Victor M. Rivera

AP24534, a pan-BCR-ABL inhibitor for chronic myeloid leukemia, potently inhibits the T315I mutant and overcomes mutation-based resistance.

Inhibition of BCR-ABL by imatinib induces durable responses in many patients with chronic myeloid leukemia (CML), but resistance attributable to kinase domain mutations can lead to relapse and a switch to second-line therapy with nilotinib or dasatinib. Despite three approved therapeutic options, the cross-resistant BCR-ABL(T315I) mutation and compound mutants selected on sequential inhibitor therapy remain major clinical challenges. We report design and preclinical evaluation of AP24534, a potent, orally available multitargeted kinase inhibitor active against T315I and other BCR-ABL mutants. AP24534 inhibited all tested BCR-ABL mutants in cellular and biochemical assays, suppressed BCR-ABL(T315I)-driven tumor growth in mice, and completely abrogated resistance in cell-based mutagenesis screens. Our work supports clinical evaluation of AP24534 as a pan-BCR-ABL inhibitor for treatment of CML.

A Phase 2 Trial of Ponatinib in Philadelphia Chromosome–Positive Leukemias

BACKGROUND Ponatinib is a potent oral tyrosine kinase inhibitor of unmutated and mutated BCR-ABL, including BCR-ABL with the tyrosine kinase inhibitor-refractory threonine-to-isoleucine mutation at position 315 (T315I). We conducted a phase 2 trial of ponatinib in patients with chronic myeloid leukemia (CML) or Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph-positive ALL). METHODS We enrolled 449 heavily pretreated patients who had CML or Ph-positive ALL with resistance to or unacceptable side effects from dasatinib or nilotinib or who had the BCR-ABL T315I mutation. Ponatinib was administered at an initial dose of 45 mg once daily. The median follow-up was 15 months. RESULTS Among 267 patients with chronic-phase CML, 56% had a major cytogenetic response (51% of patients with resistance to or unacceptable side effects from dasatinib or nilotinib and 70% of patients with the T315I mutation), 46% had a complete cytogenetic response (40% and 66% in the two subgroups, respectively), and 34% had a major molecular response (27% and 56% in the two subgroups, respectively). Responses were observed regardless of the baseline BCR-ABL kinase domain mutation status and were durable; the estimated rate of a sustained major cytogenetic response of at least 12 months was 91%. No single BCR-ABL mutation conferring resistance to ponatinib was detected. Among 83 patients with accelerated-phase CML, 55% had a major hematologic response and 39% had a major cytogenetic response. Among 62 patients with blast-phase CML, 31% had a major hematologic response and 23% had a major cytogenetic response. Among 32 patients with Ph-positive ALL, 41% had a major hematologic response and 47% had a major cytogenetic response. Common adverse events were thrombocytopenia (in 37% of patients), rash (in 34%), dry skin (in 32%), and abdominal pain (in 22%). Serious arterial thrombotic events were observed in 9% of patients; these events were considered to be treatment-related in 3%. A total of 12% of patients discontinued treatment because of an adverse event. CONCLUSIONS Ponatinib had significant antileukemic activity across categories of disease stage and mutation status. (Funded by Ariad Pharmaceuticals and others; PACE ClinicalTrials.gov number, NCT01207440 .).

Ponatinib in Refractory Philadelphia Chromosome–Positive Leukemias

BACKGROUND Resistance to tyrosine kinase inhibitors in patients with chronic myeloid leukemia (CML) and Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph-positive ALL) is frequently caused by mutations in the BCR-ABL kinase domain. Ponatinib (AP24534) is a potent oral tyrosine kinase inhibitor that blocks native and mutated BCR-ABL, including the gatekeeper mutant T315I, which is uniformly resistant to tyrosine kinase inhibitors. METHODS In this phase 1 dose-escalation study, we enrolled 81 patients with resistant hematologic cancers, including 60 with CML and 5 with Ph-positive ALL. Ponatinib was administered once daily at doses ranging from 2 to 60 mg. Median follow-up was 56 weeks (range, 2 to 140). RESULTS Dose-limiting toxic effects included elevated lipase or amylase levels and pancreatitis. Common adverse events were rash, myelosuppression, and constitutional symptoms. Among Ph-positive patients, 91% had received two or more approved tyrosine kinase inhibitors, and 51% had received all three approved tyrosine kinase inhibitors. Of 43 patients with chronic-phase CML, 98% had a complete hematologic response, 72% had a major cytogenetic response, and 44% had a major molecular response. Of 12 patients who had chronic-phase CML with the T315I mutation, 100% had a complete hematologic response and 92% had a major cytogenetic response. Of 13 patients with chronic-phase CML without detectable mutations, 100% had a complete hematologic response and 62% had a major cytogenetic response. Responses among patients with chronic-phase CML were durable. Of 22 patients with accelerated-phase or blast-phase CML or Ph-positive ALL, 36% had a major hematologic response and 32% had a major cytogenetic response. CONCLUSIONS Ponatinib was highly active in heavily pretreated patients with Ph-positive leukemias with resistance to tyrosine kinase inhibitors, including patients with the BCR-ABL T315I mutation, other mutations, or no mutations. (Funded by Ariad Pharmaceuticals and others; ClinicalTrials.gov number, NCT00660920.).

Therapeutic strategies to overcome crizotinib resistance in non-small cell lung cancers harboring the fusion oncogene EML4-ALK

The echinoderm microtubule-associated protein-like 4 (EML4)-anaplastic lymphoma kinase (ALK) fusion oncogene represents a molecular target in a small subset of non-small cell lung cancers (NSCLCs). This fusion leads to constitutive ALK activation with potent transforming activity. In a pivotal phase 1 clinical trial, the ALK tyrosine kinase inhibitor (TKI) crizotinib (PF-02341066) demonstrated impressive antitumor activity in the majority of patients with NSCLC harboring ALK fusions. However, despite these remarkable initial responses, cancers eventually develop resistance to crizotinib, usually within 1 y, thereby limiting the potential clinical benefit. To determine how cancers acquire resistance to ALK inhibitors, we established a model of acquired resistance to crizotinib by exposing a highly sensitive EML4-ALK–positive NSCLC cell line to increasing doses of crizotinib until resistance emerged. We found that cells resistant to intermediate doses of crizotinib developed amplification of the EML4-ALK gene. Cells resistant to higher doses (1 μM) also developed a gatekeeper mutation, L1196M, within the kinase domain, rendering EML4-ALK insensitive to crizotinib. This gatekeeper mutation was readily detected using a unique and highly sensitive allele-specific PCR assay. Although crizotinib was ineffectual against EML4-ALK harboring the gatekeeper mutation, we observed that two structurally different ALK inhibitors, NVP-TAE684 and AP26113, were highly active against the resistant cancer cells in vitro and in vivo. Furthermore, these resistant cells remained highly sensitive to the Hsp90 inhibitor 17-AAG. Thus, we have developed a model of acquired resistance to ALK inhibitors and have shown that second-generation ALK TKIs or Hsp90 inhibitors are effective in treating crizotinib-resistant tumors harboring secondary gatekeeper mutations.

Phase I Trial of the Novel Mammalian Target of Rapamycin Inhibitor Deforolimus (AP23573; MK-8669) Administered Intravenously Daily for 5 Days Every 2 Weeks to Patients With Advanced Malignancies

PURPOSE This phase I trial was conducted to determine the safety, tolerability, pharmacokinetics, and pharmacodynamics of deforolimus (previously known as AP23573; MK-8669), a nonprodrug rapamycin analog, in patients with advanced solid malignancies. PATIENTS AND METHODS Patients were treated using an accelerated titration design with sequential escalating flat doses of deforolimus administered as a 30-minute intravenous infusion once daily for 5 consecutive days every 2 weeks (QDx5) in a 28-day cycle. Safety, pharmacokinetic, pharmacodynamic, and tumor response assessments were performed. RESULTS Thirty-two patients received at least one dose of deforolimus (3 to 28 mg/d). Three dose-limiting toxicity events of grade 3 mouth sores were reported. The maximum-tolerated dose (MTD) was 18.75 mg/d. Common treatment-related adverse events included reversible mouth sores and rash. Whole-blood clearance increased with dose. Pharmacodynamic analyses demonstrated mammalian target of rapamycin inhibition at all dose levels. Four patients (one each with non-small-cell lung cancer, mixed müllerian tumor [carcinosarcoma], renal cell carcinoma, and Ewing sarcoma) experienced confirmed partial responses, and three additional patients had minor tumor regressions. CONCLUSION The MTD of this phase I trial using an accelerated titration design was determined to be 18.75 mg/d. Deforolimus was well tolerated and showed encouraging antitumor activity across a broad range of malignancies when administered intravenously on the QDx5 schedule. On the basis of these overall results, a dose of 12.5 mg/d is being evaluated in phase II trials.

A Phase 2 Clinical Trial of Deforolimus (AP23573, MK-8669), a Novel Mammalian Target of Rapamycin Inhibitor, in Patients with Relapsed or Refractory Hematologic Malignancies

Purpose: Deforolimus (AP23573), a novel non-prodrug rapamycin analogue, inhibits the mammalian target of rapamycin, a downstream effector of the phosphatidylinositol 3-kinase/Akt and nutrient-sensing pathways. A phase 2 trial was conducted to determine the efficacy and safety of single-agent deforolimus in patients with relapsed or refractory hematologic malignancies. Experimental Design: Eligible patients were assigned to one of five disease-specific, parallel cohorts and given 12.5 mg deforolimus as a 30-minute infusion once daily for 5 days every 2 weeks. A Simon two-stage design was used for each cohort. Safety, pharmacokinetics, pharmacodynamics, and antitumor response were assessed. Results: Fifty-five patients received deforolimus as follows: cohort 1 23 acute myelogenous leukemia, two myelodysplastic syndrome and one chronic myelogenous leukemia in nonlymphoid blast phase; cohort 2, one acute lymphocytic leukemia; cohort 3, nine agnogenic myeloid metaplasia; cohort 4, eight chronic lymphocytic leukemia; cohort 5, nine mantle cell lymphoma and two T-cell leukemia/lymphoma. Most patients were heavily pretreated. Of the 52 evaluable patients, partial responses were noted in five (10%), two of seven agnogenic myeloid metaplasia and three of nine mantle cell lymphoma. Hematologic improvement/stable disease was observed in 21 (40%). Common treatment-related adverse events, which were generally mild and reversible, were mouth sores, fatigue, nausea, and thrombocytopenia. Decreased levels of phosphorylated 4E-BP1 in 9 of 11 acute myelogenous leukemia/myelodysplastic syndrome patients after therapy showed mammalian target of rapamycin inhibition by deforolimus. Conclusions: Deforolimus was well-tolerated in patients with heavily pretreated hematologic malignancies, and antitumor activity was observed. Further investigation of deforolimus alone and in combination with other therapeutic agents is warranted in patients with selected hematologic malignancies.

Discovery of 3-[2-(imidazo[1,2-b]pyridazin-3-yl)ethynyl]-4-methyl-N-{4-[(4-methylpiperazin-1-yl)methyl]-3-(trifluoromethyl)phenyl}benzamide (AP24534), a potent, orally active pan-inhibitor of breakpoint cluster region-abelson (BCR-ABL) kinase including the T315I gatekeeper mutant.

In the treatment of chronic myeloid leukemia (CML) with BCR-ABL kinase inhibitors, the T315I gatekeeper mutant has emerged as resistant to all currently approved agents. This report describes the structure-guided design of a novel series of potent pan-inhibitors of BCR-ABL, including the T315I mutation. A key structural feature is the carbon-carbon triple bond linker which skirts the increased bulk of Ile315 side chain. Extensive SAR studies led to the discovery of development candidate 20g (AP24534), which inhibited the kinase activity of both native BCR-ABL and the T315I mutant with low nM IC(50)s, and potently inhibited proliferation of corresponding Ba/F3-derived cell lines. Daily oral administration of 20g significantly prolonged survival of mice injected intravenously with BCR-ABL(T315I) expressing Ba/F3 cells. These data, coupled with a favorable ADME profile, support the potential of 20g to be an effective treatment for CML, including patients refractory to all currently approved therapies.

Ponatinib (AP24534), a Multitargeted Pan-FGFR Inhibitor with Activity in Multiple FGFR-Amplified or Mutated Cancer Models

Members of the fibroblast growth factor receptor family of kinases (FGFR1–4) are dysregulated in multiple cancers. Ponatinib (AP24534) is an oral multitargeted tyrosine kinase inhibitor being explored in a pivotal phase II trial in patients with chronic myelogenous leukemia due to its potent activity against BCR-ABL. Ponatinib has also been shown to inhibit the in vitro kinase activity of all four FGFRs, prompting us to examine its potential as an FGFR inhibitor. In Ba/F3 cells engineered to express activated FGFR1–4, ponatinib potently inhibited FGFR-mediated signaling and viability with IC50 values <40 nmol/L, with substantial selectivity over parental Ba/F3 cells. In a panel of 14 cell lines representing multiple tumor types (endometrial, bladder, gastric, breast, lung, and colon) and containing FGFRs dysregulated by a variety of mechanisms, ponatinib inhibited FGFR-mediated signaling with IC50 values <40 nmol/L and inhibited cell growth with GI50 (concentration needed to reduce the growth of treated cells to half that of untreated cells) values of 7 to 181 nmol/L. Daily oral dosing of ponatinib (10–30 mg/kg) to mice reduced tumor growth and inhibited signaling in all three tumor models examined. Importantly, the potency of ponatinib in these models is similar to that previously observed in BCR-ABL–driven models and plasma levels of ponatinib that exceed the IC50 values for FGFR1–4 inhibition can be sustained in patients. These results show that ponatinib is a potent pan-FGFR inhibitor and provide strong rationale for its evaluation in patients with FGFR-driven cancers. Mol Cancer Ther; 11(3); 690–9. ©2012 AACR.

Phase II Study of the Mammalian Target of Rapamycin Inhibitor Ridaforolimus in Patients With Advanced Bone and Soft Tissue Sarcomas

PURPOSE Ridaforolimus is an inhibitor of mammalian target of rapamycin, an integral component of the phosphatidyl 3-kinase/AKT signaling pathway, with early evidence of activity in sarcomas. This multicenter, open-label, single-arm, phase II trial was conducted to assess the antitumor activity of ridaforolimus in patients with distinct subtypes of advanced sarcomas. PATIENTS AND METHODS Patients with metastatic or unresectable soft tissue or bone sarcomas received ridaforolimus 12.5 mg administered as a 30-minute intravenous infusion once daily for 5 days every 2 weeks. The primary end point was clinical benefit response (CBR) rate (complete response or partial response [PR] or stable disease ≥ 16 weeks). Safety, progression-free survival (PFS), overall survival (OS), time to progression, and duration of response were also evaluated. RESULTS A total of 212 patients were treated in four separate histologic cohorts. In this heavily pretreated population, 61 patients (28.8%) achieved CBR. Median PFS was 15.3 weeks; median OS was 40 weeks. Response Evaluation Criteria in Solid Tumors (RECIST) confirmed response rate was 1.9%, with four patients achieving confirmed PR (two with osteosarcoma, one with spindle cell sarcoma, and one with malignant fibrous histiocytoma). Archival tumor protein markers analyzed were not correlated with CBR. Related adverse events were generally mild or moderate and consisted primarily of stomatitis, mucosal inflammation, mouth ulceration, rash, and fatigue. CONCLUSION Single-agent ridaforolimus in patients with advanced and pretreated sarcomas led to PFS results that compare favorably with historical metrics. A phase III trial based on these data will further define ridaforolimus activity in sarcomas.

Structural Mechanism of the Pan-BCR-ABL Inhibitor Ponatinib (AP24534): Lessons for Overcoming Kinase Inhibitor Resistance.

The BCR‐ABL inhibitor imatinib has revolutionized the treatment of chronic myeloid leukemia. However, drug resistance caused by kinase domain mutations has necessitated the development of new mutation‐resistant inhibitors, most recently against the T315I gatekeeper residue mutation. Ponatinib (AP24534) inhibits both native and mutant BCR‐ABL, including T315I, acting as a pan‐BCR‐ABL inhibitor. Here, we undertook a combined crystallographic and structure–activity relationship analysis on ponatinib to understand this unique profile. While the ethynyl linker is a key inhibitor functionality that interacts with the gatekeeper, virtually all other components of ponatinib play an essential role in its T315I inhibitory activity. The extensive network of optimized molecular contacts found in the DFG‐out binding mode leads to high potency and renders binding less susceptible to disruption by single point mutations. The inhibitory mechanism exemplified by ponatinib may have broad relevance to designing inhibitors against other kinases with mutated gatekeeper residues.