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Abstract Background and aims This study sought to determine the value of patient-derived organoids (PDOs) from esophago-gastric adenocarcinoma (EGC) for response prediction to neoadjuvant chemotherapy (neoCTx). Methods Endoscopic biopsies of patients with locally advanced EGC (n = 120) were taken into culture and PDOs expanded. PDOs' response towards the single substances of the FLOT regimen and the combination treatment were correlated to patients' pathological response using tumor regression grading. A classifier based on FLOT response of PDOs was established in an exploratory cohort (n = 13) and subsequently confirmed in an independent validation cohort (n = 13). Results EGC PDOs reflected patients' diverse responses to single chemotherapeutics and the combination regimen FLOT. In the exploratory cohort, PDOs response to single 5-FU and FLOT combination treatment correlated with the patients' pathological response (5-FU: Kendall's τ = 0.411, P = 0.001; FLOT: Kendall's τ = 0.694, P = 2.541e-08). For FLOT testing, a high diagnostic precision in receiver operating characteristic (ROC) analysis was reached with an AUCROC of 0.994 (CI 0.980 to 1.000). The discriminative ability of PDO-based FLOT testing allowed the definition of a threshold, which classified in an independent validation cohort FLOT responders from non-responders with high sensitivity (90%), specificity (100%) and accuracy (92%). Conclusion In vitro drug testing of EGC PDOs has a high predictive accuracy in classifying patients' histological response to neoadjuvant FLOT treatment. Taking into account the high rate of successful PDO expansion from biopsies, the definition of a threshold that allows treatment stratification paves the way for an interventional trial exploring PDO-guided treatment of EGC patients.

Abstract Gastric cancer ranks the fifth most common and third leading cause of cancer‐related deaths worldwide. Alterations in the RTK/MAPK, WNT, cell adhesion, TP53, TGFβ, NOTCH, and NFκB signaling pathways could be identified as main oncogenic drivers. A combination of altered pathways can be associated with molecular subtypes of gastric cancer. In order to generate model systems to study the impact of different pathway alterations in a defined genetic background, we generated three murine organoid models: a RAS‐activated (KrasG12D, Tp53R172H), a WNT‐activated (Apcfl/fl, Tp53R172H), and a diffuse (Cdh1fl/fl, Apcfl/fl) model. These organoid models were morphologically and phenotypically diverse, differed in proteome expression signatures and possessed individual drug sensitivities. A differential vulnerability to RTK/MAPK pathway interference based on the different mitogenic drivers and according to the level of dependence on the pathway could be uncovered. Furthermore, an association between RTK/MAPK pathway activity and susceptibility to HDAC inhibition was observed. This finding was further validated in patient‐derived organoids from gastric adenocarcinoma, thus identifying a novel treatment approach for RTK/MAPK pathway altered gastric cancer patients.

Pathological complete response (pCR) has been correlated with overall survival in several cancer entities including colorectal cancer. Novel total neoadjuvant treatment (TNT) in rectal cancer has achieved pathological complete response in one-third of the patients. To define better treatment options for nonresponding patients, we used patient-derived organoids (PDOs) as avatars of the patient’s tumor to apply both photon- and proton-based irradiation as well as single and combined chemo(radio)therapeutic treatments. While response to photon and proton therapy was similar, PDOs revealed heterogeneous responses to irradiation and different chemotherapeutic drugs. Radiotherapeutic response of the PDOs was significantly correlated with their ability to repair irradiation-induced DNA damage. The classical combination of 5-FU and irradiation could not sensitize radioresistant tumor cells. Ataxia-telangiectasia mutated (ATM) kinase was activated upon radiation, and by inhibition of this central sensor of DNA damage, radioresistant PDOs were resensitized. The study underlined the capability of PDOs to define nonresponders to irradiation and could delineate therapeutic approaches for radioresistant patients.

Inhibitor of growth 3 (ING3) is one of five members of the ING tumour suppressor family, characterized by a highly conserved plant homeodomain (PHD) as a reader of the histone mark H3K4me3. ING3 was reported to act as a tumour suppressor in many different cancer types to regulate apoptosis. On the other hand, ING3 levels positively correlate with poor survival prognosis of prostate cancer (PCa) patients. In PCa cells, ING3 acts rather as an androgen receptor (AR) co-activator and harbours oncogenic properties in PCa. Here, we show the identification of a novel ING3 splice variant in both the human PCa cell line LNCaP and in human PCa patient specimen. The novel ING3 splice variant lacks exon 11, ING3∆ex11, which results in deletion of the PHD, providing a unique opportunity to analyse functionally the PHD of ING3 by a natural splice variant. Functionally, overexpression of ING3Δex11 induced morphological changes of LNCaP-derived 3D spheroids with generation of lumen and pore-like structures within spheroids. Since these structures are an indicator of epithelial–mesenchymal transition (EMT), key regulatory factors and markers for EMT were analysed. The data suggest that in contrast to ING3, ING3Δex11 specifically modulates the expression of key EMT-regulating upstream transcription factors and induces the expression of EMT markers, indicating that the PHD of ING3 inhibits EMT. In line with this, ING3 knockdown also induced the expression of EMT markers, confirming the impact of ING3 on EMT regulation. Further, ING3 knockdown induced cellular senescence via a pathway leading to cell cycle arrest, indicating an oncogenic role for ING3 in PCa. Thus, the data suggest that the ING3Δex11 splice variant lacking functional PHD exhibits oncogenic characteristics through triggering EMT in PCa cells.