PF-114, a Novel Selective Pan BCR/ABL Inhibitor Targets The T315I and Suppress Models Of Advanced Ph+ ALL
Authors: Mian Afsar , Rafiei Anahita , Metodieva Anna , Haberbosch Isabella , Zeifman Alexey , Titov Ilya , Stroylov Victor , Stroganov Oleg , Novikov Fedor , Ottmann Oliver G. , Chilov Ghermes , Ruthardt Martin 
Published: October 2019

Abstract

Background
Targeting BCR/ABL by ABL-directed kinase inhibitors (AKIs) induces long lasting remissions in patients with chronic myeloid leukemia (CML), and short remissions in Ph+ acute lymphatic leukemia (ALL). Notably in advanced Ph+ leukemia resistance attributable to either kinase domain mutations in BCR/ABL or non mutational mechanisms remains the major clinical challenge. With the only exception of Ponatinib, a multitargeted kinase inhibitor, all actually approved AKIs are unable to inhibit the „gatekeeper“ mutation T315I. Ponatinib is unable to overcome non mutational resistance in advanced leukemia and has an unfavorable spectrum of undesired side effects, most likely due to the broad spektrum of kinase inhibition. Thus there is the urgent need for further and more selective therapeutical options in treating therapy-resistant advanced Ph+ leukemia. PF114 is an ATP competitor, which was developed focusing on: i.) targeting all known resistance mutations in BCR/ABL but mainly the T315I; ii.) a higher selectivity as compared to Ponatinib in order to reduce undesired side effects; iii.) the ability to overcome also non mutational resistance in advanced Ph+ leukemia; iv.) an activity not only in chronic phase CML but also advanced Ph+ leukemia, Ph+ ALL or blast crisis CML (BC-CML).

Methods
An toxicity profile of PF-114 as well as its kinase selectivity was investigated. The preclinical evaluation of PF-114 was performed in direct comparison to Ponatinib on golden standard preclinical models of CML and advanced Ph+ leukemia and primary patient-derived long term cultures (PD-LTC) of Ph+ ALL patients. The effects on mutational resistance was investigated i.) on the factor dependence of Ba/F3 cells expressing BCR/ABL or its clinically most relevant resistance mutants (Y253F, E255K, T315I, F317L); ii.) on a PD-LTC of a Ph+ ALL patient harboring the T315I. As models for non mutational resistance we used PD-LTCs from Ph+ ALL patients with different levels of non mutational drug resistance and the SupB15RT, a Ph+ ALL cell line rendered resistant by the exposure to increasing doses of Imatinib and cross-resistant against all approved AKIs. The effects of PF114 in vivo were investigated on the transduction/transplantation model of BCR/ABL- and BCR/ABL-T315I-induced CML-like disease, secondary BCR/ABL-induced murine ALLs as well as on xenografts of PD-LTCs in NSG mice and K562 cells in nude mice.

Results
PF-114 is an orally available AKI, which is more selective than Dasatinib or Ponatinib (number of kinases inhibited at 100 nM of a drug: Nilotinib - 19; PF-114 - 27; Dasatinib - 48; Ponatinib - 80). It was classified as a class 4 - low-toxic - substance (Hodge/Sterner classification). It efficiently inhibited all tested BCR/ABL mutants in cellular and biochemical assays at dosages of 10-100nM and like Ponatinib it suppressed the up-coming of new resistance mutations in a mutation assay in Ba/F3 cells. It also suppressed growth of Ph+ PD-LTC with non mutational resistance as well as the BCR/ABL-T315I-positve PD-LTC in this dosage range. In all models the effect was independent of the presence of either the p210 or the p185 form of BCR/ABL. No effect of PF114 was seen in PD-LTCs of Ph- ALL. Noteworthy PF-114 (50 mg/Kg) prolonged significantly the survival of mice with both BCR/ABL- and BCR/ABL-T315I-driven CML-like disease as compared to Ponatinib (25 mg/Kg). In all in vivo models of advanced leukemia, PF-114 (50 mg/Kg) significantly inhibited leukemia to a similar extent as Ponatinib (25 mg/Kg). Like Ponatinib, PF-114 was unable to overcome the non mutational resistance in SupB15RT.

Conclusions
Our work supports clinical evaluation of PF114 as a pan BCR/ABL inhibitor for treatment not only for chronic phase CML, but also for advanced and resistant Ph+ leukemia such as Ph+ ALL or BC-CML.

Disclosures:
Mian: Fusion Pharma: Research Funding. Zeifman:Fusion Pharma: Employment. Titov:Fusion Pharma: Employment. Stroylov:Fusion Pharma: Employment. Stroganov:Fusion Pharma: Employment. Novikov:Fusion Pharma: Employment. Chilov:Fusion Pharma: Employment. Ruthardt:Fusion Pharma: Research Funding.