Abstract 22: A new transgenic mouse model to explore the role of Cripto signaling in lethal prostate cancer
Authors: Sousa Elisa Rodrigues , Zoni Eugenio , Scarpa Mario , De Menna Marta , Alexander Allen Abey , De Brot Simone , Thalmann George N. , Kruithof-de Julio Marianna 
Published: April 2023

Introduction: Prostate cancer (PCa) at its early stages is treated with surgery or androgen-deprivation therapy, but PCa can turn castration resistant possibly due to pre-existing stem cell-like cells that reinitiate tumor growth and lead to metastasis. Given that human tumors express high levels of the oncofetal Cripto, our hypothesis is that Cripto might play a role in PCa initiation and progression, so we conditionally knock out Cripto in GEMMs models representative of early and late metastatic PCa to study its oncogenic potential.

Methods: Conditional Cripto knock out (CRIPTOflox/flox) was performed on N (Nkx3.1CreERT2, R26 LSL-YFP/LSL-YFP), NP (Nkx3.1CreERT2; Ptenflox/flox, R26 LSL-YFP/LSL-YFP) and NPK (Nkx3.1CreERT2; Ptenflox/flox; KrasLSL-G12D/+, R26 LSL-YFP/LSL-YFP) to generate respectively NC, NPC and NPKC. Normal epithelium is mirrored by N animals, whereas first stage of the disease, high-grade prostatic intraepithelial neoplasia (PIN)/carcinoma lesions with local invasive epithelium, are seen in NP. The advanced stage of PCa with metastasis formation is developed in NPK. In vivo experiments presented the following workflow: 8-weeks old mice were castrated and induced one month later with 5 daily injections of tamoxifen, after 2.5 weeks animals were weekly treated with testosterone (10 weeks). Single cells were isolated from prostate tissue and the YFP+/− population recovered by FACS sorting and cultured as organoids.

Results: Published PCa models N, NP and NPK were confirmed. The histopathological evaluation of newly generated NPC and NPKC shows presence of mPIN (100%, nNPC= 6, nNPKC=5), with a dominant cribriform morphology. NPKC presents invasive PCa with an extent dominant pattern of 100%. Invasive portions in NPKC present dense stroma forming whorl-like structures and occasional sheet-like accumulations of polygonal in stroma of regions with mPIN. Compared to the published models, NPC and NPKC feature a more reactive stroma with a mild/moderate inflammation. OrganoidsYFP+ recapitulate molecular prostate tissue features, expressing luminal and basal markers. Organoids’ morphology varies consistently: N and NC organoids present a more cystic morphology, with lower densities which is consistent with low-grade PIN phenotypes, whereas NP and NPK organoids are smaller and denser which mimics an oncogenic transformation. In general, NC, NPC and NPKC organoids present a higher percentage of hollow organoids compared to N, NP and NPK respectively.

Conclusions: Histopathological evaluation of NPC and NPKC phenotype specifically, showed important stromal alterations suggesting that Cripto might play a role not only on the epithelial compartment as well as in the stroma. For this reason, studies on secreted Cripto inhibition with ALK4-Fc are on-going. Our finding support that organoids are an efficient in vitro model replicating different phenotypes seen in vivo.

Citation Format: Elisa Rodrigues Sousa, Eugenio Zoni, Mario Scarpa, Marta De Menna, Allen Abey Alexander, Simone De Brot, George N. Thalmann, Marianna Kruithof-de Julio. A new transgenic mouse model to explore the role of Cripto signaling in lethal prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 22.