Xenograft models for parental and chemoresistant SCLC tumors

Subcutaneous xenograft in NSG mice was derived from direct implantation of untreated H1436 cells or re-implantation of chemoresistant tumors after 4 cycles of Cisplatin and Etoposide (EC), or plus 4 cycles of Cisplatin (reduced from EC due to toxicity). Specifically, a million H1436 cells were resuspended in 100 µl mixture of serum-free RPMI-1640 and Matrigel (BD Bioscience #356237) at 1:1 ratio and immediately injected in the flank of 6–8-week-old female NSG mouse (Jackson Laboratory #005557). Mice were randomized after tumor cell injection. Treatment starts after a week when the tumor becomes palpable. 5 mg/kg/w Cisplatin (Sigma P4394) in saline, 10 mg/kg/w Etoposide (Sigma E1383) in 30% PEG 300 (Sigma 202371) were freshly prepared and administered by intraperitoneal injection, for 4 cycles in total to obtain the first group of chemoresistant tumors. An additional 4 cycles of Cisplatin were administered in the second group of mice to obtain tumors with further potentiated chemoresistance. To harvest the tumor, 10 ml digestion media was used per mouse. This was prepared freshly by supplementing 9 ml HBSS with 1 ml type IV collagenase, 50 µl DNase II and 50 µl 1 M CaCl₂. Tumors were collected and placed in HBSS immediately following dissection. A fraction of the tumor was cut into a few pieces and flash-frozen in liquid nitrogen to be saved in aliquots for molecular assays. The remaining chunk was finely minced with a sterile scalpel blade. For re-implantation, the minced tissue was resuspended in digestion media, rotated at 37 °C for 20 min, filtered through a 40 µm filter, centrifuged at 300 × g for 5 min.