Cancer Cell Lines SW480 Research Articles

Preparation and properties of the immunoconjugate composed of anti-human colon cancer monoclonal antibody and mitomycin C-dextran conjugate.

Monoclonal antibody (mAb) A7, produced against human colon cancer, was conjugated with a polymeric prodrug of mitomycin C (MMC), the MMC-dextran conjugate with an anionic charge (MMCDan) and a molecular weight of 70,000. The amino groups were introduced into the MMCDan by reacting ethylenediamine with the carboxyl group in the spacer arm of the dextran bridge by a carbodiimide-catalyzed reaction. The coupling to mAb A7 was performed using SPDP. A 15 M excess of ethylenediamine produced an optimal MMCDan with amino groups, which resulted in a homogenous conjugate (A7-MMCD) with minimal formation of high-molecular-weight aggregates in about a 30% yield of both IgG and MMC. The molar binding ratio of IgG:dextran:MMC in A7-MMCD was estimated to be 1:1.2:40. A7-MMCD, having MMC prodrug properties, released active MMC with a half-life of 29.1 h and had an almost neutral electric charge under physiological conditions. A competitive binding assay using 125I-labeled A7 revealed that the A7-MMCD almost fully retained its antibody-binding activity. The cytotoxicity of A7-MMCD was assayed by determining the degree of inhibition of [3H]-thymidine in corporation in two different ways using the human colon cancer cell line SW1116. A 48-h continuous exposure test revealed that the pharmacological activity of MMC in A7-MMCD was completely preserved. In addition, A7-MMCD exhibited about a 14-fold greater cytotoxicity than MMCDan when the IC50 values determined using a 2-h pretreatment exposure system were compared. These results suggest that A7-MMCD could be useful in immunotargeting chemotherapy for colorectal cancer.

Cytokine effects and role of adhesive proteins and Fc receptors in human macrophage-mediated antibody dependent cellular cytotoxicity

Mononuclear phagocytes participate in host immunological defense against tumors. We have investigated the role of selected recombinant cytokines on human macrophage-mediated tumor cytotoxicity in vitro utilizing a human colon cancer cell line target, SW1116, and murine monoclonal antibody 17-1A. Blood monocytes were kept in continuous culture to allow differentiation into macrophages. Maximum antibody dependent cellular cytotoxicity (ADCC) as measured in a 3H-thymidine release assay occurred after culturing the monocytes for 5-7 days. Human recombinant macrophage colony stimulating factor (CSF) (1,000 U/ml) did not increase ADCC above control levels whereas recombinant human granulocyte-macrophage colony stimulating factor, interleukin 4, and interleukin 3 were all capable of increasing ADCC. Antibodies to the CD11/CD18 integrin receptors did not significantly inhibit ADCC. When the ADCC incubation occurred in the presence of antibodies to the human Fc receptors, ADCC was inhibited significantly only by anti-FcRIII (3G8). A role for tumor necrosis factor alpha or other soluble mediators of cytotoxicity was not demonstrable in this system. These studies suggest avenues for manipulation and augmentation of macrophage-mediated antitumor ADCC.

Alterations in Monoclonal Antibody Affinity and Antigenic Receptor Site Expression on Mycoplasma-Infected Human Colorectal Cancer Cells

The affinity of MoAb CO 17-1A and expression of its antigenic target were studied on uninfected and mycoplasma-infected colorectal cancer cell lines SW 1116 and SW 948. Binding of 125I-labeled CO 17-1A to SW 1116 cells was quantified at 37 degrees C by determination of the affinity constant (Ka) and the number of antigenic receptor sites (r) per cell using Scatchard plots. When mycoplasma-free SW 1116 cells were used as targets, Ka was 0.92 +/- 0.06 x 10(8) M-1 and r = 1.32 +/- 0.14 x 10(6) at 37 degrees C. One batch of unspeciated, mycoplasma-infected SW 116 cells had reduced affinity and a decreased number of antigenic receptor sites per cell for 125I-labeled 17-1A, while another batch of infected SW 1116 cells had a 4- to 5-fold increase in r and diminished Ka for the antibody compared with uninfected cells. When unspeciated, mycoplasma-infected SW 948 cells were exposed to 125I-labeled 17-1A and the data subjected to Scatchard analysis, the affinity of the antibody deviated markedly from linearity and rendered analysis for Ka and r meaningless. These data indicate that mycoplasma infection can produce variable effects on the cellular expression of antigenic receptor sites and the affinity of antibody for its target, and emphasize the importance of using mycoplasma-free cell lines in studies of these parameters.

Influence of Cocktails of Labeled Monoclonal Antibodies on the Localization of Antibodies in Human Tumor Xenografts

In order to evaluate the usefulness of cocktails of labeled monoclonal antibodies (MoAbs) recognizing different antigen molecules to localize human cancer xenografts, we have compared the potential of three MoAbs recognizing representative cancer‐associated CA 19–9, 17–1A and CEA antigens when administered alone or in combination. Specific binding of radioiodinated F(ab′)2 fragments of these three MoAbs was observed to human colorectal cancer cell lines SW1116, LS180 and Co‐3. The percentage of in vitro cell binding of a cocktail of any two MoAbs to cancer cells was equal to the average of those obtained with the two MoAbs alone. The three MoAbs were preferentially localized in tumor tissues xenografted in nude mice. When cocktails of any two MoAbs were used, the obtained tumor‐to‐normal tissue ratios and percent of injected dose per gram of tumor were between the levels obtained for each MoAb when administered alone, in all three tumors transplanted in nude mice. These data suggest that, although cocktails of labeled MoAbs recognizing different antigens may extend the spectrum of tumor specificities, their use does not improve the tumor localization ability of MoAb‐conjugates.