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Purpose: Presence of tumor-associated macrophages (TAM) and high levels of ferritin and lipocalin 2 (Lcn2) in the tumor microenvironment are associated with poor prognosis in many types of cancer. Here we investigate whether iron deprivation influences TAM phenotype and chemotherapy resistance in tumor slice cultures (TSC) of gastric cancer. Results: TAM remained morphologically and functionally stable for four DIV. DFO treatment for 72 h decreased ferritin expression in TAM and in the tumor stroma but did not alter Lcn2 expression. TAM phenotype was altered after 72 h of cisplatin or DFO treatment compared with control conditions. Single DFO treatment and combined treatment with cytotoxic drugs significantly increased tumor cell apoptosis in TSC of gastric cancer. Methods: TSC were manufactured by cutting tissue of gastric cancer resection specimens in 350 μm thick slices and cultivating them under standard conditions on a filter membrane, at an air-liquid interface. After 24 h ex vivo , TSC were treated with irinotecan (100 nM) or cisplatin (10 μM) alone and in combination with deferoxamine (DFO; 10 μM, 100 μM), respectively, for 72 h. After four days in vitro (DIV) the TSC were fixated with paraformaldehyde, paraffin embedded and analyzed by immunohistochemistry for apoptosis (cPARP), proliferation (Ki67), TAM (CD68, CD163), ferritin, and Lcn2 expression. Conclusions: TAM are well preserved and can be studied in TSC of gastric cancer. Iron deprivation significantly increased tumor cell apoptosis.

Despite novel immunotherapies being approved and established for the treatment of non-small cell lung cancer (NSCLC), ex vivo models predicting individual patients’ responses to immunotherapies are missing. Especially immune modulating therapies with moderate response rates urge for biomarkers and/or assays to determine individual prediction of treatment response and investigate resistance mechanisms. Here, we describe a standardized ex vivo tissue culture model to investigate individual tumor responses. NSCLC tissue cultures preserve morphological characteristics of the baseline tumor specimen for up to 12 days ex vivo and also maintain T-cell function for up to 10 days ex vivo. A semi-automated analysis of proliferating and apoptotic tumor cells was used to evaluate tissue responses to the PD-1 inhibitor nivolumab (n = 12), from which two cases could be successfully correlated to the clinical outcome. T-cell responses upon nivolumab treatment were investigated by flow cytometry and multispectral imaging. Alterations in the frequency of the Treg population and reorganization of tumor tissues could be correlated to nivolumab responsiveness ex vivo. Thus, our findings not only demonstrate the functionality of T cells in NSCLC slice cultures up to 10 days ex vivo, but also suggests this model for stratifying patients for treatment selection and to investigate in depth the tumor-associated T-cell regulation.

Emerging immunotherapies quest for better patient stratification in cancer treatment decisions. Moderate response rates of PD-1 inhibition in gastric and esophagogastric junction cancers urge for meaningful human model systems that allow for investigating immune responses ex vivo . Here, the standardized patient-derived tissue culture (PDTC) model was applied to investigate tumor response to the PD-1 inhibitor Nivolumab and the CD3/CD28 t-lymphocyte activator ImmunoCult TM . Resident t-lymphocytes, tumor proliferation and apoptosis, as well as bulk gene expression data were analyzed after 72 h of PD-1 inhibition either as monotherapy or combined with Oxaliplatin or ImmunoCult TM . Individual responses to PD-1 inhibition were found ex vivo and combination with chemotherapy or t-lymphocyte activation led to enhanced antitumoral effects in PDTCs. T-lymphocyte activation as well as the addition of pre-cultured peripheral blood mononuclear cells improved PDTC for studying t-lymphocyte and tumor cell communication. These data support the potential of PDTC to investigate immunotherapy ex vivo in gastric and esophagogastric junction cancer.