A new pathway against glioblastoma

MONTREAL—Researchers at McGill University have identified a new cellular pathway that limits the growth and spread of brain tumors by controlling the recycling of cell surface receptor proteins. The article, published today in the Journal of Cell Biology, suggests that the pathway — which involves a protein called Rab35 — is defective in many patients with glioblastoma multiforme (GBM).

Glioblastoma is the most aggressive type of brain cancer. As in other cancers, the proliferation and spread of glioblastoma cells depends on various receptor proteins on the outside of the cell. The levels of these receptor proteins are controlled by cellular transport pathways that internalize the receptors, and then either degrade them or return them to the cell surface. In healthy cells, these transport pathways are regulated by a class of protein known as small GTPases. 

“We previously demonstrated that the levels of a small GTPase called Rab35 are decreased in human glioblastomas. We thus sought to investigate the role of Rab35 in the progression of glioblastoma,” noted Peter S. McPherson, director of The Neurodegenerative Disease Research Group at The Neuro (Montreal Neurological Institute-Hospital), McGill University.

“There have been several links between Rab proteins and GBM,” says the article. “For example, Rab27a has been linked to lysosomal exocytosis regulating glioma cell migration and invasion (Liu et al., 2012), Rab3a is highly expressed in glioma cells and human GBM samples and is involved in glioma initiation and progression (Kim et al., 2014), and the expression of Rab38 is significantly increased in GBM (Wang and Jiang, 2013).”

“Rab35 has been extensively studied based on its function in the endosomal system (Chaineau et al., 2013). For example, we demonstrated that Rab35 is responsible for the recycling of cadherins from early endosomes to the plasma membrane and therefore regulates cell adhesion (Allaire et al., 2013),” the article continues. “Furthermore, by recruiting ACAP2, a GTPase activating protein (GAP) for Arf6, activated Rab35 suppresses the recycling of β1-integrin to the cell surface, decreasing cell migration (Allaire et al., 2013; Rahajeng et al., 2012; Chesneau et al., 2012; Kobayashi and Fukuda, 2012; Miyamoto et al., 2014). Thus, loss of Rab35 function leads to decreased cell adhesion and increased cell migration, processes associated with cancer progression.”

McPherson and his colleagues found that reducing the levels of Rab35 increased the growth and spread of brain tumors in mice, which shortened the animals’ life span. But elevating Rab35 levels reduced tumor growth and prolonged the animals’ survival.

“The levels of active Rab35 are controlled in part by its GEFs, members of the connecdenn/DENND1 family (Marat and McPherson, 2010; Kulasekaran et al., 2015). These proteins contain a DENN (differentially expressed in normal and neoplastic cells) domain that harbors the GEF activity,” adds the article. “Since (1) Rab35 mRNA levels are reduced in GBM (Allaire et al., 2013), (2) reduction of Rab35 protein leads to enhanced growth of GBM and decreased life span (Fig. 1), and (3) Rab35 expression reduces growth and increases lifespan (Fig. 2), stimulating GEF activity toward Rab35 could provide a mechanism to improve prognosis.”

The researchers discovered that Rab35 is activated by a pathway involving another small GTPase called Arf5. Together Arf5 and Rab35 appear to control the transport of various cell surface receptors, restricting the ability of cells to migrate and invade through tissues, and limiting the brain tumor-initiating cells’ ability to replicate.

“While the influence of Rab35 knockdown on cell migration, invasion, proliferation, and tumor growth in vivo are mostly phenocopied by Arf5 knockdown, the degree of influence is not always identical. This suggests that these two GTPases may have additional functions in these phenotypes unrelated to their role in the common Arf5/connecdenn/Rab35 axis described here,” the article reports.

One of the cell surface receptors controlled by this pathway is the signaling protein epidermal growth factor receptor (EGFR). In glioblastoma cells lacking Rab35, EGFR is increasingly recycled to the cell surface instead of being degraded, which enhances the receptor’s signaling activity. 

Inhibiting EGFR with the cancer drug erlotinib reduced the production of a protein called SPOCD1, which is known to promote proliferation and/or metastasis of multiple cancers. The researchers suggest that restoring the activity of Rab35 might limit the development of glioblastomas by altering the degradation and recycling of multiple cell surface receptors, including EGFR.

“Rab GTPases are emerging as an important new set of drug targets in cancer,” McPherson stated. “Our study reveals an unprecedented link between Rab and Arf proteins, and identifies new loci for therapeutic intervention in glioblastoma.”