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Dear Sir,

I read with interest the report of Balza et al.1 describing the expression of endoglin (EDG) in human tissues and cultured cell lines using a new monoclonal antibody termed A11. Although the report contains interesting data, I wish to address and clarify obvious deficiencies and questionable conclusions. These problems are important because they are concerned with the potential clinical application of anti-EDG monoclonal antibodies (MAbs) for antiangiogenic therapy of patients with various angiogenesis-associated diseases including cancer.

First, one needs to include an isotype-matched control IgG (or MAb) as a negative control when a MAb is used in various immunoassays. However, the authors used cells treated with fresh medium as a negative control in their immunoassay. Such a control is inadequate. The cells should have been treated with medium containing an isotype-matched control murine IgG (i.e., IgG1). The authors did not report use of an isotype-matched control IgG in any of their immunoassays. Use of such a proper control is particularly important when a weak reactivity of a MAb with tissues and cells is tested. It is commonly observed that IgGs bind to tissues and cells via Fc receptors and other interactions. Therefore, some of the reactivities of A11 with normal tissues and cell lines could be background reactivities or weaker reactions than they observed.

Second, reactivity of an anti-EDG MAb with cultured cell lines does not represent reactivity of the MAb with corresponding fresh tissues, although the authors used such reactivity of A11 as evidence for the lack of specificity of anti-EDG MAbs in general. Anti-EDG MAbs reacted with vascular endothelium but not with tumor cells per se of solid tumor tissues in our tests of several anti-EDG MAbs with more than 100 malignant tissue samples.2, 3 However, these and other anti-EDG MAbs reacted with many malignant epithelial cell lines.4, 5 Therefore, reactivity of anti-EDG MAbs with epithelial cell lines is not relevant to the reactivity of the MAbs in vivo. In contrast, the reactivity of anti-EDG MAbs with hematologic cell lines is clinically relevant because the expression profile of EDG is consistent between the cultured cell lines and fresh leukemia/lymphoma cells of the corresponding phenotypes, e.g., between immature B-lineage leukemia cells and cell lines.6

Third, there are substantial differences among different anti-EDG MAbs in their specificity for tumor vasculature compared with normal tissue vasculature. In our experience, certain anti-EDG MAbs defining different epitopes show differences in fine specificity.7 Balza et al.1 studied reactivity of a single anti-EDG MAb A11 and emphasized the lack of specificity of EDG expression for tumor blood vessels. Despite this assertion, the reactivity of A11 with tumor vasculature was stronger than that with the blood vessels of normal tissues.1 They stated that clinical use of EDG as a target marker should be reevaluated. However, their report does not present new data that may lead us to reevaluate our approach to potential clinical application of our anti-EDG MAbs. In addition, the previously reported reactivity of anti-EDG MAbs with several minor cells4 may not preclude selected anti-EDG MAbs from therapeutic application. We are aware of the lack of strict specificity of anti-EDG MAbs for tumor vasculature. We recently7 stated that EDG is not a tumor-specific marker and that it is expressed in varying degrees in the vasculature of normal tissues. Despite this limitation, we could effectively target tumor-associated vasculature using selected anti-EDG MAbs and immunoconjugates.2, 3, 7, 8 We believe that this effective targeting of tumor-associated vascular endothelium is attributable to the combined effect of the following: 1. Certain anti-EDG MAbs show a highly restricted reactivity among different tissues and cells. 2. EDG is a proliferation-associated antigen on endothelial cells and its expression is upregulated in tumor-associated vascular endothelium.2, 7, 9–12 3. Turnover of endothelial cells of normal adult tissue vasculature is very slow (e.g., more than 1,000 days), whereas these endothelial cells undergo rapid proliferation during spurts of angiogenesis in tumors.13, 14 Therefore, the rapidly dividing endothelial cells of tumor vasculature are much more susceptible to killing by anti-EDG MAbs and immunoconjugates than the quiescent vascular endothelium of normal tissues. Other, undefined, factors may be involved as well.

I would like to point out that most, if not all, of the MAbs that are being successfully applied for cancer therapy in patients are not tumor-specific MAbs. These MAbs include rituximab (anti-CD20 MAb), trastuzumab (anti-HER2 MAb) and C225 (anti-epidermal growth factor receptor MAb). In the therapeutic application of a MAb, we need to find an appropriate therapeutic window in which the MAb can achieve therapeutic efficacy without exerting severe side effects in the host. We were able to find such therapeutic windows for our selected anti-EDG MAbs in animal studies.2, 3, 7, 8 We need to find such windows in patients for the clinical application of these MAbs.

Yours sincerely,

Ben K. SEON

REFERENCES

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  • 1
    Balza E, Castellani P, Zijlstra A, Neri D, Zardi L, Siri A. Lack of specificity of endoglin expression for tumor blood vessels. Int J Cancer 2001;94: 57985.
  • 2
    Seon BK, Matsuno F, Haruta Y, Kondo M, Barcos M. Long-lasting complete inhibition of human solid tumors in SCID mice by targeting endothelial cells of tumor vasculature with antihuman endoglin immunotoxin. Clin Cancer Res 1997;3: 103144.
  • 3
    Matsuno F, Haruta Y, Kondo M, Tsai H, Barcos M, Seon BK. Induction of lasting complete regression of preformed distinct solid tumors by targeting the tumor vasculature using two new anti-endoglin monoclonal antibodies. Clin Cancer Res 1999;5: 37182.
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    Seon BK, Kumar S. CD105 antibody for targeting of tumor vascular endothelial cells. In: FanT-PD, KohnEC, eds. The new angiotherapy. Totowa, NJ: Humana Press, 2001. 499515.
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    Letamendia A, Lastres P, Almendro N, Raab U, Buhring HJ, Kumar S, Bernabeu C. Endoglin, a component of the TGF-beta receptor system, is a differentiation marker of human choriocarcinoma cells. Int J Cancer 1998;76: 5416
  • 6
    Haruta Y, Seon BK. Distinct human leukemia-associated cell surface glycoprotein GP160 defined by monoclonal antibody SN6. Proc Natl Acad Sci USA 1986;83: 7898902.
  • 7
    Takahashi N, Haba A, Matsuno F, Seon BK. Antiangiogenic therapy of established tumors in human skin/severe combined immunodeficiency mouse chimeras by anti-endoglin (CD105) monoclonal antibodies, and synergy between anti-endoglin antibody and cyclophosphamide. Cancer Res 2001;61: 784654.
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    Tabata M, Kondo M, Haruta Y, Seon BK. Antiangiogenic radioimmunotherapy of human solid tumors in SCID mice using (125)I-labeled anti-endoglin monoclonal antibodies. Int J Cancer 1999;82: 73742.
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    Wang JM, Kumar S, Pye D, van Agthoven AJ, Krupinski J, Hunter RD. A monoclonal antibody detects heterogeneity in vascular endothelium of tumours and normal tissues. Int J Cancer 1993;54: 36370.
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    Burrows FJ, Derbyshire EJ, Tazzari PL, Amlot P, Gazdar AF, King SW, Letarte M, Vitetta ES, Thorpe PE. Up-regulation of endoglin on vascular endothelial cells in human solid tumors: implications for diagnosis and therapy. Clin Cancer Res 1995;1: 162334.
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    Miller DW, Graulich W, Karges B, Stahl S, Ernst M, Ramaswamy A, Sedlacek HH, Muller R, Adamkiewicz J. Elevated expression of endoglin, a component of the TGF-beta-receptor complex, correlates with proliferation of tumor endothelial cells. Int J Cancer 1999;81: 56872.
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    Folkman J. Antiangiogenesis agents. In: DeVitaVT, HellmanS, RosenbergSA, eds. Cancer: principles & practice of oncology. Philadelphia: Lippincott, Williams & Wilkins, 2001. 50919.
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    Denekamp J. 1990. Vascular attack as a therapeutic strategy for cancer. Cancer Metastasis Rev 1990;9: 26782.