Folate receptor-targeted immunotherapy: Induction of humoral and cellular immunity against hapten-decorated cancer cells

Abstract

We previously exploited the frequent overexpression of folate receptors on cancer cells to decorate malignant cell surfaces selectively with folate-hapten conjugates. In antihapten-immunized hosts, this targeted localization of foreign haptens to tumor cells led to rapid accumulation of autologous antihapten IgG, which in turn yielded potent antitumor activity upon stimulation with cytokines (IL-2, IFN-a). In an effort to understand the effector mechanisms responsible for tumor regression, we have now investigated the involvement of both humoral and cellular immune components in the tumor destruction process. We report that the dependence of therapeutic efficacy on folate-hapten concentration is bimodal, suggesting that the conjugate must bridge between a cell surface FR and an antihapten IgG in order to mediate killing. Studies with cancer cells in vitro further demonstrate that folatefluorescein-marked tumor cells are killed primarily by antibodydependent cellular cytotoxicity and phagocytosis, with no contribution from complement-dependent mechanisms. Investigations of specific immune cell involvement also reveal that asialo-GM1 1 -natural killer cells, macrophages, CD41 T cells and CD8 1 T cells contribute significantly to recognition/removal of the cancer mass, and that elimination of these cell types markedly compromises the therapy. Because the initial antibody-dependent stage of tumor cell killing is shown to lead to a long-term antibody-independent cellular immunity that involves both CD4 1 and CD8 1 T cells, we propose that F c receptor-expressing immune cells not only initiate destruction of the IgG-marked tumor cells, but also participate in presentation of endogenous tumor antigens in a manner that leads to long-term cellular immunity. ' 2005 Wiley-Liss, Inc.

Preclinical pharmacokinetics, tissue distribution, and antitumor activity of a folate-hapten conjugate–targeted immunotherapy in hapten-immunized mice

“…3C) revealed that although systemically given 3 H-folate-FITC could localize quickly to a folate receptor -positive tumor, the majority of the compound was simultaneously bound to anti-FITC antibody in the plasma. We have previously observed that folate-FITC present in vast excess over anti-FITC antibody may actually avert the ''anchoring'' of anti-FITC antibody to folate receptor -positive tumor (7,8). In an effort to consolidate our findings and to further enhance antitumor activity, immunized mice bearing s.c. M109 tumors (f100 mm 3 ) were given a 3-week regimen (dosing five times per week) consisting of folate-FITC at three dose levels (500, 2,000, and 5,000 nmol/kg), alone or in combination with IL-2 and IFN-a.…”

“…Consequently, folate-FITC can promote antibody-dependent cellular cytotoxicity, phagocytosis, and perhaps T cell -mediated tumor cell killing (8). In the tumor destruction process, however, costimulation with the proinflammatory cytokines IL-2 and IFN-a seemed to be important in promoting a more effective and longlasting antitumor response (8). Because high cytokine doses are often associated with unwanted toxicity, we investigated the effect of using lower doses of IL-2 and IFN-a in support of a fixed dosing regimen of folate-FITC.…”

“…Based on experiments with mouse effector cells in vitro, folate-targeted immunotherapy was suggested to function via multiple tumor-killing mechanisms, including antibody-dependent cellular cytotoxicity and phagocytosis (8). Additional studies of immune effector cell involvement in vivo showed the participation of asialo-GM1 + natural killer cells, macrophages, and CD4 + and CD8 + T cells in tumor regression (8).…”

“…Additional studies of immune effector cell involvement in vivo showed the participation of asialo-GM1 + natural killer cells, macrophages, and CD4 + and CD8 + T cells in tumor regression (8). More importantly, a tumor-protective immunity was established in cured animals, and it was associated with the detection of memory CD4 + and CD8 + T cells in vivo and tumor-specific cytotoxic T-cell activity ex vivo (6,8). Accordingly, the induction of humoral and cellular immune responses may have been complementary to each other in the process of both hapten-dependent and hapten-independent tumor cell destruction.…”

“…Preclinical work has thus far been focused on the therapeutic components that include (a) the fluorescein hapten (FITC), (b) keyhole limpet hemocyanin (KLH) as the carrier protein, (c) GPI-0100 as the adjuvant, (d) folateethylenediamine-FITC (folate-FITC) as the targeted hapten, and (e) the proinflammatory cytokines IL-2 and IFN-a (8). FITC was chosen as the hapten primarily because of its low molecular weight, ready availability, and ease of coupling to KLH or folate (to make folate-FITC).…”