Calcitriol Promotes Differentiation of Glioma Stem-Like Cells and Increases Their Susceptibility to Temozolomide

Abstract

Glioblastoma (GBM) is the most common and most aggressive primary brain tumor, with a very high rate of recurrence and a median survival of 15 months after diagnosis. Abundant evidence suggests that a certain sub-population of cancer cells harbors a stem-like phenotype and is likely responsible for disease recurrence, treatment resistance and potentially even for the infiltrative growth of GBM. GBM incidence has been negatively correlated with the serum levels of 25-hydroxy-vitamin D3, while the low pH within tumors has been shown to promote the expression of the vitamin D3-degrading enzyme 24-hydroxylase, encoded by the CYP24A1 gene. Therefore, we hypothesized that calcitriol can specifically target stem-like glioblastoma cells and induce their differentiation. Here, we show, using in vitro limiting dilution assays, quantitative real-time PCR, quantitative proteomics and ex vivo adult organotypic brain slice transplantation cultures, that therapeutic doses of calcitriol, the hormonally active form of vitamin D3, reduce stemness to varying extents in a panel of investigated GSC lines, and that it effectively hinders tumor growth of responding GSCs ex vivo. We further show that calcitriol synergizes with Temozolomide ex vivo to completely eliminate some GSC tumors. These findings indicate that calcitriol carries potential as an adjuvant therapy for a subgroup of GBM patients and should be analyzed in more detail in follow-up studies.

Antitumor Effects of a New Retinoate of the Fungal Cytotoxin Illudin M in Brain Tumor Models

“…For further investigation of drug efficacy and selectivity, an ex vivo tumor growth assay employing organotypic brain slice cultures of adult murine brains were used. For this purpose, GFP-expressing NCH644 (NCH644 GFP+ ) glioma stem-like cells were transplanted onto these slices, and after 1 day, the treatment was started and renewed three times per week, similar to our previous studies [ 25 , 26 ]. These cultures are suitable ex vivo models, which simulate original brain architecture and the presence of vessels, including angiogenesis and microvascular proliferation as well as specific homing of GBM cells toward those vessels, while requiring much fewer laboratory animals when compared with common in vivo animal drug tests [ 27 , 28 , 29 ].…”

“…Adult organotypic brain slice culture was carried out as described previously [ 26 , 45 , 46 ]. Briefly, the brains from adult C57BL/6 J (Envigo, Horst, The Netherlands) were dissected, and dura mater was removed after the mice were euthanized.…”

CHRDL1 Regulates Stemness in Glioma Stem-Like Cells

“…Lentiviral transduction of shRNA targeting CHRDL1 (shCHRDL1) or non-mammalian targeting Control-shRNA (shCtrl) was performed as described previously [29,30] after transfection of the shRNA-containing vector (pLKO.1-puro, Sigma-Aldrich), gag/polplasmid (psPAX2, addgene #12260) and VSV-G envelope plasmid (pMD2.G, addgene #12259) into HEK293T cells. Viral supernatants were collected after 16h and additionally after 24h pooled and mixed with fresh medium (ratio: 1:1) supplied with 8 µg/mL protamine sulfate (Sigma-Aldrich) immediately prior to transduction.…”

“…Briefly, 500 and 1,000 dissociated NCH644 and GS-5, respectively, were seeded in 96 well plates and measured after 7 days. Images were acquired using a Tecan Spark plate reader and analyzed using a self-developed macro via FIJI (v1.52p) [35] enhancing contrasts and identifying sphere area and properties as described previously [29].…”

Combinations of Calcitriol with Anticancer Treatments for Breast Cancer: An Update

“…The mechanisms so far reported by which calcitriol acts in cancer cells are diverse. Standing out among these are the promotion of cell differentiation, anti-inflammatory effects, apoptotic actions, among others [ 15 , 16 , 17 ]. Diverse preclinical and clinical studies have focused on studying combinations of calcitriol with a variety of agents with and without chemotherapeutic action in different cancer cells, including breast cancer [ 18 , 19 , 20 , 21 , 22 ].…”