Role of Wnt/β-Catenin Signalling in Acute Myeloid Leukemia (AML) Cell Response to Chemotherapy

Abstract

Background: Growing evidences from both preclinical and clinical investigations reveal the critical role of Wnt signalling for the development of many cancers and for their response to chemotherapy. Although recent studies suggest that aberrant Wnt signalling can be involved in the neoplastic myeloid cell growth, the contribution of the Wnt/β-catenin pathway to AML survival and chemoresistance is still unclear. Aims: In this study, we investigated the contribution of WNT/β-CATENIN signalling to AML survival and chemoresistance. For this purpose we tested different modulators of Wnt/β-Catenin pathway for their ability to influence AML cells proliferation and response to Cytarabine (Ara-C) or Idarubicin treatment. Methods: AML primary blast cells(30 samples) or AML cell lines cultured alone or in presence of human bone marrow mesenchymal stromal cells (hBM-MSCs), were treated with with Cytarabine (Ara-C) or Idarubicin, in presence or absence of Wnt modulators, including ligands (Wnt3a, Wnt5a/5b), Porcupine inhibitors (IWP-2), LRP6 inhibitors (Niclosamide), or antagonists of TCF/β-catenin (PKF118-310, PNU-74654). Results: In silico analysis showed the enrichment of Wnt signalling components in AML samples. Western Blot and flow cytometry showed the presence of total β-catenin only in about 2/3 of primary samples analyzed, while . β-catenin positive samples had different degree of activation of the pathway, as revealed by the expression of active forms of β-catenin, including (Ser675)β-catenin and non-phospho-(Ser33/37/Thr41) β-catenin. Notably, we found that active forms of β-catenin increased in AML samples in co-culture with hBM-MSCs, thus suggesting that Wnt signalling could be involved in the crosstalk between bone marrow stroma and AML cells. The addition of Wnt or pharmacological inhibitors, such as IWP-2, PNU-74654 and Niclosamide, to the culture medium of β-catenin-positive AML samples, either cultured alone or in co-culture with hBM-MSCs, reduced AML cell proliferation with slight effect on cell death. When associated to Idarubicin, all Wnt inhibitors except IWP-2 synergycally induced a dramatic cell death in AML cells in both culture conditions. However, when Idarubicin was replaced by Ara-C the synergism was observed only with Niclosamide and PKF. Cell death was mainly due to apoptosis, as shown by Annexin-V staining. Conclusion: Overall our data show that Wnt inhibitors reduce proliferation and chemoresistance of AML cells in culture or co-culture with bone marrow stroma cells. Wnt/β-catenin signalling may represent a potential therapeutic strategy to improve AML treatment, overcoming bone marrow stromal-mediated anti-apoptotic and chemoresistance effects. Disclosures Bonifacio: Ariad Pharmaceuticals: Consultancy; Pfizer: Consultancy; Bristol Myers Squibb: Consultancy; Novartis: Research Funding; Amgen: Consultancy.

Adjuvant Epigenetic Therapy of Decitabine and Suberoylanilide Hydroxamic Acid Exerts Anti-Neoplastic Effects in Acute Myeloid Leukemia Cells

“…Aiming to further decode the molecular mechanism underlying the higher anti-neoplastic effect of combined DAC and SAHA observed in our study, we postulated that the combinatorial treatment will modify the expression of Wnt/β-catenin and PI3K/AKT signaling pathways. The upregulation of these two signaling pathways is detectable in a high percentage of AML samples, and they have been implicated in the induction and progression of various cancers, including AML [35,60,61]. Our results showed that among the assayed proteins-pPI3K, pAKT, Wnt1, pGSK3, and p-βcatenin-SAHA treatment significantly upregulated pPI3K and pAKT expression, and the combination treatment significantly upregulated pGSK3 expression in U937 cells relative to control cells.…”

The Role of Notch and Wnt Signaling in MSC Communication in Normal and Leukemic Bone Marrow Niche

“…Transcriptomic and proteomic approaches, such as flow cytometry, ELISA, Western immunobloting, and mass spectrometry, showed in MSCs the enrichment in both canonical and non-canonical Wnt pathway components (Kuljanin et al, 2017). Using phosphospecific antibodies, we observed that Ser33/37/Thr41-phospho β-catenin (inactive) is totally absent in MSC cell lysate, thus suggesting that the Wnt/β-catenin is fully active in MSCs (Takam Kamga et al, 2016b;Wang et al, 2019). The requirement of a functional β-catenin-independent Wnt signaling, such as Wnt/Ca 2+ , Wn/Jnk, Wnt/Ryk, Wnt/Ror2, was also described in MSCs (Qiu et al, 2011;Qu et al, 2013;Jeong et al, 2020).…”

“…Accordingly, the addition of exogenous ligands of the two pathways, such as Jagged-1 or DLL-1 (Notch signaling), and Wnt3a (canonical Wnt signaling), to cultures of purified primitive human blood progenitors induces selfrenewal, survival and expansion of stem cells provided with pluripotent repopulating capacity in mouse models (Karanu et al, 2000;Willert et al, 2003;Delaney, 2005). Our and other groups have thoroughly described the expression of Notch and Wnt signaling in MSCs (Kamdje et al, 2011;Kamdje et al, 2012;Takam Kamga et al, 2016b), but other MSC-derived stromal components, including osteoblasts, and endothelial cells, can be the source of paracrine Wnt and Notch signaling in the BM (Nemeth et al, 2009;Wang et al, 2016). Moreover, MSCs can reconstitute the complete human BMME in irradiated mice (Muguruma et al, 2006) and therefore improve HSC engraftment following transplantation (Zhao et al, 2019).…”

“…-Overexpression of Notch1 and Jagged1 in AML-MSCs (Takam Kamga et al, 2016a). -Overexpression of Wnt molecules in AML-MSCs (Takam Kamga et al, 2016b).…”

“…-Notch signaling is required for β-catenin-mediated oncogenesis in mouse models of AML (Kode et al, 2014). -MSC-derived Notch signaling supports growth and survival of leukemic cells (Takam Kamga et al, 2016b). -MSC-derived Notch signaling supports growth and survival of leukemic cells (Takam Kamga et al, 2016a).…”