Material collected from a parkway in the city of Chicago afforded the isolation of a Nostoc species (UIC 10022A). The extract of this strain displayed significant inhibition of the 20S proteasome as well as antiproliferative activity against HT29, MCF7, NCI-H460, and SF268 cancer cell lines. A standardized dereplication protocol allowed for the rapid identification of three known (11-13) and nine new (1-9) chlorinated cylindrocyclophanes from less than 100 mg of organic extract. Scale-up isolation of 1-9 and 11-13 from a larger extract was guided by LC-UV-MS data. In addition, KBr enrichment of the culture media afforded the isolation of a brominated cylindrocyclophane (10). Biological evaluation of 1-5, 9, and 10-13 revealed a large range of activity against the 20S proteasome and allowed the determination of preliminary structure-activity relationships of the cylindrocyclophane pharmacophore.