Reversal of acquired drug resistance in FLT3-mutated acute myeloid leukemia cells via distinct drug combination strategies.

FMS-like tyrosine kinase-3 (FLT3) internal tandem duplication (FLT3-ITD) mutations are common in patients with acute myeloid leukemia (AML). These patients regularly develop resistance to FLT3 inhibitors suggesting that targeted combination drug strategies are needed to enhance AML therapy efficacy. Acquired point mutations of FLT3-ITD gene were screened using cDNA-based sequencing approach in vitro sorafenib-resistant cells, which were developed by long-term exposure of Ba/F3-ITD to increasing doses of sorafenib, and in FLT3-ITD mutated AML patients, who developed relapse following sorafenib therapy. Drug effects (e.g., proliferation inhibition, apoptosis induction, and changes in signal transduction protein expression) were assessed in AML cells harboring the point mutations in vitro and in FLT3-ITD-mutated AML patient samples. We identified several acquired point mutations in the tyrosine kinase domains (TKD) of the FLT3 gene in sorafenib-resistant murine leukemia cell line carrying human FLT3-ITD mutations, which were also detected in two of four sorafenib-resistant patient samples. Engineering these point mutations into Ba/F3-ITD cells generated sublines that demonstrated varying degrees of sorafenib [a type II tyrosine kinase inhibitor (TKI)] resistance. A similar pattern of resistance could be observed by exposing these sublines to the other type II TKIs AC220 and MLN518. However, these sublines retained sensitivity to the type I TKIs PKC412 or crenolanib. The combination of crenolanib with sorafenib demonstrated marked cytotoxic effects in all of the sorafenib-resistant sublines. These combination strategies could be clinically important in reversing acquired resistance to FLT3 inhibition in AML.

Zhang W ，Gao C ，Konopleva M ，Chen Y ，Jacamo RO ，Borthakur G ，Cortes JE ，Ravandi F ，Ramachandran A ，Andreeff M ... -  《-》

Crenolanib is active against models of drug-resistant FLT3-ITD-positive acute myeloid leukemia.

Zimmerman EI ，Turner DC ，Buaboonnam J ，Hu S ，Orwick S ，Roberts MS ，Janke LJ ，Ramachandran A ，Stewart CF ，Inaba H ，Baker SD ... -  《-》

Emergence of polyclonal FLT3 tyrosine kinase domain mutations during sequential therapy with sorafenib and sunitinib in FLT3-ITD-positive acute myeloid leukemia.

To evaluate the clinical activity of sequential therapy with sorafenib and sunitinib in FMS-like tyrosine kinase 3 (FLT3)-internal tandem duplication (ITD)-positive acute myelogenous leukemia (AML) and monitor the emergence of secondary FLT3 tyrosine kinase domain (TKD) mutations during treatment. Six children with relapsed/refractory AML were treated with sorafenib in combination with clofarabine and cytarabine, followed by single-agent sorafenib if not a candidate for transplantation. Sunitinib was initiated after sorafenib relapse. Bone marrow samples were obtained for assessment of FLT3 TKD mutations by deep amplicon sequencing. The phase of secondary mutations with ITD alleles was assessed by cloning and sequencing of FLT3 exons 14 through 20. Identified mutations were modeled in Ba/F3 cells, and the effect of kinase inhibitors on FLT3 signaling and cell viability was assessed. Four patients achieved complete remission, but 3 receiving maintenance therapy with sorafenib relapsed after 14 to 37 weeks. Sunitinib reduced circulating blasts in two patients and marrow blasts in one. Two patients did not respond to sorafenib combination therapy or sunitinib. FLT3 mutations at residues D835 and F691 were observed in sorafenib resistance samples on both ITD-positive and -negative alleles. Deep sequencing revealed low-level mutations and their evolution during sorafenib treatment. Sunitinib suppressed leukemic clones with D835H and F691L mutations, but not D835Y. Cells expressing sorafenib-resistant FLT3 mutations were sensitive to sunitinib in vitro. Sunitinib has activity in patients that are resistant to sorafenib and harbor secondary FLT3 TKD mutations. The use of sensitive methods to monitor FLT3 mutations during therapy may allow individualized treatment with the currently available kinase inhibitors.

Baker SD ，Zimmerman EI ，Wang YD ，Orwick S ，Zatechka DS ，Buaboonnam J ，Neale GA ，Olsen SR ，Enemark EJ ，Shurtleff S ，Rubnitz JE ，Mullighan CG ，Inaba H ... -  《-》

Selective FLT3 inhibition of FLT3-ITD+ acute myeloid leukaemia resulting in secondary D835Y mutation: a model for emerging clinical resistance patterns.

Moore AS ，Faisal A ，Gonzalez de Castro D ，Bavetsias V ，Sun C ，Atrash B ，Valenti M ，de Haven Brandon A ，Avery S ，Mair D ，Mirabella F ，Swansbury J ，Pearson AD ，Workman P ，Blagg J ，Raynaud FI ，Eccles SA ，Linardopoulos S ... -  《-》

Activity of ponatinib against clinically-relevant AC220-resistant kinase domain mutants of FLT3-ITD.

Smith CC ，Lasater EA ，Zhu X ，Lin KC ，Stewart WK ，Damon LE ，Salerno S ，Shah NP ... -  《-》