The immune checkpoint molecules such as PD-L1 and PD-L2 have a substantial contribution to cancer immunotherapy including breast cancer. Microarray expression profiling identified several molecular subtypes, namely luminal-type (with a good-prognosis), HER2-type (with an intermediate-prognosis), and triple-negative breast cancer (TNBC)-type (with a poor-prognosis). We found that PD-L1 and PD-L2 mRNA expressions were highly expressed in TNBC-type cell lines (HCC1937, MDA-MB-231), moderately expressed in HER2-type cell line (SK-BR-3), and poorly expressed in luminal-type cell lines (MDA-MB-361, MCF7). The PD-L1 and PD-L2 expression in SK-BR-3 cells, but not those in HCC1937 and MDA-MB-231 cells, decreased by nicotine stimulation in a dose-dependent manner. In addition, nicotine treatment decreased the phosphorylation of Akt in SK-BR-3 cells, but not in other cell lines. These results show that nicotine regulates the expression of immune checkpoint molecules, PD-L1 and PD-L2, via inhibition of Akt phosphorylation. This findings may provide the new therapeutic strategies for the treatment of breast cancer.