Ab120554

Restek phytocannabinoid standards were used in this study: ∆-9 tetrahydrocannabinol (Δ-9 THC, 34067), cannabichromene (CBC, 34092), cannabidiol (CBD, 34011) at a concentration of 1 mg/mL, originally dissolved in methanol. For quantification of phytocannabinoids the standards were dissolved in methanol at various concentrations from 0–25 μg/mL. Mitomycin-C (MMC) was dissolved in water at a stock concentration of 800 μg/mL. Inverse agonists (IA) CB1 (AM251, Abcam ab120088), CB2 (SR144528, Abcam ab146185), TRPA1 blocker (HC-030031, Abcam ab120554) and TRPV1 and TRPV2 antagonists (Abcam ab141772 and Tranilast 1098/10, respectively; Tranilast is a TRPV2 inhibitor [40 (link)]) were dissolved in dimethyl sulfoxide (DMSO) at a concentration of 10mM.

Standard/Material preparation and use was done, as described previously [19 (link),20 (link)]. Briefly, the cannabinoid standards at a concentration of 1 mg/mL in methanol used in this study included THC (34067; Restek, Bellefonte, PA, USA), CBC (34092; Restek), CBG (34091; Restek) and CBN (34010; Restek). Inverse agonists (IA) to CB1 and CB2 were AM251 (ab120088; Abcam, Cambridge, UK) and SR144528 (ab146185; Abcam), respectively. The TRPA1 blocker used was HC-030031 (ab120554; Abcam). TRPV1 and TRPV2 antagonists were ab141772 (Abcam) and Tranilast 1098/10 (Abcam), respectively. All IAs, the blocker and antagonists were dissolved in dimethyl sulfoxide (DMSO) at a concentration of 10 mM and used for cell treatment at a final concentration of 10 µM. Chemotherapy solutions for synergy tests included niraparib (AG0038ZU; Angene, Nanjing, China) and gemcitabine (461060010; Acros Organics, Beijing, China) both dissolved in DMSO. Solutions for induction of malignant features [21 (link)] included IL-1β (200-01B; PeproTech, Cranbury, NJ, USA) and TNFα (300-01A; PeproTech) both dissolved in pure water. Solvents (methanol and/or DMSO) were used as a negative control and niraparib was used as a positive control in all the biological assays. The control in any given experiment was set at a concentration to match the vehicle concentration in the highest concentration treatment.

The cannabinoid standards (at a concentration of 1 mg/mL in methanol) used in this study included cannabidiol (CBD, Restek catalog no. 34011), cannabigerol (CBG, Restek catalog no. 34091), tetrahydrocannabivarin (THCV, Restek catalog no. 34100), cannabinol (CBN, Restek catalog no. 34010),a and delta-9-tetrahydrocannabidiol (Δ-9 THC, Restek catalog no. 34067). Inverse agonists (IA) to CB1 and CB2 used included AM251 (ab120088; Abcam) and SR144528 (ab146185; Abcam), respectively. The transient receptor potential ankyrin subtype 1 protein (TRPA1) blocker used was HC-030031 (ab120554; Abcam). Transient receptor potential vanilloid receptor 1 (TRPV1) and 2 (TRPV2) antagonists were SB-366791 (ab141772-5-B Abcam) and Tranilast (1098/10 Abcam), respectively. All IAs were dissolved in dimethyl sulfoxide (DMSO) at a concentration of 10 mM. Doxorubicin (D1515; Sigma Aldrich, St. Louis, MO, USA) served as a positive control in concentrations of 0.5 µg/mL on A172 cells and 50 µg/mL on U87. Temozolomid (TMZ, T2577; Sigma Aldrich, St. Louis, MO, USA) was tested as a positive control. Analytical grade methanol was used according to the indicated concentration of the treatment. Ultra-pure deionized water (MS grade) was used as received without further purification.