Dedifferentiation of Glioma Cells to Glioma Stem-like Cells By Therapeutic Stress-induced HIF Signaling in the Recurrent GBM Model

Abstract

Increasing evidence exposes a subpopulation of cancer cells, known as cancer stem cells (CSCs), to be critical for the progression of several human malignancies, including glioblastoma multiforme (GBM). CSCs are highly tumorigenic, capable of self-renewal, and resistant to conventional therapies, and thus considered to be one of the key contributors to disease recurrence. In order to elucidate the poorly understood evolutionary path of tumor recurrence and the role of CSCs in this process, we developed patient-derived xenograft GBM recurrent models induced by anti-glioma chemotherapy, temozolomide (TMZ). In this model, we observed a significant phenotypic shift towards an undifferentiated population. We confirmed these findings in vitro as sorted CD133-negative populations cultured in differentiation-forcing media were found to acquire CD133 expression following chemotherapy treatment. To investigate this phenotypic switch at the single cell level, GSC-specific promoter-based reporter systems were engineered to track changes in the GSC population in real time. We observed the active phenotypic and functional switch of single non-stem glioma cells to a stem-like state and that TMZ therapy significantly increased the rate of single-cell conversions. Importantly, we showed the therapy induced hypoxia inducible factors (HIF) 1α and HIF2α play key roles in allowing non-stem glioma cells to acquire stem-like traits, as the expression of both HIFs increase upon TMZ therapy and knockdown of HIFs expression inhibits the interconversion between non-stem glioma cells and GSCs post-therapy. Based on our results, we propose that anti-glioma chemotherapy promotes the accumulation of HIFs in the GBM cells that induces the formation of therapy-resistant GSCs responsible for recurrence.

De-novo purine biosynthesis is a major driver of chemoresistance in glioblastoma

“…Previous reporting demonstrated that Hypoxia-inducible factors (HIFs) are critical for maintenance of the BTIC niche and that HIF-1α (HIF1A) can directly modulate EZH2 expression via promoter binding 30,31 . We have shown that TMZ therapy increases the BTIC frequency in a HIF dependent manner 20,32 . Now we show that the TMZ-mediated increase in HIF1A is specific to the CD133+ BTIC compartment…”

“…The loss of stemness can be rescued by re-expressing ARL13B in KD cells ( Fig. 3B 32,37 . Immunoblot analysis indicated that when PDX lines cultured in NEB, the expression of ARL13B was significantly elevated in three different subtypes of GBM ( Fig.…”

TMZ regulates GBM stemness via MMP14‐DLL4‐Notch3 pathway

“…The human glioma U87MG (CVCL_0022) and U251 (CVCL_0021) cell lines were purchased from the American Type Culture Collection (Manassas, VA). The primary patient‐derived GBM1, GBM7 and GBM13 cells were prepared from specimens obtained at the N.N.…”

Radiotherapy and Glioma Stem Cells: Searching for Chinks in Cellular Armor

“…This alteration is an example of cellular plasticity, a mechanism by which GBM cells can transition from differentiated, treatment-sensitive states to treatment-resistant, undifferentiated GSC states. Hypoxia-induced factors (HIF) have been shown to participate in the regulation of the GSC niche(17, 50, 53–55). This ability of HIF to influence cellular plasticity has been confirmed in breast cancer stem cells(56).…”

“…We have previously shown that therapeutic stress induced by chemotherapy can activate cellular plasticity via HIF signaling under normoxic conditions(17). Several studies have recently confirmed that radiation also alters cellular plasticity.…”