Recently, the search for novel therapeutic agents against Acanthamoeba species has been focused on the evaluation of natural resources. Among them, marine microorganisms have risen as a source of bioactive compounds with the advantage of the ability to obtain unlimited and constant amounts of the compounds in contrast to other natural sources such as plants. Furthermore, marine actinomycetes have recently been reported as highly rich in bioactive agents including salinosporamides, xiamycines, indolocarbazoles, naphtyridines, phenols, dilactones such as antimycines and macrolides among others. In this study, staurosporine (STS) was isolated from a strain of Streptomyces sanyensis and tested against Acanthamoeba to characterize the therapeutic potential of STS against this protozoan parasite. We have established that STS is active against both stages of the Acanthamoeba life cycle, by the activation of Programmed Cell Death via the mitochondrial pathway of the trophozoite. We have also established that STS has relatively low toxicity towards a macrophage cell line. However, previous studies have highlighted higher toxicity levels induced on other vertebrate cell lines and future research to lower these toxicity issues should be developed.