Andrographis Reverses Gemcitabine Resistance through Regulation of ERBB3 and Calcium Signaling Pathway in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies, primarily due to intrinsic or acquired resistance to chemotherapy, such as Gemcitabine (Gem). Naturally occurring botanicals, including Andrographis (Andro), can help enhance the anti-tumorigenic therapeutic efficacy of conventional chemotherapy through time-tested safety and cost-effectiveness. Accordingly, we hypothesized that Andro might reverse Gem resistance in PDAC. The critical regulatory pathways associated with Gem resistance in PDAC were identified by analyzing publicly available transcriptomic profiling and PDAC tissue specimens. A series of systematic in vitro experiments were performed using Gem-resistant (Gem-R) PDAC cells and patient-derived 3D-organoids to evaluate the Andro-mediated reversal of Gem resistance in PDAC. Transcriptomic profiling identified the calcium signaling pathway as a critical regulator of Gem-resistance (Fold enrichment: 2.8, p = 0.002). Within this pathway, high ERBB3 expression was significantly associated with poor prognosis in PDAC patients. The combination of Andro and Gem exhibited superior anti-cancer potential in Gem-R PDAC cells through potentiating cellular apoptosis. The combined treatment down-regulated ERBB3 and decreased intracellular calcium concentration in Gem-R PDAC cells. Finally, these findings were successfully interrogated in patient-derived 3D-organoids. In conclusion, we demonstrate novel evidence for Andro-mediated reversal of chemoresistance to Gem in PDAC cells through the regulation of ERBB3 and calcium signaling.

Berberine Overcomes Gemcitabine-Associated Chemoresistance through Regulation of Rap1/PI3K-Akt Signaling in Pancreatic Ductal Adenocarcinoma.

Gemcitabine (Gem)-based chemotherapy is one of the first-line treatments for pancreatic ductal adenocarcinoma (PDAC). However, its clinical effect is limited due to development of chemoresistance. Various naturally occurring compounds, including Berberine (BBR), provide an anti-cancer efficacy with time-tested safety, individually and in combination with chemotherapeutic drugs. Accordingly, we hypothesized that BBR might enhance the chemosensitivity to Gem in PDAC. In this study, cell culture studies using MIA PaCa-2 and BxPC-3 cells, followed by analysis in patient-derived organoids were performed to evaluate the anti-cancer effects of BBR in PDAC. Considering that cancer is a significant manifestation of increased chronic inflammatory stress, systems biology approaches are prudent for the identification of molecular pathways and networks responsible for phytochemical-induced anti-cancer activity, we used these approaches for BBR-mediated chemosensitization to Gem. Firstly, Gem-resistant (Gem-R) PDAC cells were established, and the combination of BBR and Gem revealed superior anti-cancer efficacy in Gem-R cells. Furthermore, the combination treatment induced cell cycle arrest and apoptosis in Gem-R PDAC cells. Transcriptomic profiling investigated the Rap1 and PI3K-Akt signaling pathway as a key regulator of Gem-resistance and was a key mediator for BBR-mediated chemosensitization in PDAC cells. All cell culture-based findings were successfully validated in patient-derived organoids. In conclusion, we demonstrate that BBR-mediated reversal of chemoresistance to Gem manifests through Rap1/PI3K-Akt signaling in PDAC.

Preclinical testing of oncolytic adenovirus sensitivity in patient-derived tumor organoids.

Patient-derived organoids (PDOs) have shown the potential to reflect patient sensitivity to chemotherapeutic or targeted drugs. Recently, we showed that organoid models can also serve as a platform to screen for selectivity and potency of oncolytic adenoviruses (OAds). In this protocol, we describe the steps for tumor organoid adenoviral infection and functional assessment of patient-specific responses to OAds. We provide methods to determine OAd relative efficacy by evaluation of PDO viability after infection and adenoviral replication within cancer cells. For complete details on the use and execution of this protocol, please refer to Raimondi et al. (2020).

Patient-derived pancreatic tumour organoids identify therapeutic responses to oncolytic adenoviruses.

BACKGROUND: Pancreatic patient-derived organoids (PDOs) are a well-established model for studying pancreatic ductal adenocarcinoma (PDAC) carcinogenesis and are potential predictors of clinical responses to chemotherapy. Oncolytic virotherapy is envisioned as a novel treatment modality for pancreatic cancer, and candidate viruses are being tested in clinical trials. Here, we explore the feasibility of using PDOs as a screening platform for the oncolytic adenovirus (OA) response. METHODS: Organoids were established from healthy pancreas and PDAC tissues and assessed for infectivity, oncoselectivity, and patient-dependent sensitivity to OA. Antitumour effects were studied in vivo in organoid xenografts. Further evaluation of oncolytic responses was conducted in organoids derived from orthotopic models or metastastic tissues. FINDINGS: Oncolytic adenoviruses display good selectivity, with replication only in organoids derived from PDAC tumours. Furthermore, responses of PDOs to a set of OAs reveal individual differences in cytotoxicity as well as in synergism with standard chemotherapy. Adenoviral cytotoxicity in PDOs is predictive of antitumour efficacy in a subcutaneous xenograft setting. Organoids from orthotopic tumours and metastases in nude mice mirror the viral preference of PDOs, indicating that PDO sensitivity to OAs could be informative about responses in both primary tumours and metastatic foci. INTERPRETATION: Our data imply that pancreatic PDOs can serve as predictive tools for screening for sensitivity to OA.