APX cpds result in significant decrease in leukemia blasts in the peripheral blood and in the BM.
In summary, we developed novel, first-in-class inhibitors of Ref-1, which show acceptable in vivo PK and toxicity, and that potently inhibit T-ALL, including patient's specimens and drug-resistant leukemia T-cells. These new chemical entities target a unique molecular regulator, as Ref-1 redox function controls multiple TFs involved in leukemogenesis and disease progression. Studies are underway to further define the therapeutic efficacy and long-term remission potential of ApeX compounds in animal models of T-ALL, including a model of leukemia recurrence post-frontline chemotherapy. These studies should support the selection of a candidate for development and progression to clinical trials in pediatric patients with refractory, relapsed ALL.
Citation Format: Angelo A. Cardoso, James H. Wikel, Jixin Ding, April M. Reed, Meihua Luo, Mark R. Kelley. First-in-class Ref-1 redox inhibitors for the multipathway targeting of survival signals for relapsed childhood acute lymphoblastic leukemia. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr B32.