RHO GTPases in cancer: known facts, open questions, and therapeutic challenges

Abstract

RHO GTPases have been traditionally associated with protumorigenic functions. While this paradigm is still valid in many cases, recent data have unexpectedly revealed that RHO proteins can also play tumor suppressor roles. RHO signaling elements can also promote both pro- and antitumorigenic effects using GTPase-independent mechanisms, thus giving an extra layer of complexity to the role of these proteins in cancer. Consistent with these variegated roles, both gain- and loss-of-function mutations in RHO pathway genes have been found in cancer patients. Collectively, these observations challenge long-held functional archetypes for RHO proteins in both normal and cancer cells. In this review, I will summarize these data and discuss new questions arising from them such as the functional and clinical relevance of the mutations found in patients, the mechanistic orchestration of those antagonistic functions in tumors, and the pros and cons that these results represent for the development of RHO-based anticancer drugs.

Keywords: CDC42; GTPase; RAC; RHO; RHO GAP; RHO GEF.

Figure 1. Main regulatory cycle of RHO proteins.

GTPases, regulators, and effectors are shown in brown, green, and blue boxes, respectively. Genetic alterations found in some of these signaling elements in tumors are shown (inset). Inactivation steps are shown with blunted lanes. The indicated mutations have been collected from information present in the cBioPortal (http://www.cbioportal.org), St. Jude Cloud PeCan (https://pecan.stjude.cloud/home), and recent publications on this topic. iRHO, immature RHO; RHO^GDP, GDP-bound RHO; RHO^GTP, GTP-bound RHO; CIT, citron kinase; PI45K, phosphatidylinositol-4-phosphate 5-kinase; MLC, myosin light chain; MLCK, myosin light chain kinase; MLCP, myosin light chain phosphatase; MYH, myosin heavy chain; PIP₂, phosphatidylinositol (4,5) biphosphate; PIP₃, phosphatidylinositol (3,4,5) triphosphate.

Figure 2. Mutational pattern of RAC1, RHOA, and CDC42 genes in human tumors.

(A) cBioPortal-generated data depicting the genetic alterations found both in human tumors and cancer cell lines for the indicated RHO family genes (left). The type of mutation is indicated at the bottom. The frequency of genetic alterations found for each gene in the total number of samples that have been sequenced is indicated on the left. (B) Depiction of the main mutations found in RAC1 (top) and RHOA (bottom) genes. The most frequently mutated amino acid positions are shown in larger font. See inset for further information. (C) Examples of main mutations found in RAC1 (left) and RHOA (right) genes in the indicated tumors. The most frequent mutations are shown in larger fonts. These data were collected as in Figure 1. AITL, angioimmunoblastic T-cell lymphoma; PTCL-NOS, peripheral T-cell lymphoma-not otherwise specified.

Figure 3. Effects of indicated alterations in RHO signaling elements in tumorigenic processes in mouse models.

Protumorigenic (top-pointing red arrows), antitumorigenic (bottom-pointing blue arrows), and no significant effects (green equal sign) are indicated. ArhGEF4 and Spata13 are also known as Asef1 and Asef2 GEFs, respectively. DMBA, 7,12-dimethylbenz(a) anthracene; TPA, 12-O-tetradecanoylphorbol-13-acetate; APC, adenomatous polyposis coli; MLL-AF9, mixed lineage leukemia-MLLT3 (AF9) fusion oncoprotein; TCR^–, T-cell receptor negative.

Figure 4. Type of putative alterations in loss- and gain-of-function mutations in RHO signaling elements.

(A) Potential signaling effects induced by wild-type (a), nucleotide-free mutant (b and d), and signaling branch defective mutant (c) versions of RHOA in cells. In model b, the expression of the dominant negative RHOA mutant leads to the disruption of downstream signaling that can favor tumorigenesis by either the elimination of tumor suppressor functions or the dysfunction of normal biological processes. In addition, the binding of this mutant to upstream RHO GEFs can disrupt the signaling of the protein product of the wild-type RHOA allele present in cancer cells and of additional GTPase substrates of those RHOA GEFs. This latter defect should not occur in the case of signaling branch-deficient RHOA mutants (model c). These latter mutants should also have a minor impact on the overall downstream signaling of the normal protein (model c). In model d, the expression of the nucleotide-free RHOA mutant protein elicits a neomorphic, gain-of-function effect on upstream RHO GEFs. This model does not exclude the cooperativity between these pathways and the defective RHOA signaling proposed in the other models. This possibility was not included for the sake of simplicity. (B) Depiction of the possible segregation of the tumor promoter and suppression functions of RHO pathways in either clinically different subtypes of the same tumor (left) or in different progression stages of the same tumor type (right) according to current experimental evidence. See further details in the main text. In A and B, loss- and gain-of-function mutations are depicted as blue and red 10 point stars, respectively.

Figure 5. Pharmacological inhibition of RHO-regulated pathways.

Inhibitors targeting the indicated regulatory and effector steps are shown in light red speech bubbles. Secramine only works on CDC42, favoring its interaction with RHO GDIs. EHT1864 traps RHO proteins in the GDP-bound form. The rest of compounds are inhibitory. The list of PI3Kβ drugs includes both isoform-specific and pan-specific compounds.