The paper reviews the current approaches to cytotoxic effect screening and mode of action of natural and synthetic compounds usable in tumour chemotherapy. Oncological diseases belong to the most frequently occurring and the most serious diseases that threaten millions of human lives. A serious complication of anticancer therapy is an inadequate therapeutic answer which is caused by the resistance of the human organism to the employed drugs, chemotherapeutic agents. Therefore the resistance is one of the driving forces which constantly force us to search for new anticancer effective drugs of natural or synthetic origin. Primary screening in vitro, in which by different methods the sensitivity of cancer cells growing in vitro to cytotoxic compounds is monitored, is commonly and routinely used for searching for new potential cytostatics today. The properties of a potential anticancer compound are characterized by different parameters, which include antiproliferative activity, monitoring of structural and functional changes in the cytoplasmic membrane, changes in cell proteins and nucleic acids content, cell metabolism changes, cell cycle changes, induction of apoptosis, enzymatic activity (dihydrofolate reductase, proteinases, proteinkinases, topoisomerases, tymidylatsynthetase), effect on mitochondria, cell cytoskeleton, telomerase activity, etc. By monitoring these parameters, the mode of action of a cytotoxically effective compound can be followed up.