Immunotherapy with Keyhole Limpet Hemocyanin: Efficacy and Safety in the MB-49 Intravesical Murine Bladder Tumor Model

Abstract

The antitumor activity and potential toxicity of a clinical-grade keyhole limpet hemocyanin preparation (KLH-Immune Activator; KLH-IA) were determined in the MB-49 intravesical murine bladder tumor model. Mice were immunized subcutaneously with KLH-IA two weeks prior to intravesical implantation of MB-49 tumor cells. Treatment consisted of intravesical KLH-IA (10 or 100 micrograms.) 1, 4, 7, 14 and 21 days after implantation. Control animals either were not immunized prior to tumor implantation and KLH-IA treatment, or were immunized with KLH-IA and treated with the vehicle. By 4 weeks after implantation tumor outgrowth in the treated groups was significantly decreased (p < 0.01, Fisher's Exact) relative to the control groups. Prior subcutaneous immunization was required to elicit antitumor activity of KLH-IA; thus, the mechanism of action is immune-mediated and not due to spurious interference with tumor implantation by intravesical instillations. Animals treated with a dissociated form of KLH exhibited decreased tumor outgrowth approaching, but not attaining, significance (p < 0.09, Fisher's Exact). A separate toxicity study in which KLH-IA was given subcutaneously (4 mg./kg.), intraperitoneally (40 mg./kg.), or intravesically (40 mg./kg.) disclosed no significant gross or histopathologic abnormalities except for mild-to-moderate papillary hyperplasia in all catheterized animals. These results establish the efficacy and safety of KLH-IA in mice and suggest that clinical trials for intravesical treatment of superficial bladder cancer may be warranted.

Keyhole Limpet Haemocyanin in Experimental Bladder Cancer

“…Subsequent clinical and experimental studies were initiated to confirm this effect [8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21]. In the following, we give an overview of the current available experimental data and compare these with our own results.…”

“…The antitumour activity and potential toxicity of a clinical-grade KLH preparation (KLH immune activator; KLH-IA) were determined in the MB-49 intravesical murine bladder tumour model by Swerdlow et al [19]. Mice were immunised subcutaneously with KLH-IA 2 weeks prior to intravesical implantation of 2 × 10 4 MB-49 tumour cells.…”

“…Animals treated with a dissociated form of KLH exhibited decreased tumour growth approaching, but not attaining, significance (p < 0.09, Fisher’s exact test). A separate toxicity study in which KLH-IA was given subcutaneously (4 mg/kg), intraperitoneally (40 mg/kg) or intravesically (40 mg/kg) disclosed no significant gross or histopathologic abnormalities except for mild to moderate papillary hyperplasia in all catheterised animals [19]. …”

Hemocyanins in the Immunotherapy of Superficial Bladder Cancer

“…Animals treated with a dissociated form of KLH showed decreased tumor outgrowth, but this was not significant. A separate toxicity study in which KLH-IA was given subcutaneously (4 mg/kg), intraperitoneally (40 mg/kg) or intravesically (40 mg/kg) reported no significant gross or histopathological abnormalities, except for mild to moderate papillary hyperplasia in all catheterized animals (Swerdlow et al, 1994). A third model developed by Recker and collaborators also showed the effectiveness of KLH.…”

Cytotoxic Effects of Rapana venosa Hemocyanin on Bladder Cancer Permanent Cell Lines

“…Briefly, the culture medium was removed, and the cells were washed twice with PBS (Gibco) for 5 min and then covered with Annexin V labeling solution. After adding 1 µl of PI solution and 2 µl of fluorescein-annexin solution, the plates were incubated for 15 min at room temperature (15)(16)(17)(18)(19)(20) o C) and then analyzed by fluorescent microscopy.…”