GLI2 inhibition abrogates human leukemia stem cell dormancy

Abstract

BackgroundDormant leukemia stem cells (LSC) promote therapeutic resistance and leukemic progression as a result of unbridled activation of stem cell gene expression programs. Thus, we hypothesized that 1) deregulation of the hedgehog (Hh) stem cell self-renewal and cell cycle regulatory pathway would promote dormant human LSC generation and 2) that PF-04449913, a clinical antagonist of the GLI2 transcriptional activator, smoothened (SMO), would enhance dormant human LSC eradication.MethodsTo test these postulates, whole transcriptome RNA sequencing (RNA-seq), microarray, qRT-PCR, stromal co-culture, confocal fluorescence microscopic, nanoproteomic, serial transplantation and cell cycle analyses were performed on FACS purified normal, chronic phase (CP) chronic myeloid leukemia (CML), blast crisis (BC) phase CML progenitors with or without PF-04449913 treatment.ResultsNotably, RNA-seq analyses revealed that Hh pathway and cell cycle regulatory gene overexpression correlated with leukemic progression. While lentivirally enforced GLI2 expression enhanced leukemic progenitor dormancy in stromal co-cultures, this was not observed with a mutant GLI2 lacking a transactivation domain, suggesting that GLI2 expression prevented cell cycle transit. Selective SMO inhibition with PF-04449913 in humanized stromal co-cultures and LSC xenografts reduced downstream GLI2 protein and cell cycle regulatory gene expression. Moreover, SMO inhibition enhanced cell cycle transit and sensitized BC LSC to tyrosine kinase inhibition in vivo at doses that spare normal HSC.ConclusionIn summary, while GLI2, forms part of a core HH pathway transcriptional regulatory network that promotes human myeloid leukemic progression and dormant LSC generation, selective inhibition with PF-04449913 reduces the dormant LSC burden thereby providing a strong rationale for clinical trials predicated on SMO inhibition in combination with TKIs or chemotherapeutic agents with the ultimate aim of obviating leukemic therapeutic resistance, persistence and progression.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-015-0453-9) contains supplementary material, which is available to authorized users.

Small‐molecule Hedgehog inhibitor attenuates the leukemia‐initiation potential of acute myeloid leukemia cells

“…It was previously reported that Hh signaling modulates cell cycle regulators in stem cells to control normal hematopoietic regeneration . In the BCR‐ABL‐positive leukemia model, selective SMO inhibition abrogates human LSC dormancy . The authors showed that treatment with PF‐913 in humanized stromal cocultures reduced downstream cell cycle gene expression and sensitized BCR‐ABL‐positive LSCs to tyrosine kinase inhibition.…”

“…(27) In the BCR-ABL-positive leukemia model, selective SMO inhibition abrogates human LSC dormancy. (28) The authors showed that treatment with PF-913 in humanized stromal cocultures reduced downstream cell cycle gene expression and sensitized BCR-ABL-positive LSCs to tyrosine kinase inhibition. A recent study also reported that inhibition of Hh signaling induced apoptosis by decreasing c-Myc protein expression in OPM1 myeloma cells.…”

Glasdegib in combination with cytarabine and daunorubicin in patients with AML or high‐risk MDS: Phase 2 study results

“…Glasdegib prevents the translocation of SMO into primary cilia and prevents SMO‐mediated activation of downstream Hh targets . Previous studies have reported glasdegib inhibition of SMO reduced the expression of key intracellular leukemia stem cell regulators (eg, GLI2) and enhanced cell cycle transit . These results and preclinical evidence that Hh inhibition may sensitize cells to cytarabine or azacitidine provided rationale for evaluating glasdegib in combination with chemotherapeutic agents to reduce resistance and leukemic persistence or progression.…”

The Wnt signaling pathway in cancer

“…CSCs, represent a rare population of cells amongst the tumor bulk that are able to maintain the tumor via proliferation and self-renewal 13 capabilities and telomerase expression 13 . They are also known to be more resistant to conventional treatment (chemotherapy and radiotherapy) and responsible for cancer relapse and metastasis 14,15 . Their quiescent state and specific interactions with their microenvironment play a significant role in their drug resistance properties 16 .…”