In vitro anticancer effects of a novel immunostimulant: keyhole limpet hemocyanin

Keyhole limpet hemocyanin (KLH) is a recently described immune stimulant and hapten carrier derived from a circulating glycoprotein of the marine mollusk Megathura crenulata. It has been reported to be a potent form of intravesical immunotherapy for the treatment of transitional cell carcinoma of the bladder and has been used in a variety of genitourinary tumors. We hypothesized that KLH would be effective against other cancer cells in vitro.

Multiple cancer cell lines were tested, including estrogen-dependent breast (MCF-7), estrogen-independent breast (ZR75-1), pancreas (PANC-1, MIA-PaCa), and prostate (DU145). Serial twofold dilutions of KLH were prepared in sterile 96-well plates. Dose-response curves were performed beginning with a concentration of 100 microg of KLH/well and ending at a concentration of 0.8 ng/well. Cells were added at concentrations of 5 x 10(4) cells per well. Cell viability was evaluated at 24 and 72 h by MTT assay at an absorbance of 570 nm.

Significant (P < 0.05) cancer cell growth inhibition was observed in four of the five cell lines tested at both time treatment intervals. The breast cancer line ZR75-1 exhibited a mean growth inhibition of 43 +/- 1.1% (range 37 to 59%) at 72 h, whereas treated MCF-7 cells had an average of 39 +/- 9.1% growth inhibition (range 35 to 44%) at these same concentrations. Treated PANC-1 cells had a mean growth inhibition of 19 +/- 0.8% (range 4 to 46%) at 72 h. The DU145 prostate cancer cell line averaged a 6 +/- 1.3% growth inhibition (range -19 to 55%) over the concentrations tested.

The direct growth inhibition of multiple tumor cell-lines exhibited by KLH is significant and warrants further in vitro mechanistic studies and in vivo experiments. Investigation into the efficacy and mechanism of response could directly lead to more effective treatment regimens for patients suffering from these diseases.