Abstract
Current therapies for treating pancreatic ductal adenocarcinoma (PDAC) are largely ineffective, with the desmoplastic environment established within these tumors being considered a central issue. We established a 3D spheroid co-culture in vitro model using a PDAC cell line (either PANC-1 or Capan-2), combined with stellate cells freshly isolated from pancreatic tumors (PSC) or hepatic lesions (HSC), and human type I collagen to analyze the efficiency of the chemotherapeutic gemcitabine (GEM) as well as two novel drug candidates derived from natural products: pseudopterosin (PsA-D) and O-methyltylophorinidine (TYLO). Traditional 2D in vitro testing of these agents for cytotoxicity on PANC-1 demonstrated IC50 values of 4.6 (±0.47) nM, 34.02 (±1.35) µM, and 1.99 (±0.13) µM for Tylo, PsA-D, and GEM, respectively; these values were comparable for Capan-2: 5.58 (±1.74) nM, 33.94 (±1.02) µM, and 0.41 (±0.06) µM for Tylo, PsA-D, and GEM, respectively. Importantly, by assessing the extent of viable cells within 3D co-culture spheroids of PANC-1 with PSC or HSC, we could demonstrate a significant lack of efficacy for GEM, while TYLO remained active and PsA-D showed slightly reduced efficacy: GEM in PANC-1/PSC (IC50 = >100 µM) or PANC-1/HSC (IC50 = >100 µM) spheroids, TYLO in PANC-1/PSC (IC50 = 3.57 ± 1.30 nM) or PANC-1/HSC (IC50 = 6.39 ± 2.28 nM) spheroids, and to PsA-D in PANC-1/PSC (IC50 = 54.42 ± 12.79 µM) or PANC-1/HSC (IC50 = 51.75 ± 0.60 µM). Microscopic 3D rendering supported these cytotoxicity outcomes, showing little or no morphological spheroid structure change during this period of rapid cell death. Our results support the use of this 3D spheroid co-culture in vitro model having a desmoplastic microenvironment for the identification of possible novel chemotherapeutic drug candidates for PDAC, such as TYLO and PsA-D.
Highlights
Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death globally, having an extremely high mortality to incidence ratio [1]
We examined the efficacy of pseudopterosin A-D (PsA-D), O-methyltylophorinidine (TYLO), or gemcitabine (GEM) at 48 h in the pancreatic ductal adenocarcinoma (PDAC) cells Capan-2 or PANC-1, when grown in a standard 2D format (Figure 3, Table 2)
Since PDAC tumors are typically poorly vascularized and desmoplastic, it is possible that the lack of clinical benefit following the administration of the currently approved chemotherapeutics could be due to insufficient penetration and/or reduced cancer cell susceptibility
Summary
Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death globally, having an extremely high mortality to incidence ratio [1]. In a Phase 1b/2 clinical trial (NCT01130142), the nonselective elimination of tumor stroma by sonic hedgehog inhibitor (IPI-926) has led to the rapid dissemination of cancer cells, causing accelerated patient death [6]. The elucidation of this unexpected result showed that the fibrotic stroma serves as a barrier to restrain the cancer [7]. Chemotherapeutics to improve the unmet medical need of effectively targeting the unique PDAC microenvironment are unlikely to be readily identified using 2D high-throughput in vitro screens with cancer cells grown on plastic substrates, urging the use of 3D cell-based models including elements of the fibrotic stroma microenvironment and reflecting the 3D tumor architecture [8,9]
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