ABSTRACT
Drug-tolerant persister (DTP) cells remain following chemotherapy and can cause cancer relapse. However, it is unclear when acquired resistance to chemotherapy emerges. Here, we compared the gene expression profiles of gastric cancer patient-derived cells (GC PDCs) and their respective xenograft tumors with different sensitivities to 5-fluorouracil (5-FU) by using immunodeficient female BALB/c-nu mice. RNA sequencing analysis of 5-FU-treated PDCs demonstrated that DNA replication/cell cycle-related genes were transiently induced in the earlier phase of DTP cell emergence, while extracellular matrix (ECM)-related genes were sustainably upregulated during long-term cell survival in 5-FU-resistant residual tumors. NicheNet analysis, which uncovers cell-cell signal interactions, indicated the transforming growth factor-ß (TGF-ß) pathway as the upstream regulator in response to 5-FU treatment. This induced ECM-related gene expression in the 5-FU-resistant tumor model. In the 5-FU-resistant residual tumors, there was a marked upregulation of cancer cell-derived TGF-ß1 expression and increased phosphorylation of SMAD3, a downstream regulator of the TGF-ß receptor. By contrast, these responses were not observed in a 5-FU-sensitive tumor model. We further found that TGF-ß-related upregulation of ECM genes was preferentially observed in non-responders to chemotherapy with 5-FU and/or oxaliplatin among 22 patient-derived xenograft tumors. These observations suggest that chemotherapy-induced activation of the TGF-ß1/SMAD3/ECM-related gene axis is a potential biomarker for the emergence of drug resistance in GCs.
Subject(s)
Drug Resistance, Neoplasm , Extracellular Matrix , Fluorouracil , Gene Expression Regulation, Neoplastic , Mice, Inbred BALB C , Signal Transduction , Stomach Neoplasms , Transforming Growth Factor beta , Stomach Neoplasms/genetics , Stomach Neoplasms/metabolism , Stomach Neoplasms/drug therapy , Stomach Neoplasms/pathology , Drug Resistance, Neoplasm/genetics , Humans , Animals , Fluorouracil/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , Female , Signal Transduction/drug effects , Extracellular Matrix/metabolism , Extracellular Matrix/drug effects , Mice , Transforming Growth Factor beta/metabolism , Mice, Nude , Cell Line, Tumor , Smad3 Protein/metabolism , Smad3 Protein/genetics , Xenograft Model Antitumor AssaysABSTRACT
Anticancer drug-tolerant persister (DTP) cells at an early phase of chemotherapy reshape refractory tumors. Aldehyde dehydrogenase 1 family member A3 (ALDH1A3) is commonly upregulated by various anticancer drugs in gastric cancer patient-derived cells (PDC) and promotes tumor growth. However, the mechanism underlying the generation of ALDH1A3-positive DTP cells remains elusive. Here, we investigated the mechanism of ALDH1A3 expression and a combination therapy targeting gastric cancer DTP cells. We found that gastric cancer tissues treated with neoadjuvant chemotherapy showed high ALDH1A3 expression. Chromatin immunoprecipitation (ChIP)-PCR and ChIP sequencing analyses revealed that histone H3 lysine 27 acetylation was enriched in the ALDH1A3 promoter in 5-fluorouracil (5-FU)-tolerant persister PDCs. By chemical library screening, we found that the bromodomain and extraterminal (BET) inhibitors OTX015/birabresib and I-BET-762/molibresib suppressed DTP-related ALDH1A3 expression and preferentially inhibited DTP cell growth. In DTP cells, BRD4, but not BRD2/3, was recruited to the ALDH1A3 promoter and BRD4 knockdown decreased drug-induced ALDH1A3 upregulation. Combination therapy with 5-FU and OTX015 significantly suppressed in vivo tumor growth. These observations suggest that BET inhibitors are efficient DTP cell-targeting agents for gastric cancer treatment. SIGNIFICANCE: Drug resistance hampers the cure of patients with cancer. To prevent stable drug resistance, DTP cancer cells are rational therapeutic targets that emerge during the early phase of chemotherapy. This study proposes that the epigenetic regulation by BET inhibitors may be a rational therapeutic strategy to eliminate DTP cells.
Subject(s)
Aldehyde Oxidoreductases , Drug Resistance, Neoplasm , Fluorouracil , Histones , Stomach Neoplasms , Transcription Factors , Animals , Female , Humans , Male , Mice , Acetylation/drug effects , Aldehyde Oxidoreductases/drug effects , Aldehyde Oxidoreductases/metabolism , Antineoplastic Agents/pharmacology , Bromodomain Containing Proteins/drug effects , Bromodomain Containing Proteins/metabolism , Cell Cycle Proteins/drug effects , Cell Cycle Proteins/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , Fluorouracil/pharmacology , Fluorouracil/therapeutic use , Gene Expression Regulation, Neoplastic/drug effects , Histones/drug effects , Histones/metabolism , Mice, Inbred BALB C , Mice, Nude , Promoter Regions, Genetic/drug effects , Stomach Neoplasms/drug therapy , Stomach Neoplasms/pathology , Stomach Neoplasms/metabolism , Stomach Neoplasms/genetics , Transcription Factors/drug effects , Transcription Factors/metabolism , Xenograft Model Antitumor AssaysABSTRACT
WNT/ß-catenin signaling is aberrantly activated in colorectal cancer (CRC) mainly by loss-of-function mutations in adenomatous polyposis coli (APC) and is involved in tumor progression. Tankyrase inhibitors, which suppress WNT/ß-catenin signaling, are currently in pre-clinical and clinical trials. However, the mechanisms of resistance to tankyrase inhibitors remain unclear. In this study, we established tankyrase inhibitor-resistant CRC cells, JC73-RK100, from APC-mutated patient-derived CRC cells. JC73-RK100 cells and several CRC cell lines were sensitive to tankyrase inhibitors at low concentrations but were resistant at high concentrations, showing an intrinsic/acquired bell-shaped dose response. Mechanistically, tankyrase inhibitors at high concentrations promoted BRD3/4-dependent E2F target gene transcription and over-activated cell cycle progression in these cells. BET inhibitors canceled the bell-shaped dose response to tankyrase inhibitors. Combination of tankyrase and BET inhibitors significantly suppressed tumor growth in a mouse xenograft model. These observations suggest that the combination of tankyrase and BET inhibitors may be a useful therapeutic approach to overcome the resistance of a subset of CRCs to tankyrase inhibitors.
Subject(s)
Adenomatous Polyposis Coli , Antineoplastic Agents , Colorectal Neoplasms , Tankyrases , Animals , Humans , Mice , Adenomatous Polyposis Coli/drug therapy , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , beta Catenin/metabolism , Cell Line, Tumor , Colorectal Neoplasms/drug therapy , Colorectal Neoplasms/genetics , Colorectal Neoplasms/pathology , Disease Models, Animal , Wnt Signaling PathwayABSTRACT
BACKGROUND: Aberrant WNT/ß-catenin signaling drives carcinogenesis. Tankyrases poly(ADP-ribosyl)ate and destabilize AXINs, ß-catenin repressors. Tankyrase inhibitors block WNT/ß-catenin signaling and colorectal cancer (CRC) growth. We previously reported that 'short' APC mutations, lacking all seven ß-catenin-binding 20-amino acid repeats (20-AARs), are potential predictive biomarkers for CRC cell sensitivity to tankyrase inhibitors. Meanwhile, 'Long' APC mutations, which possess more than one 20-AAR, do not predict inhibitor-resistant cells. Thus, additional biomarkers are needed to precisely predict the inhibitor sensitivity. METHODS: Using 47 CRC patient-derived cells (PDCs), we examined correlations between the sensitivity to tankyrase inhibitors (G007-LK and RK-582), driver mutations, and the expressions of signaling factors. NOD.CB17-Prkdcscid/J and BALB/c-nu/nu xenograft mice were treated with RK-582. RESULTS: Short APC mutant CRC cells exhibited high/intermediate sensitivities to tankyrase inhibitors in vitro and in vivo. Active ß-catenin levels correlated with inhibitor sensitivity in both short and long APC mutant PDCs. PIK3CA mutations, but not KRAS/BRAF mutations, were more frequent in inhibitor-resistant PDCs. Some wild-type APC PDCs showed inhibitor sensitivity in a ß-catenin-independent manner. CONCLUSIONS: APC/PIK3CA mutations and ß-catenin predict the sensitivity of APC-mutated CRC PDCs to tankyrase inhibitors. These observations may help inform the strategy of patient selection in future clinical trials of tankyrase inhibitors.