Approximately 5x103 T3M4 and 2x104 Capan-2 cells were seeded in each well of a 96-well plate. After overnight incubation, culture medium was replaced by dilution range of chemotherapies for 72 hours. The chemotherapies used were 5-Fluorouracil (5-FU), irinotecan, oxaliplatine and gemcitabine (Accord Healthcare, Lille, France). Then, cells were fixed with 70% ethanol for 10 minutes, followed by 0.2% crystal violet in 20% ethanol for 15 minutes. After three washes, 0.1% acetic acid in 50% ethanol was added in each well and optical density was measured at 540 nm using a microplate reader (Multiskan As-cent; Thermo Fisher Scientific). The half-maximal inhibitory concentration (IC50) values of each chemotherapy were determined by a non-linear regression using GraphPad Prism 9® (GraphPad Software, La Jolla, CA, USA). All experiments were performed in triplicates in three independent assays.
Irinotecan
Manufactured by Accord Healthcare
Sourced in France, Spain, Switzerland, Netherlands
Irinotecan is a synthetic chemical compound used as a pharmaceutical active ingredient in drug formulations. It is a topoisomerase I inhibitor, which means it interferes with the enzyme topoisomerase I that is involved in the replication and transcription of DNA. Irinotecan is commonly used in the treatment of various types of cancer.
Automatically generated - may contain errors
Lab products found in correlation
Multiskan ascent, by Thermo Fisher Scientific (1 mentions)
Gemcitabine, by Accord Healthcare (1 mentions)
5 fluorouracil, by Accord Healthcare (1 mentions)
Prism 9, by GraphPad (1 mentions)
El10a, by Biobase (1 mentions)
Bleomycin sulfate, by Merck Group (1 mentions)
Doxorubicin, by Merck Group (1 mentions)
Dulbecco s phosphate, by Thermo Fisher Scientific (1 mentions)
Fluorouracil, by Accord Healthcare (1 mentions)
4 protocols using irinotecan
1
Chemotherapeutic Sensitivity Assay for T3M4 and Capan-2 Cells
2
Evaluating Chemotherapeutic Drug Cytotoxicity
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A thiazolyl blue tetrazolium bromide (MTT, BioChem, PanreacApplichem, Barcelona, Spain) assay was performed to detect the viability of the previously oxaliplatin-treated and non-treated cells exposed to different concentrations of oxaliplatin (050 µg/mL, 10 serial 1:2 dilutions starting from the maximum doses) and irinotecan (Accord Healthcare S.L.U., Barcelona, Spain) (030 µM, four serial 1:2 dilutions from the maximum doses). Briefly, cells were seeded in 96-well plates (Deltalab S.L, Barcelona, Spain) at a concentration of 200,000 cells/mL in a volume of 100 µL. Next, the chosen chemotherapeutic drug was added to each well. Three replicates per condition were performed in each plate with three independent experiments in total. After 72 h of the treatment, the medium was removed, and 50 µL of 0.5 mg/mL MTT was added. Then, plates were incubated for 4 h at 37 °C, and finally, the medium was removed and resuspended in 100 µL of dimethyl sulfoxide (DMSO, Labkem, Barcelona, Spain) before detection of absorbance in a BIOBASE-EL 10A (Biobase, Shandong, China) spectrophotometer at 546 nm.
3
Microbubble Delivery of Anti-Cancer Drugs
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Bleomycin sulfate (B1141000; Sigma-Aldrich, St Louis, MO) and doxorubicin (D2975000; Sigma Aldrich, St Louis, MO) were dissolved in Dulbecco's phosphate-buffered saline solution (DPBS; Thermo Fisher Scientific) at a 1 mM stock solution. Irinotecan (Accord Health care AG, Bottmingen, Switzerland) was a generous gift from Dr. Virginie André (Regional Center of Cancerology, Henry Kaplan, CHRU de Tours, France), and its concentration was 20 mg/mL. Vevo MicroMarker ® contrast agents were purchased from VisualSonics-Fujifilm Inc.
(Toronto, Canada) [20, (link)21] (link). These agents are MBs consisting of a gaseous core of nitrogen and a perfluorobutane mixture encapsulated in by a PEGylated phospholipid shell [22] (link). As previously reported in our in-vitro and in-vivo studies, these MBs are the most effective in delivering therapeutic molecules including anticancer drugs and plasmid DNA [21] (link)[22] (link)[23] (link). They were prepared according to the manufacturer's instructions at a final concentration of 2.10 9 MB/mL.
(Toronto, Canada) [20, (link)21] (link). These agents are MBs consisting of a gaseous core of nitrogen and a perfluorobutane mixture encapsulated in by a PEGylated phospholipid shell [22] (link). As previously reported in our in-vitro and in-vivo studies, these MBs are the most effective in delivering therapeutic molecules including anticancer drugs and plasmid DNA [21] (link)[22] (link)[23] (link). They were prepared according to the manufacturer's instructions at a final concentration of 2.10 9 MB/mL.
4
Optimizing FOLFIRINOX Dosing for Tumors
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The cytotoxicity of the drugs toward the KPC cell line was confirmed by AlamarBlue assay (Supplementary Section S.1, online only). To determine the optimal dose for therapeutic effect and tolerable toxicity, five doses of FOLFIRONOX were injected into mice with heterotopic tumors. The administration of FOLFIRINOX differed from clinical practice as all drugs were administered mixed in one vial as a bolus, whereas clinically the drugs are given sequentially as a 46-h infusion. On the basis of published results with a comparable tumor model [29] , calcium folinate (Pfizer, Zaventem, Belgium), oxaliplatin (SUN Pharma, Hoofddorp, Netherlands), irinotecan (Accord Healthcare, Middlesex, UK) and fluorouracil (Accord Healthcare, Middlesex, UK) were mixed and 100 μL was injected using a 24-gauge lateral tail vain catheter. The doses used are given in Table 1 (doses A-C). Treatment was administered on days 9 and 13 after tumor cell inoculation. All animals were killed 19 d after inoculation. Next, an additional two doses of FOLFIRINOX were injected based on calculations from clinical doses given in established protocols at St. Olav's hospital to equivalent mouse doses using a converting table [30] (doses E and F, Table 1). Treatment was administered on days 9
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