Intercellular Transfer of Oncogenic KRAS via Tunneling Nanotubes Introduces Intracellular Mutational Heterogeneity in Colon Cancer Cells

Abstract

Mutated forms of the RAS oncogene drive 30% of all cancers, but they cannot be targeted therapeutically using currently available drugs. The molecular and cellular mechanisms that create a heterogenous tumor environment harboring both mutant and wild-type RAS have not been elucidated. In this study, we examined horizontal transfer of mutant KRAS (Kirsten Rat Sarcoma Virus) between colorectal cancer (CRC) cells via a direct form of cell-to-cell communication called tunneling nanotubes (TNTs). TNT formation was significantly higher in CRC cell lines expressing mutant KRAS than CRC cell lines expressing wild-type RAS; this effect was most pronounced in metastatic CRC cell lines with both mutant KRAS and deficiency in mismatch repair proteins. Using inverted and confocal fluorescence time-lapse and fluorescence recovery after photobleaching (FRAP)-based microscopy, we observed GFP-tagged mutant KRASG12D protein trafficking between CRC cells through TNTs within a span of seconds to several minutes. Notably, acquisition of mutant KRAS increased Extracellular Signal-regulated Kinase (ERK) phosphorylation and upregulated tunneling nanotube formation in recipient wildtype CRC cells. In conclusion, these findings suggest that intercellular horizontal transfer of RAS can occur by TNTs. We propose that intercellular transfer of mutant RAS can potentially induce intratumoral heterogeneity and result in a more invasive phenotype in recipient cells.

Neuronal signatures in cancer

“…Here the neurite-like protrusions are induced by the sodium channel β1 subunit and promoted metastasis and tumor cell growth. 33 Tunneling nanotube (TNT)-like protrusions, which are thinner, shorter and less stable then TMs 23 and neurite-like protrusions, have been described in a variety of cell types, including colon, 34 pancreatic 35 and prostate cancer 36 and also GB 17 in vitro. Although the in vivo relevance of these structures still remains to be elucidated, in vitro studies suggest that TNT-like structures promote resistance against cytotoxic therapies, [35][36][37] much as described for TMs.…”

Investigating Tunneling Nanotubes in Cancer Cells: Guidelines for Structural and Functional Studies through Cell Imaging

“…In this regard, TNTs have been increasingly recognized as qualified signals carriers, which can be involved in tumorigenic microenvironment setup. For instance, it has been demonstrated recently that TNTs spread the K-RAS mutation oncogenic message between colon cancer cells, contributing to the tumor heterogeneity and probably to the acquisition of an invasive phenotype of these recipient cells [7]. In addition, several teams describe that TNTs also allow tumor cells to reprogram the healthy neighboring cells to make them more conducive to the formation of a tumor niche.…”

ΕGFR/ERβ-Mediated Cell Morphology and Invasion Capacity Are Associated with Matrix Culture Substrates in Breast Cancer

“…Direct intercellular cytoplasmic connections, like TNTs, develop when traditional intercellular communication via cell junctions is not available, mainly due to cells being isolated [ 75 ]. TNTs are related to intercellular molecular exchange, including exosomes and mitochondria, and, therefore, they could be the expression of enhanced cell–cell interplay [ 76 , 77 , 78 ]. On the other hand, EGFR inhibition induced cell contacts and grouped cell growth, while it bestowed less aggressive morphological traits on the cells, including a flat polygonal or globular shape and less EVs and TNTs.…”