Cancer cells depend on specific oncogenic pathways or present a genetic alteration that leads to a particular disturbance. Still, personalized and targeted biological therapy remains challenging, with current efforts generally yielding disappointing results. Carefully assessing onco-target molecular pathways can, however, potently assist with such efforts for the selection of patient populations that would best respond to a given drug treatment. RNF43, an E3 ubiquitin ligase that negatively regulates Wnt/frizzled (FZD) receptors by their ubiquitination, internalization, and degradation, controls a key pathway in cancer. Recently, additional target proteins of RNF43 were described, including p85 of the PI3K/AKT/mTOR signaling pathway and protease-activated receptor 2 (PAR₂), a G-protein-coupled receptor that potently induces β-catenin stabilization, independent of Wnts. RNF43 mutations with impaired E3 ligase activity were found in several types of cancers (e.g., gastrointestinal system tumors and endometrial and ovarian cancer), pointing to a high dependency on FZD receptors and possibly PAR₂ and the PI3K/AKT/mTOR signaling pathway. The development of drugs toward these targets is essential for improved treatment of cancer patients.

Studying primordial events in cancer is pivotal for identifying predictive molecular indicators and for targeted intervention. While the involvement of G-protein-coupled receptors (GPCRs) in cancer is growing, GPCR-based therapies are yet rare. Here, we demonstrate the overexpression of protease-activated receptor 2 (PAR2), a GPCR member in the fallopian tubes (FTs) of high-risk BRCA carriers as compared to null in healthy tissues of FT. FTs, the origin of ovarian cancer, are known to express genes of serous tubal intraepithelial carcinoma (STICs), a precursor lesion of high-grade serous carcinoma (HGSC). PAR2 expression in FTs may serve as an early prediction sensor for ovarian cancer. We show now that knocking down Par2 inhibits ovarian cancer peritoneal dissemination in vivo, pointing to the central role of PAR2. Previously we identified pleckstrin homology (PH) binding domains within PAR1,2&4 as critical sites for cancer-growth. These motifs associate with PH-signal proteins via launching a discrete signaling network in cancer. Subsequently, we selected a compound from a library of backbone cyclic peptides generated toward the PAR PH binding motif, namely the lead compound, Pc(4-4). Pc(4-4) binds to the PAR PH binding domain and blocks the association of PH-signal proteins, such as Akt or Etk/Bmx with PAR2. It attenuates PAR2 oncogenic activity. The potent inhibitory function of Pc(4-4) is demonstrated via inhibition of ovarian cancer peritoneal spread in mice. While the detection of PAR2 may serve as a predictor for ovarian cancer, the novel Pc(4-4) compound may serve as a powerful medicament in STICs and ovarian cancer. This is the first demonstration of the involvement of PAR PH binding motif signaling in ovarian cancer and Pc(4-4) as a potential therapy treatment.

Although the role of G-protein-coupled receptors (GPCRs) in cancer is acknowledged, GPCR-based cancer therapy is rare. Mammalian protease-activated receptors (PARs), a sub-group of GPCRs, comprise four family members, termed PAR_1–4. Here, we demonstrate that PAR₂ is dominant over PAR₄ oncogene in cancer. We performed a knockdown of Par2/f2rl1 and expressed C-terminally truncated PAR₂ (TrPAR₂), incapable of inducing signaling, to assess the impact of PAR₂ on PAR₄ oncogenic function by β-catenin stabilization assessment, immunoprecipitation, and xenograft tumor generation in Nude/Nude mice. PAR₂ and PAR₄ act together to promote tumor generation. Knockdown Par2 and TrPAR₂ inhibited the PAR₂ and PAR₄-induced β-catenin levels, nuclear dishevelled 1(DVL1), and TOPflash reporter activity. Likewise, PAR₂ and PAR₄-induced invasion and migration were inhibited when Par2 was knocked down or in the presence of TrPAR₂. PAR cyclic (4-4) [Pc(4-4)], a PAR-based compound directed toward the PAR pleckstrin homology (PH)-binding site, effectively inhibited PAR₂ oncogenic activity. Pc(4-4) inhibition is mediated via the increase in p53 level and the up-regulation of p21 as caspase-3 as well. Overall, we showed that in the absence of PAR₂ signaling, the PAR₄ pro-tumor functions are significantly inhibited. Pc(4-4) inhibits PAR₂ acting via the modification of wt p53, thus offering a powerful drug measure for fighting cancer.

Hierarchical organization transpires within a tumor. It is appreciated that cancer stem cells (CSCs) are the cornerstone primary cells of such assemblies, fueled by molecular mechanisms that allow resistance to chemotherapy, vigorous self-renewal capacity, and resistance to immune therapy. Due to their powerful self-renewal capability, CSCs constantly initiate tumor cell proliferation, maintaining the growth state of the tumor. Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a known stem cell niche protein, is a G protein-coupled receptor (GPCR) that is involved in Wnt signaling. It was previously shown that members of protease-activated receptors (PARs) and another affiliate of GPCRs, the G protein-coupled receptor 25 (GPR25), elicit β-catenin stabilization and transcriptional activity. PARs and GPR25 can associate with LGR5 and markedly enhance β-catenin stabilization. The prospect of identifying additional cancer driver GPCRs partners of LGR5, such as the prostaglandin receptors, EP2 and EP4, and the lysophosphatidic acid receptors (LPARs), which have been demonstrated as capable of inducing β-catenin stabilization yet pending experimental association with LGR5, is discussed. The existence of an LGR5-GPCRs-β-catenin hub may potentially overcome tumor plasticity and alleviate drug resistance. It certainly elucidates new opportunities in the fight to eradicate cancer.