A New Target in Gastric Cancer Paves the Way for Promising Novel Therapies

Findings from a recent study by Toby J. Phesse, PhD, European Cancer Stem Cell Research Institute, Cardiff University, yield new data that could lead to the development of promising novel therapies in gastric cancer.¹

In an interview with Oncology Learning Network, Dr Phesse discussed the basis for the study, his team’s results, and what these findings mean for this patient population.

What are Wnt and Fzd in the context of gastric cancers?

Wnt is the ligand for the Wnt signaling pathway, and Fzd is one of the receptors for this pathway. When Wnt binds to Fzd receptors they form a complex with other co-receptors to transmit the Wnt signal into the cell. Wnt signaling regulates several cellular functions, including proliferation, differentiation, apoptosis, and migration, and plays a critical role in the embryonic development and homeostasis of several epithelial tissues, including the gastrointestinal tract.

Aberrant activation of Wnt signaling promotes increased cell proliferation and is frequently observed in many cancers, including gastric cancer. This deregulation can occur in several ways, including mutations to cytoplasmic Wnt regulators (such as APC) or upregulation of Wnt ligands and/or Frizzled (Fzd) Wnt receptors. Indeed, several genes that act to repress Wnt signaling at the level of the receptor/ligand, such as sFRP or DKK1, are methylated and consequently downregulated in gastric cancer, which leads to activation of Wnt signaling.

Several reports also demonstrate that Fzd7 is upregulated in gastric cancer and associated with poor clinical outcome, and that the E3 ubiquitin ligase which degrades Fzd proteins on the cell surface, RNF43, is frequently mutated in gastric cancer.

Together these data suggest that targeting Wnt signaling at the level of the receptor, and specifically Fzd7, is an attractive target for the treatment of gastric cancer.

What existing data led you to evaluate the targeting of Fzd receptors to inhibit Wnt in this patient population?

I became interested in Fzd receptors after meeting my co-senior author for this paper, Professor Elizabeth Vincan whilst we were both working in Melbourne. The majority of research on Wnt signaling in cancer and stem cells had focused on the cytoplasmic regulators of the pathway such as APC, given their frequently observed mutations in cancer.

Professor Vincan had already published some interesting data showing that Fzd7 was required for mesenchymal to epithelial transition in colon cancer cells, and a paper from Karl Willert’s lab subsequently showed that Fzd7 was required to maintain the pluripotency of human embryonic stem cells.

These prompted us to explore the role of Fzd7 during homeostasis in the gastrointestinal tract, where, in collaboration with Professor Nick Barker and Professor Hans Clevers, we identified that Fzd7 was the predominant Wnt receptor regulating stem cell function in the intestinal and gastric epithelium.

Many of the signaling pathways identified as important regulators of development and stem cells are often deregulated in cancer, and as Wnt signaling was active in gastric cancer, we next explored the therapeutic benefit of targeting Fzd7 in gastric cancer.

Please briefly describe your study and its findings. Were any outcomes particularly surprising?

We first established that gastric cancer cells expressed Fzd receptors, with Fzd7 expression the highest of the 10 Fzd genes in mammals, and that secretion of Wnt ligands were required for their growth.

Using several preclinical platforms, we next showed that pharmacologic inhibition of Fzd receptors using vantictumab was able to inhibit the growth of gastric tumors in vivo, and in human gastric cancer cells using xenografts. Vantictumab inhibits 5 Fzd proteins, including Fzd7, so we next showed that the anti-tumor effects of vantictumab could be phenocopied by genetic deletion of Fzd7 alone.

Given that Wnts and Fzds can be expressed by non-epithelial cells, we employed gastric organoids—which are cultured as pure epithelial populations—to demonstrate that Wnt ligands and Fzd receptors are required cell intrinsically for the growth of gastric tumor cells.

One set of outcomes (which seem counterintuitive on first inspection, but are in fact consistent with the literature in the field) were our results when targeting Fzd receptors in APC-mutant gastric cancer cells. APC is part of the cytoplasmic degradation complex which targets the co-transcription factor β-catenin for degradation. Upon Wnt binding to Fzd the degradation complex is perturbed and thus β-catenin can translocate into the nucleus to regulate Wnt target genes.

As APC is down stream of Fzd in the Wnt pathway it might seem illogical to target Fzd in cells with mutant APC. However, mutant Apc is transcribed and translated into a protein that is still able to function in the degradation complex to turn-over β-catenin, albeit at a reduced level. Thus, we demonstrated that pharmacologic inhibition of Fzd or genetic deletion of Fzd7 was still able to block the grow of gastric cancer cells, even if they had mutations to APC.

What role does APC mutation play in this setting, and in the outcomes of these patients?

Analysis of published databases identified that APC is mutated in approximately 18% of human gastric tumors, and thus it was important for us to establish if our novel therapeutic approach targeting Fzd receptors was effective in these tumors.

We show that targeting Fzd receptors, and Fzd7 specifically, is able to markedly inhibit the growth of human gastric cancer cells, which have many mutations, including those to APC, thus illustrating this therapy is not constricted to APC wildtype tumors.

What are the real-world applications of these findings in clinical practice?

Our results now open the way for patients with gastric cancer to potentially be treated by therapies that target Fzd receptors in the future, and will directly inform any new clinical trials in this area. Indeed, vantictumab is already in phase Ib clinical trials for other solid tumors, and therefore our data indicate that patients with gastric cancer would also be suitable to be included in any potential future trials with this drug, or the new generation of PORCN inhibitors which prevent Wnt secretion.

In addition to identifying Fzd7 as an attractive therapeutic target for gastric cancer, we also established that the transcription factor Myc was required downstream of Fzd7 for gastric tumor growth. Myc has been the subject of intense research due to its upregulation in several cancers, and our work now shows that gastric tumors would also be sensitive to any Myc-specific inhibitor drugs (which, to date, have been challenging to develop).

With regard to APC-mutant tumors, we have shown that gastric tumors are still sensitive to inhibition of Fzd receptors, but is this true of other cancers with APC mutations such as colon cancer?

This is still an area of controversy and active research in the field, with conflicting sets of data on the ability of APC-mutant colon cancer cells to respond to Fzd inhibition. Interestingly, approximately 37% of APC-mutant gastric tumors are mutant for RNF43, which regulates Fzd on the cell surface, whilst in colon tumors these mutations are mutually exclusive, indicating that gastric cells and colon cells preferentially select different Wnt mutations that confer optimal levels of Wnt signaling required for tumor growth.

Thus, the level of Wnt inhibition in each cancer setting may have to be different to elicit a response which is something we are researching presently.

Do you and your co-investigators intend to expand upon this research? If so, will you be incorporating any new methods, end points, or patient populations?

Yes, a recent award from the Medical Research Council has enabled my lab to continue to investigate the role of Fzd receptors in gastric cancer, and I continue to work closely with my international co-authors—Dr Flanagan, Professor Vincan, Professor Barker, and Professor Clevers—using CRISPR and powerful preclinical platforms to study Fzd in additional cancer settings.

Reference

1.  Flanagan Dj, Barker N, Di Costanzo NS, et al. Frizzled-7 is required for Wnt signaling in gastric tumors with and without Apc mutations. Cancer Res. 2019 Jan 8. Epub ahead of print.