Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher. Acknowledgments We thank Mara de Souza Junqueira and Brbara Amlia Aparecida Santana Lemos for assistance with mouse irradiation, and Patrcia Vianna Bonini Palma for guidance regarding flow cytometry experiments. in control samples. retinoic acid (ATRA) and arsenic trioxide (ATO) revolutionized the treatment of acute promyelocytic leukemia (APL), leading to a disease-free survival rate of 80-90% (1). Nevertheless, 5-10% of APL patients still relapse due to ATRA or ATO resistance (2). Despite the cytotoxic activities of ATRA and ATO in APL cells, low doses of those agents result in induction of terminal myeloid cell differentiation (3, 4). In this context, previous reports exhibited that Ondansetron Hydrochloride Dihydrate inhibitors of the epidermal growth factor receptor (EGFR) increased ATRA and ATO-induced expression of the myeloid differentiation marker CD11b in AML cells (3C7). Nonetheless, the use of EGFR inhibitors in combination with standard therapy was not previously explored in APL cells resistant to ATRA and ATO. Non-small cell lung cancer (NSCLC) exhibited constitutive activation of the epidermal PAPA1 growth factor (EGF)/EGFR pathway, due to mutations around the (8). Although mutations are rare in AML (9C11), the level of EGFthe main EGFR ligandwas elevated in the urine of patients diagnosed with APL and decreased after ATRA-induced complete remission (12). Hence, it is conceivable that this activation of the EGF/EGFR signaling pathway could also confer APL leukemic cells with a survival advantage. However, the prevalence and clinical significance of EGFR and its interactors in APL patients remains unknown. It has been well established that this distinct dimer interfaces formed between the extracellular domain name of the EGF receptor and its respective ligands EGF and amphiregulin (AREG) differentially activate intracellular signaling cascades to regulate cell proliferation and differentiation (13). The EGFR tyrosine kinase inhibitors gefitinib (ZD1839) and erlotinib (CP-358774) are small-molecule compounds that prevent the binding of ATP to the intracellular Ondansetron Hydrochloride Dihydrate domain name of EGFR, thus impairing autophosphorylation and downstream signal transduction (14). The efficacy and safety of Ondansetron Hydrochloride Dihydrate gefitinib and erlotinib as first-line therapies for NSCLC have been demonstrated in several clinical trials and retrospective studies (15). Although there is usually evidence of patients with co-occurrence of acute myeloid leukemia (AML) and NSCLC, which achieved complete hematological remission when treated with erlotinib monotherapy (16, 17), subsequent studies evaluating the response of AML patients to EGFR inhibitors alone could not corroborate these findings (18C20). In a phase II trial, 26/29 (90%) patients with refractory or relapsed AML who received erlotinib monotherapy discontinued treatment because of disease progression. Nevertheless, combination therapies between differentiation brokers with EGFR inhibitors have not been evaluated in AML patients (18). Here, we evaluated the effects of EGFR pharmacological inhibition in distinct APL models. Gefitinib monotherapy induced apoptosis and inhibited the proliferation of NB4 (ATRA-sensitive) and NB4-R2 (ATRA-resistant) APL cells. Additionally, the combination between gefitinib with ATRA and ATO rewired NB4-R2 and NB4 ATOr (ATO-resistant) cells into sensitivity to standard therapy for APL. experiments, NB4, NB4-R2, NB4-ATOr, and NB4 clone 21 cells were collected 72?h after drug treatment, washed, and resuspended in 100 L PBS and incubated with CD11b-PE (#347557, clone: D12), CD11c-APC (#559877, clone: B-ly6), CD15 (#562371, clone: 7C3.rMAb), and CD16 (#557758, clone: 3G8) (BD Biosciences). Cells obtained from BM, or the spleen of leukemia model mice were labeled with antibodies against CD11b-PE (#553311, clone: M1/70), CD117-FITC (#561680, clone: 2B8), Gr1-FITC (#551460, clone: 1A8; all from BD Biosciences), then collected and washed and resuspended in PBS. The percentage of positive cells and MFI were determined by flow cytometry. Western Blotting Whole-cell.