Two previously undescribed indolocarbazole alkaloids, bresmycins A (1) and B (2) were isolated from the culture extract of the marine mesophotic zone sponge-associated Streptomyces sp. NBU3142. Their structures were assigned by using the detailed NMR and MS spectroscopic analyses. Their relative and absolute configurations were established by coupling constants, NOESY correlations and comparison of experimental ECD spectra. Biological evaluation revealed that compounds 1 and 2 exhibited significant cytotoxicity against breast cancer cell line MCF-7 with IC50 values in the range of 2.01-7.08 μM. In addition, compound 2 was found to display a strong inhibitory effect (IC50 = 4.47 μM) against adriamycin-resistant MCF-7 (MCF-7/ADR) cells. The structure−activity differences of bresmycins A and B suggested that an additional N-methyl group in O-acetylhydroxylamine moiety should be the key factors to their anticancer activities.