We have utilized a recently developed human tumor cloning system to screen for antitumor effects in vitro of a new anthracenedione derivative, Mitoxantrone. The object was to determine if the system is useful for pinpointing the types of tumors in patients which should be studied in early Phase II clinical trials. Tumors from 267 patients were placed in culture (20 different histological tumor types). One hundred seventy tumors both grew and formed enough colonies for drug sensitivity assays. Excellent in vitro antitumor activity was noted for Mitoxantrone against human adenocarcinoma of the lung, small cell lung cancer, melanoma, and biliary tree cancer. Good antitumor activity was noted against breast cancer, ovarian cancer, non-Hodgkin's lymphoma, head and neck cancer, squamous cell lung cancer, soft tissue sarcoma, gastric cancer, and hepatomas. The drug showed no in vitro activity against colon cancer. These data indicate that Mitoxantrone has a wide spectrum of in vitro antitumor activity. A comparison of these in vitro results with the results of Phase II clinical trials with the drug should allow an evaluation of the utility of the human tumor cloning system for predicting clinical antitumor activity of a new compound.