Axitinib (Inlyta, Pfizer Inc), already approved for the second-line treatment of advanced renal cell carcinoma (RCC), has surprisingly been shown to inhibit the BCR-ABL1 (T315I) mutation in cells of patients with chronic myelogenous leukemia (CML) and Philadelphia chromosome–positive B-cell acute lymphoblastic leukemia (B-ALL). This mutation renders these diseases resistant to most current treatments, so axitinib offers hope as a new therapy.
This was an unexpected finding and makes axitinib the first vascular endothelial growth factor receptor (VEGFR) inhibitor to show activity in BCR-ABL (T315I) CML and B-ALL.
The new observations were reported in a study published onlineFebruary 9 in Nature and resulted from a collaboration between researchers from the Institute for Molecular Medicine Finland (FIMM) at the University of Helsinki, the Helsinki University Central Hospital Comprehensive Cancer Center, and Pfizer's research team.
Although the study was research oriented, the results also indicated that one patient with BCR-ABL (T315I) CML was successfully treated with axitinib after experiencing disease progression on approved therapies. Now clinical trials are planned.
With a view to personalizing cancer care, the research group at the FIMM uses its drug sensitivity and resistance testing (DSRT) to profile patients' primary cancer samples and feeds the information to the clinician with a potential to help patients, Krister Wennerberg, PhD, FIMM-EMBL group leader at the Institute for Molecular Medicine Finland, University of Helsinki, and one of the senior authors of the article, told Medscape Medical News. That is how one patient was successfully treated with axitinib.
This finding has potential clinical applicability. It offers hope for patients with BCR-ABL1 (T315I) CML whose disease is resistant to treatment with the approved selective BCR-ABL tyrosine kinase inhibitors (TKIs), imatinib (Gleevec, Novartis Pharmaceuticals Corporation), dasatinib (Sprycel, Bristol-Myers Squibb Company), nilotinib (Tasigna, Novartis Pharmaceuticals Corporation), and bosutinib (Bosulif, Pfizer Inc).
The only TKI inhibitor active in CML with BCR-ABL1 (T315I) mutation, ponatinib (Iclusig, ARIAD Pharmaceuticals, Inc), has vascular side effects that prompted a temporary withdrawal from the market, although it is now back on the market with warnings.
Jorge Cortes, MD, the D. B. Lane Cancer Research Distinguished Professor for Leukemia Research, Department of Leukemia, Division of Cancer Medicine, the University of Texas MD Anderson Cancer Center, Houston, agreed that the new findings suggest potential clinical use for axitinib in these patients.
"These are very interesting observations," Dr Cortes told Medscape Medical News. "We have a lot of treatment options for patients with CML, and for a large group of patients, we are doing well. However, ponatinib is the only option for some patients with the T315I mutation; because it may have serious side effects in some patients, identifying another drug for these patients is useful," he added.
Drug Sensitivity and Resistance Testing
The Finnish researchers used cancer cells ex vivo from patients with T315I CML and B-ALL who had developed resistance to currently available treatments. Using the DSRT developed at FIMM, they determined the growth response of the cells to a panel of 252 drugs — 125 already approved, and 127 investigative.
The growth of T315I cancer cells was not inhibited by imatinib, nilotinib, and dasatinib but was inhibited by ponatinib and drugs that targeted molecules downstream of BCR-ABL signaling — as expected. The big surprise was that growth of the cells was inhibited by axitinib, a VEGFR inhibitor. Indeed, axitinib was the only VEGFR inhibitor shown to inhibit the growth of these cancer cells ex vivo.
In addition, the kinase activity of the T315I mutation was inhibited at a very low concentration of axitinib — 100 picomolar — which compared favorably with the concentration required to inhibit the tyrosine kinase activity of VEGFR (20 picomolar). The activity of wild-type BCR-ABL kinase was inhibited with a significantly higher concentration of axitinib (3800 picomolar). These observations suggested a specific and potent inhibition of BCR-ABL (T315I).
This screening method is a key component of FIMM's Individualized Systems Medicine strategy. The drug panel covers all approved and many emerging cancer therapeutics and thus enables an individualized selection of potentially effective therapies for leukemia patients, explained Dr Wennerberg in a FIMM press release.
Preclinical and Clinical Validation
The researchers next looked for preclinical evidence to confirm these results. They examined the biochemical activity of BCR-ABL1 and cell proliferation in a strain of murine cells called pro-B Ba/F3 cells after wild-type BCR-ABL1 or BCR-ABL1 (T315I) was stably introduced in the cells through transfection.
In a growth proliferation assay, cells with the BCR-ABL1 (T315I) were shown to be growth inhibited with axitinib in a dose-dependent manner much in the same way as with ponatinib. In addition, the biochemical activity through which tyrosine residues in BCR are "autophosphorylated" — an activity important for cell growth — was potently inhibited in these cells.
The researchers went back to the clinic and obtained mononuclear cells from the bone marrow of patients with CML — two did not have the T315I mutations, and three did. When the growth of these cells was compared with that of cells from healthy individuals and from patients with acute myeloid leukemia, growth of cells from patients with T315I mutations was again potently inhibited.
Ex vivo observations of one patient's cells showed potent growth inhibition by axitinib. All lines of treatment for this patient had been exhausted. This patient was then treated with axitinib (5 mg twice daily, the dose approved for use in advanced RCC) for 2 weeks and showed a rapid clearance of cells expressing T315I cells.
Defining the Mechanism
But how does one explain the unexpected inhibition of BCR-ABL1 (T315I) by a VEGFR inhibitor?
The structure-based drug discovery expertise at Pfizer provided the answer. The group looked at co-crystals of axitinib with VEFGR, ABL, and ABL (T315I). When compared with the binding of other BCR-ABL kinase inhibitors (eg, imatinib, dasatinib), they showed that axitinib filled different binding spaces, making the binding to the T315I protein tighter.
In addition, axitinib was shown to bind wild-type ABL1 in its inactive conformation in much the same way it does VEGFR. In contrast, the axitinib-ABL1 (T315I) complex had an active or open conformation.
"If you think of the targeted protein as a lock into which the cancer drug fits in as a key, the resistant protein changes in such a way that we need a different key," senior author Brion W. Murray, PhD, Pfizer Worldwide Research and Development, commented in a press release. In the case of axitinib, it acts as two distinct keys — one for renal cell carcinoma and one for leukemia, he explained.
The x-ray crystallographic coordinates and structure factor files for axitinib-ABL1 (T315I) and axitinib-wild-type ABL1 complexes have been deposited in the Protein Data Bank under accession numbers 4TWP and 4WA9, respectively, Dr Murray told Medscape Medical News.
Dr Murray explained the collaborative nature of the work to Medscape Medical News. The FIMM screening was a valuable asset because it is done with bona fide patient samples and provided strong evidence of clinical relevance. Researchers at Pfizer contributed biochemical data, cellular studies with engineered models, and structural biology components to the study, he elucidated.
The study provides new molecular insights into how cancer-causing kinases can be blocked, and it highlights the power and value of collaboration between Pfizer and independent academic research groups, Dr Murray said.
What's Next for Axitinib?
Because axitinib is already used to treat cancer, its safety is known, and a formal exploration of its clinical utility in drug-resistant leukemia can now be done in a fast-track mode, commented coauthor Kimmo Porkka, MD, PhD, head of hematology at Helsinki University Central Hospital Comprehensive Cancer Center. "Thus, the normally very long path from lab bench to bedside is now significantly shortened," he said in a press release.
The efficacy in the laboratory is attractive, and we must now think about bringing the findings into the clinic, Dr Cortes told Medscape Medical News.
Dr Cortes said he is involved in the development of a phase II study to test the safety and efficacy of axitinib in patients with T315I CML. Because the drug is already approved for advanced RCC, we do not need a full phase 1 study, but there will be sufficient safety measures incorporated into the protocol, he said.
In summarizing their findings, the researchers said: "Our results on axitinib in T315I-mutant CML provide a powerful example of how unbiased drug sensitivity testing of patient-derived cancer cells can lead to the discovery of an unexpected drug-target interaction with mechanistic, structural, and clinical implications."
"Our study highlights the value of drug repositioning, that is, searching for novel indications for existing, emerging, and abandoned drugs," they concluded.
Some authors of the publication, including Dr Murray, are employees of Pfizer Inc. Dr Wennerberg receives investigator-initiated research funding from Pfizer for a separate project. Dr Cortes is involved in designing Pfizer's phase II study for axitinib in T315I CML and has received research support from and is a consultant (compensated) for several pharmaceutical companies.
Nature. Published online February 9, 2015. Abstract
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