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Maisel et al. J Cancer Metastasis Treat 2019;5:7 I http://dx.doi.org/10.20517/2394-4722.2018.82 Page 3 of 14

and stabilize membrane curvature and a PX domain capable of interacting with phosphoinositides (PtdIns)
to determine the localization of early endosomes [18-20] . Alteration of SNX proteins can directly affect receptor
tyrosine kinase activity, as loss of SNX1 activity is correlated to a reduction in lysosomal degradation of
epidermal growth factor receptor (EGFR) [21-23] .

Rab GTPases and retrotranslocation
Whether cargo enters the system via CME or SNXs, it will be found associated with Rab GTPases. The Rab
family of proteins - highly specialized in location and function, are frequently amplified but not mutated
in metastatic cancers, leading to increased availability of retrograde trafficking vesicle fusion components,
thereby promoting unabrogated receptor signal transduction and uninhibited nuclear colocalization [24-27] .

Rab proteins are GTPases involved in vesicular transport, providing membrane identity and tethering
vesicles prior to fusion [26,28] . They are capable of interacting with other coat components, motor proteins
including kinesins and myosins, and soluble N-ethylmaleimide-sensitive factor attachment protein receptors
[26]
(SNAREs) . Despite strong homology among the family members, each Rab confers a unique function to
[29]
the vesicles they mediate in response to specific activators . Two Rabs in particular are known mediators of
retrotranslocation: Rab22A which acts in early endosome-TGN trafficking, and Rab6A which is involved in
Golgi-ER transport.

Rab22A is involved in clathrin-independent endocytosis, associating with both early and late endosomes,
but not lysosomes. When bound to GTP and active, Rab22A is capable of binding to early endosome antigen 1
(EEA1; a marker unique to the endosomal sorting complex where most transmembrane receptors pass
through during endocytosis), directing the activator for Rab5 to promote early endosome fusion, a necessary
[30]
step in retrotranslocation . Rab22A expression is increased in lung, liver, ovarian, renal cell carcinoma,
and melanoma [30,31] . Rab22A is also commonly overexpressed in breast cancer and associated with decreased
patient survival. A HIF1α-dependent Rab, Rab22A is responsible for generating hypoxia-induced vesicles,
[32]
that when present promote breast cancer cell invasion through cytoskeletal alterations . Using miRNAs
to suppress the expression of Rab22A, studies have demonstrated that miR-373 can inhibit ovarian cancer,
miR-203 inhibits osteosarcoma, and miR-204 inhibits renal cell carcinomas, all in a Rab22A-dependent
manner [31,33,34] .

Rab6A is a GTPase associated with highly dynamic vesicles subject to retrograde transport from the Golgi
[35]
to the ER . Part of the recycling endocytic pathway, Rab6A is known to function within the cis- and trans-
Golgi, involved in both intra-Golgi and coat protein complex-1 (COP-1)-mediated Golgi-ER transport [36-38] .
Frequently dysregulated and associated with poor prognosis in cancer, it also presents with increased
expression in human epidermal growth factor 2 (HER2+; ErbB2) breast cancer patients [35,39] .

One of the primary functions of the Golgi is to aid in post-translational modifications (such as glycosylation)
of proteins, particularly for activation of receptor tyrosine kinases, which allows for ligand binding,
[40]
receptor trafficking, and internalization . As previously discussed, a key element of retrotranslocation is
the ability of vesicles to fuse with the TGN, instead of trafficking for lysosomal degradation. GOLPH3, a
resident Golgi protein, localizes to the TGN and can directly interact with the retromer complex through
Vsp35. Responsible for driving cell proliferation, tumor development, and anchorage independent growth
both in vitro and in vivo, GOLPH3 is an oncogene amplified in more than 30 percent of lung, ovarian,
and breast cancer cases (as high as 56 percent of lung cancers). Previously demonstrated in yeast that the
retromer complex is involved in mammalian target of rapamycin (mTOR) signaling, high expression levels
of mTOR have been similarly observed in GOLPH3-amplified human tumor tissues, resulting in increased
[40]
pAKT activity . Not only is increased pAKT associated with high metastatic potential in breast cancers,
but mutations in an anchor protein such as GOLPH3 could result in endosomal retention of receptors by
inhibiting their transition out of endosomes, leading to prolonged signal transduction and metastasis [41,42] .