Increased human cathepsin L activity is linked to invasive and metastatic cancers where it promotes degradation of the extracellular matrix. This major cysteine protease found in cell lysosomes and secreted from tissues, also plays a role in the pathology of degenerative cartilage and neurological disorders, and is reported to be required for the SARS coronavirus infection. A library of 59 small non-peptidic thiosemicarbazone and α, β-unsaturated carbonyl derivatives of benzophenone, propiophenone, α- and β-tetralone, 4-chromanone, and 4-thiochromenone were evaluated as inhibitors of human cathepsin L. While most of the compounds had IC50 values in the range of 0.4 μM or greater, four were very effective inhibitors of cathepsin L: the benzophenone thiosemicarbazones 2 (IC₅₀ = 1.5 nM), 55 (IC₅₀ = 44 nM), 38 (IC₅₀ = 60 nM), 32 (IC₅₀ = 66 nM), and 37 (IC₅₀ = 140 nM) and a sulfone analog of the bromo substituted thiochroman-4-one 22 (IC₅₀ = 1 nM). Kinetics studies were used to gain understanding in enzymeinhibitor interactions of the most potent compounds (2 and 22) and they were found to be reversible, slow, tight binding inhibitors of cathepsin L. These data support formation of a transient covalent intermediate between thiosemicarbazone inhibitors and the cathepsin L active site thiolate. Ten of the most promising lead compounds were also tested for cytotoxicity in HEK-293 cells and generated no toxicity after 24 hours. Exposure of the prostate cancer cell line DU-145 to the most promising lead compounds successfully decreased the invasiveness and mobility properties of these cells in vitro. The non-peptidic nature of these inhibitors, coupled with their cell-based activity, makes these compounds very promising leads for the development of selective cathepsin L inhibitors. A separate research project consisted of recombinant cruzain purification and evaluation of thiosemicarbazone derivatives as potential inhibitors of this parasitic cysteine protease. Cruzain is the major cysteine protease of Trypanosoma cruzi organism and is a validated therapeutic target for the development of new chemotherapy. Chagas disease, a result of Trypanosoma cruzi infection, is the third largest parasitic disease challenge worldwide after malaria and leishmania and there is an urgent need for development of new therapeutic agents against Chagas disease. From the same library of thiosemicarbazone derivatives evaluated against cathepsin L, 25 compounds were evaluated against cruzain from which six compounds were in the nanomolar range with IC50 values ranging from 170 nM to 622 nM.