Subtype-specific kinase dependency regulates growth and metastasis of poor-prognosis mesenchymal colorectal cancer

Abstract

Background Colorectal cancer (CRC) can be divided into four consensus molecular subtypes (CMS), each with distinct biological features. CMS4 is associated with epithelial-mesenchymal transition and stromal infiltration (Guinney et al., Nat Med 21:1350–6, 2015; Linnekamp et al., Cell Death Differ 25:616–33, 2018), whereas clinically it is characterized by lower responses to adjuvant therapy, higher incidence of metastatic spreading and hence dismal prognosis (Buikhuisen et al., Oncogenesis 9:66, 2020). Methods To understand the biology of the mesenchymal subtype and unveil specific vulnerabilities, a large CRISPR-Cas9 drop-out screen was performed on 14 subtyped CRC cell lines to uncover essential kinases in all CMSs. Dependency of CMS4 cells on p21-activated kinase 2 (PAK2) was validated in independent 2D and 3D in vitro cultures and in vivo models assessing primary and metastatic outgrowth in liver and peritoneum. TIRF microscopy was used to uncover actin cytoskeleton dynamics and focal adhesion localization upon PAK2 loss. Subsequent functional assays were performed to determine altered growth and invasion patterns. Results PAK2 was identified as a key kinase uniquely required for growth of the mesenchymal subtype CMS4, both in vitro and in vivo. PAK2 plays an important role in cellular attachment and cytoskeletal rearrangements (Coniglio et al., Mol Cell Biol 28:4162–72, 2008; Grebenova et al., Sci Rep 9:17171, 2019). In agreement, deletion or inhibition of PAK2 impaired actin cytoskeleton dynamics in CMS4 cells and, as a consequence, significantly reduced invasive capacity, while it was dispensable for CMS2 cells. Clinical relevance of these findings was supported by the observation that deletion of PAK2 from CMS4 cells prevented metastatic spreading in vivo. Moreover, growth in a model for peritoneal metastasis was hampered when CMS4 tumor cells were deficient for PAK2. Conclusion Our data reveal a unique dependency of mesenchymal CRC and provide a rationale for PAK2 inhibition to target this aggressive subgroup of colorectal cancer.

Clinical Challenges of Consensus Molecular Subtype CMS4 Colon Cancer in the Era of Precision Medicine

“…Other candidate targets have emerged from preclinical studies, including HSP90, combined with 5-FU ( 7 ), PAK2 ( 67 ) or AKT3 ( 42 ), or PrP C , whose neutralization could have the combined advantage of reducing both TGFβ and PDGF signaling ( 68 ). From gene expression signatures studies, KRAS -mutated CMS4 colorectal cancer was further predicted to respond to combined MEK and SRC inhibition ( 69 ).…”

Kinase-targeted therapy in subsets of colorectal cancer

“…We identified PAK2 to be a key kinase in invasion and proliferation for CMS4, which is the poor-prognosis and mesenchymal subtype of CRC. Additionally, we unveiled a number of kinases that require further validation for the other subtypes, with CMS2 (the epithelial and Wnt active subtype) showing sensitivity to the largest number of kinases [ 12 ]. In a different study, we used a bioinformatical analysis of gene expression from both tumor and in vitro models that had been previously CMS-classified.…”