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Loss of the tumor suppressor phosphatase and tensin homologue deleted on chromosome 10 (PTEN), which negatively regulates the PI3K–AKT–mTOR pathway, is strongly linked to advanced prostate cancer progression and poor clinical outcome. Accordingly, several therapeutic approaches are currently being explored to combat PTEN-deficient tumors. These include classical inhibition of the PI3K–AKT–mTOR signaling network, as well as new approaches that restore PTEN function, or target PTEN regulation of chromosome stability, DNA damage repair and the tumor microenvironment. While targeting PTEN-deficient prostate cancer remains a clinical challenge, new advances in the field of precision medicine indicate that PTEN loss provides a valuable biomarker to stratify prostate cancer patients for treatments, which may improve overall outcome. Here, we discuss the clinical implications of PTEN loss in the management of prostate cancer and review recent therapeutic advances in targeting PTEN-deficient prostate cancer. Deepening our understanding of how PTEN loss contributes to prostate cancer growth and therapeutic resistance will inform the design of future clinical studies and precision-medicine strategies that will ultimately improve patient care. View Full-Text\ud Keywords: PTEN; PI3K; targeted therapy; prostate cancer

Oncogenic activation of the phosphatidylinositol-3-kinase (PI3K), protein kinase B (PKB/AKT), and mammalian target of rapamycin (mTOR) pathway is a frequent event in prostate cancer that facilitates tumor formation, disease progression and therapeutic resistance. Recent discoveries indicate that the complex crosstalk between the PI3K-AKT-mTOR pathway and multiple interacting cell signaling cascades can further promote prostate cancer progression and influence the sensitivity of prostate cancer cells to PI3K-AKT-mTOR-targeted therapies being explored in the clinic, as well as standard treatment approaches such as androgen-deprivation therapy (ADT). However, the full extent of the PI3K-AKT-mTOR signaling network during prostate tumorigenesis, invasive progression and disease recurrence remains to be determined. In this review, we outline the emerging diversity of the genetic alterations that lead to activated PI3K-AKT-mTOR signaling in prostate cancer, and discuss new mechanistic insights into the interplay between the PI3K-AKT-mTOR pathway and several key interacting oncogenic signaling cascades that can cooperate to facilitate prostate cancer growth and drug-resistance, specifically the androgen receptor (AR), mitogen-activated protein kinase (MAPK), and WNT signaling cascades. Ultimately, deepening our understanding of the broader PI3K-AKT-mTOR signaling network is crucial to aid patient stratification for PI3K-AKT-mTOR pathway-directed therapies, and to discover new therapeutic approaches for prostate cancer that improve patient outcome.

Purpose: The MYC oncogene is frequently overexpressed in prostate cancer. Upregulation of ribosome biogenesis and function is characteristic of MYC-driven tumors. In addition, PIM kinases activate MYC signaling and mRNA translation in prostate cancer and cooperate with MYC to accelerate tumorigenesis. Here, we investigate the efficacy of a single and dual approach targeting ribosome biogenesis and function to treat prostate cancer. Experimental Design:The inhibition of ribosomal RNA (rRNA) synthesis with CX-5461, a potent, selective, and orally bioavailable inhibitor of RNA polymerase I (Pol I) transcription, has been successfully exploited therapeutically but only in models of hematologic malignancy. CX-5461 and CX-6258, a pan-PIM kinase inhibitor, were tested alone and in combination in prostate cancer cell lines, in Hi-MYC- and PTEN-deficient mouse models and in patient-derived xenografts (PDX) of metastatic tissue obtained from a patient with castration-resistant prostate cancer. Results: CX-5461 inhibited anchorage-independent growth and induced cell-cycle arrest in prostate cancer cell lines at nanomolar concentrations. Oral administration of 50 mg/kg CX-5461 induced TP53 expression and activity and reduced proliferation (MKI67) and invasion (loss of ductal actin) in Hi-MYC tumors, but not in PTEN-null (low MYC) tumors. While 100 mg/kg CX-6258 showed limited effect alone, its combination with CX-5461 further suppressed proliferation and dramatically reduced large invasive lesions in both models. This rational combination strategy significantly inhibited proliferation and induced cell death in PDX of prostate cancer. Conclusions: Our results demonstrate preclinical efficacy of targeting the ribosome at multiple levels and provide a new approach for the treatment of prostate cancer. Clin Cancer Res; 22(22); 5539–52. ©2016 AACR.

Copper-ionophores that elevate intracellular bioavailable copper display significant therapeutic utility against prostate cancer cells in vitro and in TRAMP (Transgenic Adenocarcinoma of Mouse Prostate) mice. However, the pharmacological basis for their anticancer activity remains unclear, despite impending clinical trails. Herein we show that intracellular copper levels in prostate cancer, evaluated in vitro and across disease progression in TRAMP mice, were not correlative with copper-ionophore activity and mirrored the normal levels observed in patient prostatectomy tissues (Gleason Score 7 & 9). TRAMP adenocarcinoma cells harbored markedly elevated oxidative stress and diminished glutathione (GSH)-mediated antioxidant capacity, which together conferred selective sensitivity to prooxidant ionophoric copper. Copper-ionophore treatments [CuII(gtsm), disulfiram & clioquinol] generated toxic levels of reactive oxygen species (ROS) in TRAMP adenocarcinoma cells, but not in normal mouse prostate epithelial cells (PrECs). Our results provide a basis for the pharmacological activity of copper-ionophores and suggest they are amendable for treatment of patients with prostate cancer. Additionally, recent in vitro and mouse xenograft studies have suggested an increased copper requirement by prostate cancer cells. We demonstrated that prostate adenocarcinoma development in TRAMP mice requires a functional supply of copper and is significantly impeded by altered systemic copper distribution. The presence of a mutant copper-transporting Atp7b protein (tx mutation: A4066G/Met1356Val) in TRAMP mice changed copper-integration into serum and caused a remarkable reduction in prostate cancer burden (64% reduction) and disease severity (grade), abrogating adenocarcinoma development. Implications for current clinical trials are discussed.

Genetic alterations that potentiate PI3K signaling are frequent in prostate cancer, yet how different genetic drivers of the PI3K cascade contribute to prostate cancer is unclear. Here, we report PIK3CA mutation/amplification correlates with poor survival of patients with prostate cancer. To interrogate the requirement of different PI3K genetic drivers in prostate cancer, we employed a genetic approach to mutate Pik3ca in mouse prostate epithelium. We show Pik3caH1047R mutation causes p110α-dependent invasive prostate carcinoma in vivo. Furthermore, we report that PIK3CA mutation and PTEN loss coexist in patients with prostate cancer and can cooperate in vivo to accelerate disease progression via AKT–mTORC1/2 hyperactivation. Contrasting single mutants that slowly acquire castration-resistant prostate cancer (CRPC), concomitant Pik3ca mutation and Pten loss caused de novo CRPC. Thus, Pik3ca mutation and Pten deletion are not functionally redundant. Our findings indicate that PIK3CA mutation is an attractive prognostic indicator for prostate cancer that may cooperate with PTEN loss to facilitate CRPC in patients. Significance: We show PIK3CA mutation correlates with poor prostate cancer prognosis and causes prostate cancer in mice. Moreover, PIK3CA mutation and PTEN loss coexist in prostate cancer and can cooperate in vivo to accelerate tumorigenesis and facilitate CRPC. Delineating this synergistic relationship may present new therapeutic/prognostic approaches to overcome castration/PI3K–AKT–mTORC1/2 inhibitor resistance. Cancer Discov; 8(6); 764–79. ©2018 AACR. See related commentary by Triscott and Rubin, p. 682. This article is highlighted in the In This Issue feature, p. 663

Skin integrity requires an ongoing replacement and repair orchestrated by several cell types. We previously investigated the architecture of the skin of avian myeloblastosis viral oncogene homolog (Myb) knock-out (KO) embryos and wound repair in Myb(+/)(-) mice revealing a need for Myb in the skin, attributed to fibroblast-dependent production of collagen type 1. Here, using targeted Myb deletion in keratin-14 (K14) positive cells we reveal further Myb-specific defects in epidermal cell proliferation, thickness and ultrastructural morphology. This was associated with a severe deficit in collagen type 1 production, reminiscent of that observed in patients with ichthyosis vulgaris and Ehlers-Danlos syndrome. Since collagen type 1 is a product of fibroblasts, the collagen defect observed was unexpected and appears to be directed by the loss of Myb with significantly reduced tumor growth factor beta 1 (Tgfβ-1) expression by primary keratinocytes. Our findings support a specific role for Myb in K14+ epithelial cells in the preservation of adult skin integrity and function.

The therapeutic efficacy of two bis(thiosemicarbazonato) copper complexes, glyoxalbis[N4-methylthiosemicarbazonato]Cu(II) [Cu(II)(gtsm)] and diacetylbis[N4-methylthiosemicarbazonato]Cu(II) [Cu(II)(atsm)], for the treatment of prostate cancer was assessed in cell culture and animal models. Distinctively, copper dissociates intracellularly from Cu(II)(gtsm) but is retained by Cu(II)(atsm). We further demonstrated that intracellular H2gtsm [reduced Cu(II)(gtsm)] continues to redistribute copper into a bioavailable (exchangeable) pool. Both Cu(II)(gtsm) and Cu(II)(atsm) selectively kill transformed (hyperplastic and carcinoma) prostate cell lines but, importantly, do not affect the viability of primary prostate epithelial cells. Increasing extracellular copper concentrations enhanced the therapeutic capacity of both Cu(II)(gtsm) and Cu(II)(atsm), and their ligands (H2gtsm and H2atsm) were toxic only toward cancerous prostate cells when combined with copper. Treatment of the Transgenic Adenocarcinoma of Mouse Prostate (TRAMP) model with Cu(II)(gtsm) (2.5 mg/kg) significantly reduced prostate cancer burden (∼70%) and severity (grade), while treatment with Cu(II)(atsm) (30 mg/kg) was ineffective at the given dose. However, Cu(II)(gtsm) caused mild kidney toxicity in the mice, associated primarily with interstitial nephritis and luminal distention. Mechanistically, we demonstrated that Cu(II)(gtsm) inhibits proteasomal chymotrypsin-like activity, a feature further established as being common to copper-ionophores that increase intracellular bioavailable copper. We have demonstrated that increasing intracellular bioavailable copper can selectively kill cancerous prostate cells in vitro and in vivo and have revealed the potential for bis(thiosemicarbazone) copper complexes to be developed as therapeutics for prostate cancer.

The translation of basic research into improved therapies for breast cancer patients requires relevant preclinical models that incorporate spontaneous metastasis. We have completed a functional and molecular characterisation of a new isogenic C57BL/6 mouse model of breast cancer metastasis, comparing and contrasting it with the established BALB/c 4T1 model. Metastatic EO771.LMB tumours were derived from poorly metastatic parental EO771 mammary tumours. Functional differences were evaluated using both in vitro assays and spontaneous metastasis assays in mice. Results were compared to non-metastatic 67NR and metastatic 4T1.2 tumours of the 4T1 model. Protein and transcript levels of markers of human breast cancer molecular subtypes were measured in the four tumour lines, as well as p53 (Tp53) tumour-suppressor gene status and responses to tamoxifen in vivo and in vitro. Array-based expression profiling of whole tumours identified genes and pathways that were deregulated in metastatic tumours. EO771.LMB cells metastasised spontaneously to lung in C57BL/6 mice and displayed increased invasive capacity compared with parental EO771. By immunohistochemical assessment, EO771 and EO771.LMB were basal-like, as was the 4T1.2 tumour, whereas 67NR had a luminal phenotype. Primary tumours from all lines were negative for progesterone receptor, Erb-b2/Neu and cytokeratin 5/6, but positive for epidermal growth factor receptor (EGFR). Only 67NR displayed nuclear estrogen receptor alpha (ERα) positivity. EO771 and EO771.LMB expressed mutant p53, whereas 67NR and 4T1.2 were p53-null. Integrated molecular analysis of both the EO771/EO771.LMB and 67NR/4T1.2 pairs indicated that upregulation of matrix metalloproteinase-3 (MMP-3), parathyroid hormone-like hormone (Pthlh) and S100 calcium binding protein A8 (S100a8) and downregulation of the thrombospondin receptor (Cd36) might be causally involved in metastatic dissemination of breast cancer.

Background: Prostate epithelium is exquisitely sensitive to the overexpression of the proto-oncogene MYC which causes neoplastic transformation. Indeed, MYC protein is almost universally overexpressed in metastatic castration-resistant prostate cancer (CRPC) making targeting MYC an attractive option for treating advanced stage disease. Unfortunately, the development of therapeutic agents directly targeting MYC has been largely unsuccessful, thus emphasizing the need to indirectly target MYC activity through inhibition of downstream cellular processes it regulates. One of the main effects of MYC in cancer cells is to accelerate proliferative growth via stimulation of high levels of ribosome biogenesis. Accordingly, the control of protein synthesis rate has emerged as the “Achilles' heel” of a wide array of tumors. MYC also regulates and cooperates with PIM kinases to increase the activity of the eIF4F translation initiation complex and MYC-driven tumors are addicted to eIF4E. Here, we investigate the efficacy of a single and dual approach targeting ribosome biogenesis and function to treat prostate cancer (PC). Experimental design: We employed numerous models of PC, including a novel CRPC patient derived xenograft system, which showed the pre-clinical efficacy of therapies that combine to target MYC directed signaling to the ribosome. The inhibition of ribosomal RNA (rRNA) synthesis with CX-5461, a potent, selective and orally bioavailable inhibitor of RNA polymerase I (Pol I) transcription has been successfully exploited therapeutically, but only in models of hematological malignancy. CX-5461 and CX-6258, a pan-PIM kinase inhibitor, were tested alone and in combination in PC cell lines, in Hi-MYC and PTEN-deficient mouse models and in patient derived xenografts (PDX) of metastatic tissue obtained from a castration-resistant PC patient. Results: CX-5461 inhibited anchorage-independent growth and induced cell cycle arrest in PC cell lines at nanomolar concentrations. Oral administration of 50 mg/kg CX-5461 induced p53 expression and activity and reduced proliferation (Ki-67) and invasion (loss of ductal actin) in Hi-MYC tumors, but not in PTEN null (low MYC) tumors. While 100 mg/kg CX-6258 showed limited effect alone, its combination with CX-5461 further suppressed proliferation and dramatically reduced large invasive lesions in both models. This rational combination strategy significantly inhibited proliferation and induced cell death in PDX of PC. Conclusion: Our results demonstrate preclinical efficacy of targeting the ribosome at multiple levels and provide a new approach for the treatment of PC. In addition, a key conclusion of our study is that the androgen receptor (AR) presence or activity has no significant impact on the therapeutic activity of our novel combination therapy. Therefore, we believe our new exciting combination therapy could be used in the clinic in combination with current anti-androgens or as salvage therapy in multi-drug resistant CRPC. Citation Format: Richard J. Rebello, Eric Kusnadi, Don P. Cameron, Helen B. Pearson, Analia Lesmana, Jennifer R. Devlin, Denis Drygin, Ashlee K. Clark, Laura Porter, John Pedersen, Shahneen Sandhu, Gail P. Risbridger, Richard B. Pearson, Ross D. Hannan, Luc Furic. Inhibition of ribosomal RNA synthesis as a new therapeutic approach to treat advanced prostate cancer. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr B23.

Loss of cell polarity and tissue architecture is a hallmark of aggressive epithelial cancers. In addition to serving as an initial barrier to tumorigenesis, evidence in the literature has pointed towards a highly conserved role for many polarity regulators during tumor formation and progression. Here, we review recent developments in the field that have been driven by genetically engineered mouse models that establish the tumor suppressive and context dependent oncogenic function of cell polarity regulators in vivo. These studies emphasize the complexity of the polarity network during cancer formation and progression, and reveal the need to interpret polarity protein function in a cell-type and tissue specific manner. They also highlight how aberrant polarity signaling could provide a novel route for therapeutic intervention to improve our management of malignancies in the clinic.

Lung cancer is the leading cause of cancer deaths worldwide with non small-cell lung cancer (NSCLC) accounting for 80% of all lung cancers. Although activating mutations in genes of the RAS-MAPK pathway occur in up to 30% of all NSCLC, the cooperating genetic lesions that are required for lung cancer initiation and progression remain poorly understood. Here we identify a role for the cell polarity regulator Scribble (Scrib) in NSCLC. A survey of genomic databases reveals deregulation of SCRIB in human lung cancer and we show that Scrib(+/-) mutant mice develop lung cancer by 540 days with a penetrance of 43%. To model NSCLC development in vivo, we used the extensively characterized LSL-KRas(G12D) murine model of NSCLC. We show that loss of Scrib and activated oncogenic KRas cooperate in vivo, resulting in more aggressive lung tumors, likely due to a synergistic elevation in RAS-MAPK signaling. Finally, we provide data consistent with immune infiltration having an important role in the acceleration of tumorigenesis in KRas(G12D) lung tumors following Scrib loss.

Loss of cellular polarity is a hallmark of epithelial cancers, raising the possibility that regulators of polarity have a role in suppressing tumorigenesis. The Scribble complex is one of at least three interacting protein complexes that have a critical role in establishing and maintaining epithelial polarity. In human colorectal, breast, and endometrial cancers, expression of the Scribble complex member SCRIB is often mislocalized and deregulated. Here, we report that Scrib is indispensable for prostate homeostasis in mice. Scrib heterozygosity initiated prostate hyperplasia, while targeted biallelic Scrib loss predisposed mice to prostate intraepithelial neoplasia. Mechanistically, Scrib was shown to negatively regulate the MAPK cascade to suppress tumorigenesis. Further analysis revealed that prostate-specific loss of Scrib in mice combined with expression of an oncogenic Kras mutation promoted the progression of prostate cancer that recapitulated the human disease. The clinical significance of the work in mice was highlighted by our observation that SCRIB deregulation strongly correlated with poor survival in human prostate cancer. These data suggest that the polarity network could provide a new avenue for therapeutic intervention.

Aberrant Ras and Wnt signaling are emerging as key events in the multistep nature of prostate tumorigenesis and progression. Here, we report the generation of a compound model of prostate cancer to define the synergism of activated K-ras (K-ras+/V12) and dominant stabilized β-catenin (Catnb+/lox(ex3)) in the murine prostate. Recombination of floxed alleles and subsequent expression of oncogenic transgenes was mediated by Cre recombinase expression governed by the composite Probasin (PB) promoter (termed PBCre). Concomitant with elevated mitogen-activated protein kinase (MAPK) signaling, PBCre+K-ras+/V12 mice developed AH at 100 days (100% incidence) and low-grade prostate intraepithelial neoplasia and adenocarcinoma (60% and 7% incidence) by 500 days. PBCre+Catnb+/lox(ex3) mice showed reduced longevity (average 428 days) and were predisposed to PIN-like keratinized squamous metaplasia at 100 days (100% incidence) and adenocarcinoma (100% incidence) at end-point. These lesions displayed elevated Wnt signaling and basal levels of MAPK signaling. Synchronous activation of K-ras and β-catenin significantly reduced survival (average 189 days), reflecting accelerated tumorigenesis and the development of invasive carcinoma that displayed activated Wnt and MAPK signaling. Notably, expression of the basal cell marker p63 negatively correlated with tumor grade, resembling human prostate adenocarcinoma. Taken together, our data show that combinatorial oncogenic mutations of K-ras and β-catenin drive rapid progression of prostate tumorigenesis to invasive carcinoma, characterized by the synergistic elevation of androgen receptor, cyclooxygenase-2, and c-Myc. [Cancer Res 2009;69(1):94–101]

Oncogenic activation of the PI3K cascade increases cell proliferation, survival, and migration to facilitate tumor growth in many human cancers, including prostate cancer. Prostate cancer is the second major cause of cancer-related deaths in men globally, reflecting resistance to standard clinical regimens (i.e. androgen ablation, radiation, and chemotherapy). To better understand the role of the PI3K pathway in the initiation, progression and treatment of prostate cancer we have developed a novel mouse model of prostate cancer by expressing a single activating mutation in Pik3ca, the gene coding for the p110α catalytic subunit of PI3K, in the prostate. To generate the model we have used a mouse with a conditional (Cre recombinase-inducible) knock-in of the common activating mutation, Pik3caH1047R, expressed from the endogenous locus and targeted the mutation to the prostate gland by crossing this mouse to mice expressing Cre under the control of prostate-specific promoter PB. Histological analysis of mice expressing Pik3caH1047R in their prostate revealed flat and cribriform hyperplastic lesions at 200 d, which progressed to locally invasive carcinoma at 400 d. This data demonstrates that a heterozygous activating mutation in Pik3ca is sufficient to cause invasive prostate cancer. Furthermore, we show that mice co-expressing heterozygous Pik3ca and KRas oncogenic mutations in the prostate display accelerated progression to invasive prostate carcinoma (100% incidence) compared to single mutants, demonstrating a synergistic relationship between these two oncogenic events. We then used this model to perform a preclinical trial to assess the therapeutic efficacy of TORC1, panPI3K/mTORC1 and MEK inhibitors, either alone or in combination, in prostate cancer. Our results show that the combination of MEK and PI3K/mTOR inhibitors has superior efficacy compared to all other combinations or single agents assessed and with no sign of toxicity. Our data provide strong evidence that simultaneously targeting both the PI3K and RAS pathways in advanced prostate cancer patients may prove to hold enhanced therapeutic outcome. Citation Format: Helen B. Pearson, Patrick O. Humbert, Wayne A. Phillips. Targeting PI3K and RAS pathways in a novel preclinical model of prostate cancer. [abstract]. In: Proceedings of the AACR Special Conference: Targeting the PI3K-mTOR Network in Cancer; Sep 14-17, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(7 Suppl):Abstract nr B31.

Prostate cancer, a hormone-dependent disease of aging men, occurs as the relative levels of estrogen compared to testosterone are increasing. Indeed, high doses of estrogen, in combination with androgens, can initiate prostate cancer. This effect is attributed to the activity of the estrogen receptor α (ERα), but a paucity of suitable models means that the precise role of ERα in prostate cancer is still poorly understood. In this study we observed increased ERα expression in three different models of prostate cancer, PTEN null mice, Hi-Myc mice, and high grade human tumor specimens. Within the PTEN null prostate, there was a consistent pattern of ERα expression: low in benign glands, moderate in tumors within the dorsal, lateral and ventral lobes, and high in tumors within the anterior prostate. This pattern significantly correlated with the levels of the proliferative marker Ki67. There was also a significant correlation between ERα and Ki67 within individual malignant glands in the anterior prostate. Furthermore, we demonstrated that ERα sustained the in vitro proliferation of cells derived from a PTEN null tumor in 2D and 3D assays. There was a significant decrease in proliferation in cells treated with TPSF, a non-competitive ERα antagonist, or with ERα-specific shRNA. Loss of ERα caused a significant decrease in the levels of MYC and other ERα target genes. It also reduced the activity of both the PI3K and MAPK pathways as measured by decreased levels of phosphorylated erk1/2, S6 kinase and other downstream factors. This effect was reversed in rescue experiments with expression constructs for both full length ERα, capable of genomic and non-genomic actions, and membrane-only ERα, only able to trigger rapid non-genomic signalling. Collectively, these results demonstrate that ERα levels increase in prostate cancer, which promotes proliferation through classical genomic and rapid non-genomic signalling. Citation Format: Itsuhiro Takizawa, Mitchell Lawrence, Helen Pearson, John Pedersen, Normand Pouliot, Australian Prostate Cancer BioResource, Patrick Humbert, Luc Furic, Gail Risbridger. Estrogen receptor alpha drives proliferation of prostate cancer through PI3K and MAPK signaling. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2106. doi:10.1158/1538-7445.AM2014-2106

There is increasing interest in the use of non-toxic natural products for the treatment of various pathologies, including cancer. In particular, biologically active constituents of the ginger oleoresin (Zingiber officinale Roscoe) have been shown to mediate anti-tumour activity and to contribute to the anti-inflammatory, antioxidant, antimicrobial, and antiemetic properties of ginger. Here we report on the inhibitory properties of [10]-gingerol against metastatic triple negative breast cancer (TNBC) in vitro and in vivo. We show that [10]-gingerol concentration-dependently induces apoptotic death in mouse and human TNBC cell lines in vitro. In addition, [10]-gingerol is well tolerated in vivo, induces a marked increase in caspase-3 activation and inhibits orthotopic tumour growth in a syngeneic mouse model of spontaneous breast cancer metastasis. Importantly, using both spontaneous and experimental metastasis assays, we show for the first time that [10]-gingerol significantly inhibits metastasis to multiple organs including lung, bone and brain. Remarkably, inhibition of brain metastasis was observed even when treatment was initiated after surgical removal of the primary tumour. Taken together, these results indicate that [10]-gingerol may be a safe and useful complementary therapy for the treatment of metastatic breast cancer and warrant further investigation of its efficacy, either alone or in combination with standard systemic therapies, in pre-clinical models of metastatic breast cancer and in patients.

A subset of patients with gastric cancer have mutations in genes that participate in or regulate Wnt signaling at the level of ligand (Wnt) receptor (Fzd) binding. Moreover, increased Fzd expression is associated with poor clinical outcome. Despite these findings, there are no in vivo studies investigating the potential of targeting Wnt receptors for treating gastric cancer, and the specific Wnt receptor transmitting oncogenic Wnt signaling in gastric cancer is unknown. Here, we use inhibitors of Wnt/Fzd (OMP-18R5/vantictumab) and conditional gene deletion to test the therapeutic potential of targeting Wnt signaling in preclinical models of intestinal-type gastric cancer and ex vivo organoid cultures. Pharmacologic targeting of Fzd inhibited the growth of gastric adenomas in vivo. We identified Fzd7 to be the predominant Wnt receptor responsible for transmitting Wnt signaling in human gastric cancer cells and mouse models of gastric cancer, whereby Fzd7-deficient cells were retained in gastric adenomas but were unable to respond to Wnt signals and consequently failed to proliferate. Genetic deletion of Fzd7 or treatment with vantictumab was sufficient to inhibit the growth of gastric adenomas with or without mutations to Apc. Vantictumab is currently in phase Ib clinical trials for advanced pancreatic, lung, and breast cancer. Our data extend the scope of patients that may benefit from this therapeutic approach as we demonstrate that this drug will be effective in treating patients with gastric cancer regardless of APC mutation status. Significance: The Wnt receptor Fzd7 plays an essential role in gastric tumorigenesis irrespective of Apc mutation status, therefore targeting Wnt/Fzd7 may be of therapeutic benefit to patients with gastric cancer.