TERT promoter C228T mutation in neural progenitors confers growth advantage following telomere shortening in vivo

Abstract

Background Heterozygous TERT (Telomerase reverse transcriptase) promoter mutations (TPMs) facilitate TERT expression and are the most frequent mutation in glioblastoma (GBM). A recent analysis revealed this mutation is one of the earliest events in gliomagenesis. However, no appropriate human models have been engineered to study the role of this mutation in the initiation of these tumors. Method We established GBM models by introducing the heterozygous TPM in human induced pluripotent stem cells (hiPSCs) using a two-step targeting approach in the context of GBM genetic alterations, CDKN2A/B and PTEN deletion, and EGFRvIII overexpression. The impact of the mutation was evaluated through the in vivo passage and in vitro experiment and analysis. Results Orthotopic injection of neuronal precursor cells (NPCs) derived from hiPSCs with the TPM into immunodeficient mice did not enhance tumorigenesis compared to TERT promoter wild type NPCs at initial in vivo passage presumably due to relatively long telomeres. However, the mutation recruited GA-Binding Protein and engendered low-level TERT expression resulting in enhanced tumorigenesis and maintenance of short telomeres upon secondary passage as observed in human GBM. These results provide the first insights regarding increased tumorigenesis upon introducing a TPM compared to isogenic controls without TPMs. Conclusion Our novel GBM models presented the growth advantage of heterozygous TPMs for the first time in the context of GBM driver mutations relative to isogenic controls, thereby allowing for the identification and validation of TERT promoter-specific vulnerabilities in a genetically accurate background.

RNA splicing analysis deciphers developmental hierarchies and reveals therapeutic targets in adult glioma

“…Overall, our AS signature not only contributes to the intratumor- plemental Figure 4A). To assess the impact of these genetic alternations on the AS landscape, we utilized a human iPSC-derived glioma "avatar" model (32,33). Given the difficulty of modeling 1p/19q codeletion, we focused on the latter 2 genotypes.…”

“…Generation of Genetically Engineered hiPSC Clones. The iPSC clones, including WT, iPSC-T (TP53 -/-), iPSC-C (CDKN2A/2B -/-), and iPSC-PCT (PTEN -/-, CDKN2A/2B -/-, TERTp C228T/WT ), were recently described (33,63). We introduced IDH1 R132H/WT or/and ATRX -/in iPSC-T and MTAP -/and/or EGFRvIII OE in iPSC-PCT using methods detailed below.…”

RNA splicing analysis deciphers developmental hierarchies and reveals therapeutic targets in adult glioma

“…Overall, our AS signature not only contributes to the intratumor- plemental Figure 4A). To assess the impact of these genetic alternations on the AS landscape, we utilized a human iPSC-derived glioma "avatar" model (32,33). Given the difficulty of modeling 1p/19q codeletion, we focused on the latter 2 genotypes.…”

“…Generation of Genetically Engineered hiPSC Clones. The iPSC clones, including WT, iPSC-T (TP53 -/-), iPSC-C (CDKN2A/2B -/-), and iPSC-PCT (PTEN -/-, CDKN2A/2B -/-, TERTp C228T/WT ), were recently described (33,63). We introduced IDH1 R132H/WT or/and ATRX -/in iPSC-T and MTAP -/and/or EGFRvIII OE in iPSC-PCT using methods detailed below.…”

Telomerase reactivation induces progression of mouse Braf V600E -driven thyroid cancers without telomere lengthening

“…To our knowledge, there had not been in vivo studies of TPMs in genetically engineered mouse models. The closest effort is a recent publication in which human stem cells were engineered for TERT -124C>T mutation, differentiated into neural precursors and orthotopically injected into mice to study glioblastoma in the context of EGFR overexpression and loss of CDKN2A and PTEN (60). Our results showing that murine Tert reactivation promotes cancer progression in oncogenic Braf-driven thyroid tumors, open the door at devising equivalent studies in tumors from other lineages with high prevalence of TPMs.…”

GABP couples oncogene signaling to telomere regulation in TERT promoter mutant cancer

“…Thus, our study and others are performed using a range of TERT promoter mutant and WT cell lines with different genetic backgrounds. An isogenic model derived from human-induced pluripotent stem cells has been described very recently ( Miki et al, 2022 ) and could be used in future studies to understand genetic dependencies of TERT promoter mutant cancers.…”

RNA splicing analysis deciphers developmental hierarchies and reveals therapeutic targets in adult glioma

“…Overall, our AS signature not only contributes to the intratumor- plemental Figure 4A). To assess the impact of these genetic alternations on the AS landscape, we utilized a human iPSC-derived glioma "avatar" model (32,33). Given the difficulty of modeling 1p/19q codeletion, we focused on the latter 2 genotypes.…”

“…Generation of Genetically Engineered hiPSC Clones. The iPSC clones, including WT, iPSC-T (TP53 -/-), iPSC-C (CDKN2A/2B -/-), and iPSC-PCT (PTEN -/-, CDKN2A/2B -/-, TERTp C228T/WT ), were recently described (33,63). We introduced IDH1 R132H/WT or/and ATRX -/in iPSC-T and MTAP -/and/or EGFRvIII OE in iPSC-PCT using methods detailed below.…”

Telomerase reactivation induces progression of mouse Braf V600E -driven thyroid cancers without telomere lengthening

“…To our knowledge, there had not been in vivo studies of TPMs in genetically engineered mouse models. The closest effort is a recent publication in which human stem cells were engineered for TERT -124C>T mutation, differentiated into neural precursors and orthotopically injected into mice to study glioblastoma in the context of EGFR overexpression and loss of CDKN2A and PTEN (60). Our results showing that murine Tert reactivation promotes cancer progression in oncogenic Braf-driven thyroid tumors, open the door at devising equivalent studies in tumors from other lineages with high prevalence of TPMs.…”

GABP couples oncogene signaling to telomere regulation in TERT promoter mutant cancer

“…Thus, our study and others are performed using a range of TERT promoter mutant and WT cell lines with different genetic backgrounds. An isogenic model derived from human-induced pluripotent stem cells has been described very recently ( Miki et al, 2022 ) and could be used in future studies to understand genetic dependencies of TERT promoter mutant cancers.…”

RNA splicing analysis deciphers developmental hierarchies and reveals therapeutic targets in adult glioma

“…Overall, our AS signature not only contributes to the intratumor- plemental Figure 4A). To assess the impact of these genetic alternations on the AS landscape, we utilized a human iPSC-derived glioma "avatar" model (32,33). Given the difficulty of modeling 1p/19q codeletion, we focused on the latter 2 genotypes.…”

“…Generation of Genetically Engineered hiPSC Clones. The iPSC clones, including WT, iPSC-T (TP53 -/-), iPSC-C (CDKN2A/2B -/-), and iPSC-PCT (PTEN -/-, CDKN2A/2B -/-, TERTp C228T/WT ), were recently described (33,63). We introduced IDH1 R132H/WT or/and ATRX -/in iPSC-T and MTAP -/and/or EGFRvIII OE in iPSC-PCT using methods detailed below.…”

Telomerase reactivation induces progression of mouse Braf V600E -driven thyroid cancers without telomere lengthening

“…To our knowledge, there had not been in vivo studies of TPMs in genetically engineered mouse models. The closest effort is a recent publication in which human stem cells were engineered for TERT -124C>T mutation, differentiated into neural precursors and orthotopically injected into mice to study glioblastoma in the context of EGFR overexpression and loss of CDKN2A and PTEN (60). Our results showing that murine Tert reactivation promotes cancer progression in oncogenic Braf-driven thyroid tumors, open the door at devising equivalent studies in tumors from other lineages with high prevalence of TPMs.…”

GABP couples oncogene signaling to telomere regulation in TERT promoter mutant cancer

“…Thus, our study and others are performed using a range of TERT promoter mutant and WT cell lines with different genetic backgrounds. An isogenic model derived from human-induced pluripotent stem cells has been described very recently ( Miki et al, 2022 ) and could be used in future studies to understand genetic dependencies of TERT promoter mutant cancers.…”