citations

                    Transcriptional control of subtype
switching ensures adaptation and growth
of pancreatic cancer
                

Authors:

Adams CR, Htwe HH, Marsh T, Wang AL, Montoya ML, Subbaraj L, Tward AD, Bardeesy N, Perera RM.

In:

Source: eLife
Publication Date: (2019)
Issue: :

Research Area:

Cancer Research/Cell Biology

Basic Research

Platform:

4D-Nucleofector® X-Unit

Experiment

Generation of GLI2 and SPP1 knockout cell lines using CRISPR/Cas9 GLI2 and SPP1 knockouts in KP4 cells were generated using the RNP-electroporation method as previously described (Liang et al., 2015). One million KP4 cells were used per electroporation using the Amaxa 4D Nucleofector kit (V4XC-9064, Lonza). Guide RNA and Cas9 complexes were formed using 160 mM crRNA annealed to 160 mM tracrRNA (Dharmacon) and incubated with 40 mM Cas9 protein (purchased from University of California, Berkeley). Cutting efficiency was assessed 48 hr post-electroporation using PCR and sanger sequencing. GLI2 and SPP1 knockout was confirmed using quantitative RT-PCR, immunoblotting and ELISA after clonal expansion of single cells.

Abstract

Pancreatic ductal adenocarcinoma (PDA) is a heterogeneous disease comprised of a basal-like subtype with mesenchymal gene signatures, undifferentiated histopathology and worse prognosis compared to the classical subtype. Despite their prognostic and therapeutic value, the key drivers that establish and control subtype identity remain unknown. Here, we demonstrate that PDA subtypes are not permanently encoded, and identify the GLI2 transcription factor as a master regulator of subtype inter-conversion. GLI2 is elevated in basal-like PDA lines and patient specimens, and forced GLI2 activation is sufficient to convert classical PDA cells to basal-like. Mechanistically, GLI2 upregulates expression of the pro-tumorigenic secreted protein, Osteopontin (OPN), which is especially critical for metastatic growth in vivo and adaptation to oncogenic KRAS ablation. Accordingly, elevated GLI2 and OPN levels predict shortened overall survival of PDA patients. Thus, the GLI2-OPN circuit is a driver of PDA cell plasticity that establishes and maintains an aggressive variant of this disease.

Open in PubMed

Transcriptional control of subtype switching ensures adaptation and growth of pancreatic cancer