EDITORIAL

Cytotoxins and Cancer Immunotherapy: The Dance of the Macabre?

Carmen J. Allegra, Richard W. Childs

Correspondence to: Carmen J. Allegra, MD, Network for Medical Communication and Research, North Potomac, MD 20878 (e-mail: callegra{at}nmcr.com) or Richard W. Childs, MD, Hematology Branch, National Heart, Lung, and Blood Institute, NIH Bldg. 10/CRC, Rm. 3-5140, 10 Center Dr., MSC 1202, Bethesda MD 20892 (e-mail: childsr{at}nih.gov).

The combined mortality rate for patients with breast or colorectal cancer in the United States approaches 100 000 individuals annually or approximately 270 individuals every day (1). This astounding mortality rate needs to be rapidly and positively addressed through multiple complementary efforts that must include the development of more specific, more effective, and less toxic therapeutic strategies. The present management of patients with advanced breast and colorectal cancers involves the use of broadly cytotoxic agents applied either sequentially or in various combinations and, generally, in concert with biologic agents. Although these strategies have met with laudable success, they unfortunately do not result in curing patients with disease that has spread beyond the practical limits of surgical intervention. Thus, there is an urgent need to develop more effective and better tolerated systemic therapeutic options.

In this issue of the Journal, Correale et al. describe the use of a 28-amino-acid peptide (TS/PP) that contains the sequence of three HLA-A02.01–restricted peptides derived from the enzyme thymidylate synthase (TS) to vaccinate mice against cancers that overexpress this enzyme relative to its expression in normal tissues (2). In vitro, TS/PP peptide-pulsed dendritic cells induced potent cytotoxic T-lymphocyte (CTL) antitumor activity against multiple TS-derived peptides that was enhanced by pretreating tumor cells with 5-fluorouracil (5-FU). More important, vaccination with this polypeptide in combination with relatively noncytotoxic doses of 5-FU induced a potent CTL response that inhibited human colon and breast cancers in vitro and cured or delayed the growth of a human lymphoma tumor cells inoculated into HLA-A02.01 transgenic mice. Of interest, vaccination and exposure to relatively noncytotoxic doses of 5-FU resulted in no discernable immune-mediated toxic effects on normal murine tissues by histologic examination—a remarkable result if translatable into the management of human cancer.

What is TS and why was this particular enzyme selected as a potential target for vaccination? TS is expressed in virtually every normal and malignant cell and is essential for maintenance of cell viability in the absence of a supraphysiologic source of thymidine. TS is an essential step in the de novo pyrimidine synthetic pathway and responsible for the enzymatic methylation of deoxyuridylate to form thymidylate needed for DNA repair and replication. TS has been shown by multiple laboratories to be overexpressed in tumor versus normal tissues, and its cellular levels are tightly controlled by an autoregulatory translational feedback inhibition resulting from the binding of the TS protein to its own mRNA—an interaction that is exquisitely dependent on the state of occupancy of the enzyme by its substrates and/or inhibitors (3,4). With exposure to 5-FU, the enzyme becomes tightly bound in a ternary complex with the fluoropyrimidine anabolite, 5-fluorodeoxyuridylate, and methylene tetrahydrofolate, resulting in loss of its ability to bind to and negatively regulate the translation of its own mRNA. This loss of translational repression results in increased protein levels of TS. It is this induction of TS protein along with its baseline relative overexpression in cancerous cells that provided the rationale for selecting this enzyme as a vaccine target. Overexpression of TS has also been shown to be associated with a poor prognosis in patients with colon and breast cancers and with relative insensitivity to 5-FU in patients with advanced disease (5,6). Recently, it has been demonstrated that TS overexpression is oncogenic (7). Each of these features supports TS as a potential target for vaccination.

Although in vitro and in vivo murine tumor models have provided us with tremendous insight into mechanisms to enhance antitumor immunity, relatively few humans have benefited from conventional peptide-based cancer immunotherapy. Bulky tumors outgrowing or suppressing the immune response, loss of the target antigen that is not essential for tumor survival, and poor host immunity as a consequence of immunosuppressive cytotoxic chemo/radiotherapy are all factors thought to play a role in the failure of peptide-based vaccination approaches. The ability to generate tumor antigen–specific immune responses in patients who ultimately do not achieve a disease response has led investigators to explore methods to sensitize the tumor to immune attack. Recently, radiotherapy and several cytotoxic agents have been shown to increase Fas and the expression of tumor-associated antigens such as carcinoembryonic antigen (CEA) (8). Although Correale et al. show that 5-FU sensitizes the tumor to killing by TS-specific CTL, definitive data showing that CTL killing is enhanced as a direct consequence of increased tumor surface expression of TS-derived peptide antigens are not presented in this study. An alternative explanation that needs to be considered is that the enhanced tumor susceptibility to T-cell attack is the result of another of the many cellular effects associated with exposure to 5-FU. Several laboratories have previously shown that 5-FU–exposed tumors increase the expression a variety of molecules including tumor necrosis factor–related apoptosis-inducing ligand (TRAIL), Fas, and p53—molecules that might sensitize the tumor (and possibly nonmalignant cells) more globally to other antigen-specific CTL responses (911). A more comprehensive understanding of the mechanism of interaction may permit the use of more tumor-selective antigens and provide a broader menu of cytotoxic agents associated with increasing the expression of the critical molecule(s). Although alternative mechanisms that may underpin the noted synergy need further investigation, this fact does not detract from the importance of the present study.

However, the vaccine approach described by Correale et al. does raise several concerns related to potential barriers to clinical translation that will require future exploration. To optimize efficacy while avoiding autoimmunity, investigational cancer vaccine regimens have focused largely on antigens that are both overexpressed and restricted to the tumor. Although TS is highly expressed in cancer cells, it is also expressed in virtually all normal cells and may be highly induced in normal tissues with exposure to 5-FU or other inhibitors of TS such as the antifolate inhibitor, pemetrexed (Alimta), currently approved in the United States for the management of patients with malignant pleural mesothelioma and non–small-cell lung cancer (12,13). These facts provide a cautionary note to the potential for safely translating this particular vaccine target into patients despite the apparent lack of autoimmunity noted in the murine model. An additional concern is that the mice were vaccinated prior to the infusion of malignant cells and treatment with 5-FU. The ability of tumors to suppress cancer immunity and the immunosuppressive effects of chemotherapy prevent one from extrapolating whether this vaccination approach would have resulted in the same protective effect in tumor-bearing mice treated with 5-FU prior to or simultaneously with vaccination. A lack of efficacy under these latter conditions would relegate the approach to a prevention-only strategy, where the potential risk of autoimmunity may be more problematic given the global expression of TS in normal tissues. A recent report by this same group provides evidence that T-cell immunity against CEA- and TS-derived peptides in humans with metastatic colon cancer persists when combination chemotherapy is followed by treatment with granulocyte–macrophage colony-stimulating factor and interleukin 2 (14). Clearly, additional preclinical data are needed to address whether tumor immunity would still be enhanced if this polypeptide-based vaccination approach was used concomitant with chemotherapy in animals with established tumors.

In summary, the findings of Correale et al. broadly imply that 5-FU and other cytotoxic agents have the additional benefit of synergizing with tumor immunity by sensitizing the tumor to the effects of tumor antigen–specific CTL. On the basis of these data, it would seem reasonable to investigate the efficacy of postchemotherapeutic vaccination or the adoptive infusion of in vitro expanded antigen-specific CTL. Careful exploration of the logistics of drug dosing and drug timing relative to peptide vaccination or infusion of CTL with such sequential chemoimmunotherapy regimens is needed because they will likely have a critical impact on the efficacy of such approaches. Finally, although the induction of TS may be critical to the efficacy of the described approach, further exploration of alternative mechanisms associated with exposure to 5-FU, as well as the wisdom of targeting a protein expressed in both normal and malignant cells, is needed.

REFERENCES

(1) Ries LA, Eisner MP, Kosary CL, Hankey BF, Miller BA, Clegg L, et al. (eds). SEER Cancer Statistics Review, 1975-2002, National Cancer Institute. Bethesda (MD). Available at: http://seer.cancer.gov/csr/1975_2002, based on November 2004 SEER data submission, posted to the SEER Web site 2005.

(2) Correale P, Del Vecchio MT, Di Genova G, Savellini GG, La Placa M, Terrosi C, et al. 5-Fluorouracil–based chemotherapy enhances the antitumor activity of a thymidylate synthase-directed polyepitopic peptide vaccine. J Natl Cancer Inst 2005;97:1437–45.[Abstract/Free Full Text]

(3) Johnston PG, Liang CM, Henry S, Chabner BA, Allegra CJ. The production and characterization of monoclonal antibodies that localize human thymidylate synthase in the cytoplasm of human cells and tissue. Cancer Res 1991;51:6668–76.[Abstract]

(4) Chu E, Koeller DM, Casey JL, Drake JC, Chabner BA, Elwood PC, et al. Autoregulation of human thymidylate synthase messenger RNA translation by thymidylate synthase. Proc Natl Acad Sci U S A 1991;88:8977–81.[Abstract/Free Full Text]

(5) Allegra C, Paik S, Colangelo L, Parr A, Johnston P, Wolmark N, et al. Prognostic value of thymidylate synthase, Ki-67, and p53 in patients with Dukes' B and C colon cancer: a National Cancer Institute-National Surgical Adjuvant Breast and Bowel Project collaborative study. J Clin Oncol 2003;21:241–50.[Abstract/Free Full Text]

(6) Johnston PG, Lenz HJ, Leichman CG, Danenberg KD, Allegra CJ, Danenberg PV, Leichman L. Thymidylate synthase gene and protein expression correlate and are associated with response to 5-fluorouracil in human colorectal and gastric tumors. Cancer Res 1995;55:1407–12.[Abstract]

(7) Rahman L, Voeller D, Rahman M, Lipkowitz S, Allegra C, Barrett K, et al. Thymidylate synthase as an oncogene: a novel role for an essential DNA synthesis enzyme. Cancer Cell 2004;5:341–51.[CrossRef][ISI][Medline]

(8) Chakraborty M, Abrams S, Coleman CN, Camphausen K, Schlom J, Hodge JW. External beam radiation of tumors alters phenotype of tumor cells to render them susceptible to vaccine-mediated T-cell killing. Cancer Res 2004;64:4328–37.[Abstract/Free Full Text]

(9) Yamamoto T, Nagano H, Sakon M, Wada H, Eguchi H, Kondo M, et al. Partial contribution of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)/TRAIL receptor pathway to antitumor effects of interferon-alpha/5-fluorouracil against hepatocellular carcinoma. Clin Cancer Res 2004;10:7884–95.[Abstract/Free Full Text]

(10) Longely DB, Allen WL, McDermott U, Wilson TR, Latif T, Boyer J, et al. The roles of thymidylate synthase and p53 in regulating Fas-mediated apoptosis in response to antimetabolites. Clin Cancer Res 2004;10:3562–71.[Abstract/Free Full Text]

(11) Ju J, Petersen-Lane J, Maley F, Chu E. Regulation of p53 expression by thymidylate synthase. Proc Natl Acad Sci U S A 1999;96:3769–74.[Abstract/Free Full Text]

(12) Keyomarsi K, Samet J, Molnar G et al: The thymidylate synthase inhibitor, ICI D1694, overcomes translational detainment of the enzyme. J Biol Chem 1993;268:15142–9.[Abstract/Free Full Text]

(13) Welsh SJ, Titley J, Brunton L, Valenti M, Monaghan P, Jackman AL, et al. Comparison of thymidylate synthase (TS) protein up-regulation after exposure to TS inhibitors in normal and tumor cell lines and tissues. Clin Cancer Res 2000;6:2538–46.[Abstract/Free Full Text]

(14) Correale P, Cusi MG, Tsang KY, Del Vecchio MT, Marsili S, La Placa M, et al. Chemo-immunotherapy of metastatic colorectal carcinoma with gemcitabine plus FOLFOX 4 followed by subcutaneous granulocyte macrophage colony-stimulating factor and interleukin-2 induces strong immunologic and antitumor activity in metastatic colon cancer patients. J Clin Oncol 2005 Aug 1; [Epub ahead of print].



             
Copyright © 2005 Oxford University Press (unless otherwise stated)
Oxford University Press Privacy Policy and Legal Statement