MUC1 expression in primary and metastatic pancreatic cancer cells for in vitro treatment by (213)Bi-C595 radioimmunoconjugate.

Control of micrometastatic pancreatic cancer remains a major objective in pancreatic cancer treatment. The overexpression of MUC1 mucin plays an important role in cancer metastasis. The aim of this study was to detect the expression of MUC1 in human primary tumour tissues and three pancreatic cancer cell lines (CAPAN-1, CFPAC-1 and PANC-1), and target MUC1-positive cancer cells in vitro using (213)Bi-C595 alpha-immunoconjugate (AIC). The expression of MUC1 on pancreatic tumour tissues and cancer cell lines was performed by immunohistochemistry and further confirmed by confocal microscope and flow cytometry analysis on the cell surface. Cytotoxicity of (213)Bi-C595 was tested by MTS assay. Apoptosis was documented using TUNEL assay. Overexpression of MUC1 was found in approximately 90% of tested tumour samples and the three pancreatic cancer cell lines. (213)Bi-C595 is specifically cytotoxic to pancreatic cancer cells in a concentration-dependent fashion. These results suggest that overexpression of MUC1 in pancreatic cancer is a useful target, and that the novel (213)Bi-C595 AIC selectively targets pancreatic cancer cells in vitro. (213)Bi-C595 may be a useful agent for the treatment of micrometastases or minimal residual disease (MRD) in pancreatic cancer patients with overexpression of MUC1 antigen.

Preclinical studies of targeted alpha therapy for breast cancer using 213Bi-labelled-plasminogen activator inhibitor type 2.

The control of micrometastatic breast cancer remains problematic. To this end, we are developing a new adjuvant therapy based on (213)Bi-PAI2, in which an alpha-emitting nuclide ((213)Bi) is chelated to the plasminogen activator inhibitor-2 (PAI2). PAI2 targets the cell-surface receptor bound urokinase plasminogen activator (uPA), which is involved with the metastatic spread of cancer cells. We have successfully labelled and tested recombinant human PAI2 with the alpha radioisotope (213)Bi to produce (213)Bi-PAI2, which is highly cytotoxic towards breast cancer cell lines. In this study, the 2-day postinoculation model, using MDA-MB-231 breast cancer cells, was shown to be representative of micrometastatic disease. Our in vivo efficacy experiments show that a single local injection of (213)Bi-PAI2 can completely inhibit the growth of tumour at 2 days postcell inoculation, and a single systemic (i.p.) administration at 2 days causes tumour growth inhibition in a dose-dependent manner. The specific role of uPA as the target for (213)Bi-PAI2 therapy was determined by PAI2 pretreatment blocking studies. In vivo toxicity studies in nude mice indicate that up to 100 microCi of (213)Bi-PAI2 is well tolerated. Thus, (213)Bi-PAI2 is successful in targeting isolated breast cancer cells and preangiogenic cell clusters. These results indicate the promising potential of (213)Bi-PAI2 as a novel therapeutic agent for micrometastatic breast cancer.

213Bi-PAI2 conjugate selectively induces apoptosis in PC3 metastatic prostate cancer cell line and shows anti-cancer activity in a xenograft animal model.

A novel alpha-particle emitting ((213)Bi) plasminogen activator inhibitor type 2 construct, which targets the membrane-bound urokinase plasminogen activator on prostate cancer cells, was prepared and evaluated in vitro and in a xenograft animal model. The PC3 prostate cancer cell line expresses urokinase plasminogen activator which binds to its receptor on the cell membrane; plasminogen activator inhibitor type 2 is bound to urokinase plasminogen activator/urokinase plasminogen activator receptor to form stable complexes. In vitro, the cytotoxicity of (213)Bi-plasminogen activator inhibitor type 2 against prostate cancer cells was tested using the MTS assay and apoptosis was documented using terminal deoxynucleotidyl transferase-mediated deoxyuridinetriphosphate nick end-labelling (TUNEL) assay. In vivo, antiproliferative effects for tumours and prostate cancer lymph node metastasis were carried out in an athymic nude mouse model with a subcutaneous xenograft of PC3 cells. (213)Bi-plasminogen activator inhibitor type 2 was specifically cytotoxic to PC3 cells in a concentration-dependent fashion, causing the cells to undergo apoptosis. A single local or i.p. injection of (213)Bi-plasminogen activator inhibitor type 2 was able to completely regress the growth of tumours and lymph node metastases 2 days post subcutaneous inoculation, and obvious tumour regression was achieved in the therapy groups compared with control groups with (213)Bi-plasminogen activator inhibitor type 2 when the tumours measured 30-40 mm(3) and 85-100 mm(3). All control animals and one of five (20%) mice treated with 3 mCi kg(-1) (213)Bi-plasminogen activator inhibitor type 2 developed metastases in the lymph nodes while no lymphatic spread of cancer was found in the 6 mCi kg(-1) treated groups at 2 days and 2 weeks post-cell inoculation. These results demonstrate that this novel (213)Bi-plasminogen activator inhibitor type 2 conjugate selectively targets prostate cancer in vitro and in vivo, and could be considered for further development for the therapy of prostate cancer, especially for the control of micro-metastases or in minimal residual disease.

In vitro and preclinical targeted alpha therapy of human prostate cancer with Bi-213 labeled J591 antibody against the prostate specific membrane antigen.

Limited options for the treatment of prostate cancer have spurred the search for new therapies. One innovative approach is the use of targeted alpha therapy (TAT) to inhibit cancer growth, using an alpha particle emitting radioisotope such as (213)Bi. Because of its short range and high linear energy transfer (LET), alpha-particles may be particularly effective in the treatment of cancer, especially in inhibiting the development of metastatic tumors from micro-metastases. Prostate-specific membrane antigen (PSMA) is expressed in prostate cancer cells and the neovasculature of a wide variety of malignant neoplasms including lung, colon, breast and others, but not in normal vascular endothelium. The expression is further increased in higher-grade cancers, metastatic disease and hormone-refractory prostate cancer (PCA). J591 is one of several monoclonal antibodies (mabs) to the extracellular domain of PSMA. Chelation of J591 mab with (213)Bi forms the alpha-radioimmunoconjugate (AIC). The objective of this preclinical study was to design an injectable AIC to treat human prostate tumors growing subcutaneously in mice. The anti-proliferative effects of AIC against prostate cancer were tested in vitro using the MTS assay and in vivo with the nude mice model. Apoptosis was documented using terminal deoxynucleotidyl transferase [TdT]-mediated deoxyuridinetriphosphate [dUTP] nick end-labeling (TUNEL) assay, while proliferative index was assessed using the Ki-67 marker. We show that a very high density of PSMA is expressed in an androgen-dependent human PCA cell line (LNCaP-LN3) and in tumor xenografts from nude mice. We also demonstrate that the AIC extensively inhibits the growth of LN3 cells in vitro in a concentration-dependent fashion, causing the cells to undergo apoptosis. Our in vivo studies showed that a local AIC injection of 50 microCi at 2 days post-cell inoculation gave complete inhibition of tumor growth, whereas results for a non-specific AIC were similar to those for untreated mice. Further, after 1 and 3 weeks post-tumor appearance, a single (100 microCi/100 microl) intra-lesional injection of AIC can inhibit the growth of LN3 tumor xenografts (volume<100 mm(3)) in nude mice. Tumors treated with AIC decreased in volume from a mean 46+/-14 mm(3) in the first week or 71+/-15 mm(3) in the third week to non-palpable, while in control mice treated with a non-specific AIC using the same dose, tumor volume increased from 42 to 590 mm(3). There were no observed side effects of the treatment. Because of its in vitro cytotoxicity and these anti-proliferative properties in vivo, the (213)Bi-J591 conjugate has considerable potential as a new therapeutic agent for the treatment of prostate cancer.