Irinotecan activity in vivo

We used 20 of the new PDX models to evaluate the activity of irinotecan, as a model drug with established activity against pediatric solid tumors including sarcomas [¹⁰, ²⁸, ²⁹]. We reasoned that if the PDXs were truly representative of the patients' diseases, they would likely respond to this drug and such response should remain stable over successive PDX generations. Among the patients from which these PDX models were established, 11 Ewing sarcoma PDXs resulted from eight patients who received irinotecan as part of their rescue treatment (combined with temozolomide, with or without additional vincristine or trabectedin); 7 rhabdomyosarcoma PDXs from six patients who received irinotecan as part of the upfront treatment of the primary tumor (either alone or combined with carboplatin or vincristine); and 2 osteosarcoma PDXs from two patients who did not receive irinotecan. To compare responses to treatment of patients and PDXs, we selected newly diagnosed patients treated with irinotecan as upfront treatment and with available PDX models from their primary tumor. We evaluated treatment response in such patients using the responsive evaluation criteria in solid tumors (RECIST) protocol or the metabolic tumor volume.

For all efficacy studies in mice, we inserted freshly excised PDX tumors (obtained from one mouse of the immediate earlier generation) in both flanks of athymic nude or NOD‐SCID mice. Upon engraftment (tumor volumes ranging 100–500 mm³), mice were distributed to control or treatment groups, with care that tumor volume means and standard deviations (STDEV) in both groups were similar. Treatment groups received one cycle of 10 mg/kg/day irinotecan (Hospira, Lake City, IL, USA) in a 5‐day‐on‐2‐off regimen, intraperitoneal, for two consecutive weeks, as previously described [³⁰]. Control groups received saline using the same regimen as treated groups. Tumor volume was measured three times a week, until day 14, in which response to treatment was evaluated as previously described [¹⁰]. We defined complete response (CR) as tumor mass <50 mm³ and >50% reduction at the end of treatment (day 14); partial response (PR) as tumor volume regression ≥50% at day 14 but tumor volume ≥50 mm³; stable disease (SD) as <50% regression and ≤25% increase in initial volume at day 14; and progressive disease as <50% regression from initial volume and >25% increase in initial volume at day 14.

To address whether the response to the drug changed upon consecutive passages of the PDX in mice, we evaluated the response to irinotecan of different filial generations in three of the models, one early after initial engraftment (F ≤ 2) and the second one later (F ≥ 6). After one cycle of irinotecan, we followed up all animals weekly until tumor regrowth to endpoint (1,500 mm³) or day 100 to build Kaplan–Meier curves and estimate median survival times. We used the log‐rank test to compare survival curves between treatments and between different generations from the same PDX model.