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Michigan Oncology Journal Summer 2001

A Novel Anti-angiogenesis Strategy for the Treatment of Head and Neck Cancer

Theodoros N. Teknos, M.D.,
Assistant Professor
Department of Otorhinolaryngology, Head and Neck Surgery Division

Angiogenesis is the process of new blood vessel formation. It is encountered in essential physiologic processes (e.g., endometrial proliferation, embryogenesis, wound healing), as well as in countless pathologic conditions (e.g., rheumatoid arthritis, diabetic retinopathy, neoplastic disease) (1). Dr. Judah Folkman pioneered the concept that primary or metastatic tumors must become vascularized to exceed 2 mm in size (2). Based on extensive animal data, Folkman et al., found that tumor angiogenesis must precede tumor growth and thus, tumor progression can only occur when neoplasms switch from a prevascular to an angiogenic phase (3). The process of angiogenesis, however, is complex and is mediated by the delicate balance between pro-angiogenic and anti-angiogenic molecules. The activation of the angiogenic "switch" relies on a relative preponderance of pro-angiogenic molecules, such as vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), interleukin-8 (IL-8), platelet derived growth factor (PDGF), angiogenin and hepatocyte growth factor, to name a few (4,5).


Angiogenesis in Head and Neck Cancer

The requirement of angiogenesis for tumor growth is rarely debated; however, the utility in using the level of angiogenesis as a prognostic indicator is point of great debate. Specifically in head and neck squamous cell carcinoma, most studies to date have attempted to correlate intratumoral microvessel density with tumor recurrence and survival. Although not incontrovertible, a preponderance of evidence suggests that increased tumor vascularity is associated with loco-regional recurrence, distant metastases and poor prognosis (6-12). Several studies in our laboratory have shown that advanced laryngeal squamous cell carcinomas with a high level of vascularity tend to behave more aggressively, be more resistant to cytotoxic therapy and have decreased overall survival compared to those with a low level of vascularity (13).

The degree of expression of various pro-angiogenic cytokines has also provided valuable prognostic information on various solid tumors. Overexpression of VEGF, for instance, has been correlated with poor survival rates in esophageal, gastric, colorectal, pancreatic, lung and breast carcinomas (14). Our laboratory recently studied serum VEGF levels in a large number of normal control subjects and patients with advanced laryngeal carcinoma treated in the Veterans Administration Laryngeal Cancer Study Group Protocol #268. In this protocol, patients with stage III or IV laryngeal cancer were randomized to receive either conventional therapy (total laryngectomy with postoperative radiation therapy) or induction chemotherapy followed by radiation therapy. In the latter arm, surgical salvage was performed on those patients who failed to respond to two cycles of chemotherapy or on those with persistent or recurrent disease at the completion of treatment (15). Not surprisingly, the serum levels of VEGF were significantly different between the healthy volunteers vs. the patients with advanced laryngeal carcinoma (47.8pg/ml vs. 317pg/ml, p>0.001). More importantly, however, high serum VEGF levels were predictive of poorer overall survival regardless of the treatment employed (p=0.0018). A multivariate analysis was also performed comparing VEGF levels to all known risk factors for recurrence including: T stage, N stage, tumor growth pattern, site of tumor, treatment used and patient age. This analysis revealed that serum VEGF levels are the best independent predictor of poor survival and patients with levels above the mean had a 47% increased risk of death (p=0.065).


Copper Suppression as an Anti-angiogenic Approach

Based on all the available data, the level of angiogenesis may play a critical role in the development and progression of human squamous cell carcinoma. Utilization of an anti-angiogenic compound, therefore, may prove very beneficial to patients suffering from this relentless form of cancer. To date, there have been no reports in the literature of a single agent, anti-angiogenic compound effective against head and neck squamous cell carcinoma. There is, however, a growing list of angiogenesis inhibitors under investigation in a variety of tumor types. These include angiostatin, endostatin, thalidomide, linomide, TNP-470, SU1444, SU1498 and monoclonal antibodies against fibroblast growth factor, VEGF and avB3 integrin, to name a few. Due to the numerous steps involved in angiogenesis, any disruption of this process will require innovative strategies that inhibit multiple steps in the angiogenesis pathway. Copper suppression therapy may prove to be one such strategy.

Copper plays a key role in multiple steps along the angiogenesis pathway. It is an essential co-factor for such molecules as bFGF, VEGF and angiogenin (16-18). In the absence of copper, these cytokines cannot function and neovascularization is abated. A number of intracerebral tumor models have utilized this strategy, employing penicillamine and a low copper diet to successfully reduce tumor growth (19, 20). A low-copper diet is not possible in humans due to ubiquitous nature of copper in food. Tetrathiomolybdate (TM), however, is a powerful copper chelator that is well tolerated when taken orally. Dr. George Brewer developed TM at the University of Michigan for the treatment of patients with Wilson's disease, an autosomal recessive disease of copper transport that results in abnormal copper accumulation and toxicity. TM's ability to reduce copper stores involves at least two mechanisms: the first is the formation of a complex together with food proteins in the gastrointestinal tract, blocking the absorption of copper (21). The second is that the absorbed TM forma a tripartite complex with copper and albumin in blood, rendering the copper unavailable for cellular uptake and therefore removing it from use in angiogenesis. This tripartite complex has no biologic activity and is excreted via the biliary tract and urine (21). The low toxicity profile of TM is possible because the levels of copper required for angiogenesis is higher than that required for essential copper dependent processes, such as heme synthesis, superoxide dismutase and cytochrome function (21).

Merajver and Brewer et al., showed dramatic efficacy of TM as an anti-angiogenic compound for the treatment of inflammatory breast cancer in a HER2/Neu transgenic mouse model (22). In our studies, we found that mice treated with TM after the establishment of a very aggressive murine squamous cell carcinoma had a dramatic decrease in tumor growth rate and significant reduction in tumor vascularity (Fig. 1) (23). Clearly, TM was effective after tumors had been established and the angiogenic "switch" had been thrown. To investigate its use as a chemopreventative agent, we initially made mice copper deficient compared to controls then implanted tumor cells. As Figure 2 shows, tumors remained very small in size compared to controls up to day 55. When TM was removed from the mice's drinking water, the tumors rapidly enlarged, approaching the growth rate of the untreated subjects (24). Ongoing studies have shown similar effectiveness in human tumors established in immunodeficient SCID mice (Figure 3) (25). Based on these preclinical findings, clearly there is ample evidence to support the use of TM in humans with squamous cell carcinoma of the head and neck. In collaboration with Drs. Francis Worden, Susan Urba, George Brewer, Sofia Merajver and others, a phase II clinical study using TM in patients with metastatic unresectable squamous cell carcinoma is presently being organized at the Univer-sity of Michigan Comprehensive Cancer Center.



Squamous cell carcinoma of the head and neck continues to be a difficult disease process to treat. Despite dramatic advances in surgical and non-surgical treatment options, the cure rates remain unchanged over the past 30 years. Angiogenesis inhibitors are a novel and very encouraging treatment approach to these tumors. Continued investigations and the collaborative effort of all physicians caring for head and neck cancer patients will be necessary to determine if these approaches will ultimately improve survival.



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2.Folkman J. What is the evidence that tumors are angiogenesis dependent? J Natl Cancer Inst. 1990;82:4-6.

3. Hanahan D, Folkman J. Patterns and emerging mechanisms of the angiogenic switch during tumorigenesis. Cell. 1996;86:353-64.

4.Folkman J. Tumor Angiogenesis. In: Mendelsohn J, Howley PM, Israel MA, Liotta LA, eds. The Molecular Basis of Cancer. Philadelphia: W.B. Saunders; 1995:206-32.

5.Folkman J, Klagsburn M. Angiogenic factors (review). Science. 1987;235;442-7.

6.Albo D, Granick MS, Jhala N, Atkinson B, Solomon MP. The relationship of angiogenesis to biological activity in human squamous cell carcinomas of the head and neck. Ann of Plast Surg. 1994;32(6):588-94.

7.Gasparini G, Weidner N, Maluta S, et al. Intratumoral microvessel density and p53 protein: correlation with metastasis in head-and-neck squamous-cell carcinoma. Int Journal Cancer. 1993;55(5):739-44.

8.Murray JD, Carlson GW, McLaughlin K, et al. Tumor angiogenesis as a prognostic factor in laryngeal cancer. Am J Surg. 1997;174(5):523-6.

9.Pazouki S, Chisholm DM, Adi MM, et al. The association between tumour progression and vascularity in the oral mucosa. J Pathol. 1997;183(1):39-43.

10.Eisma RJ, Spiro JD, Kreutzer DL. Vascular endothelial growth factor expression in head and neck squamous cell carcinoma. Am J Surg. 1997;174(5):513-7.

11.Maeda T, Matsumura S, Hiranuma H, Jikko A, Furukawa S, Ishida T, Fuchihata H. Expression of vascular endothelial growth factor in human oral squamous cell carcinoma: Its association with tumour progression and p53 gene status. J Clin Pathol. 1998;51(10):771-5.

12.Moriyama M, Kumagai S, Kawashiri S, Kojima K, Kakihara K, Yamamoto E. Immunohistochemical study of tumour angiogenesis in oral squamous cell carcinoma. Oral Oncology. 1997;33(5):369-74.

13.Teknos TN, Cox C-, Barrios MA, et al. Tumor angiogenesis as a predictive marker for organ preservation in patients with advanced laryngeal carcinoma. The Laryngoscope. In press.

14.Tae K, El-Naggar AK, Yoo E, et al. Expression of vascular endothelial growth factor and microvessel density in head and neck tumorigenesis. Clin Cancer Res. 2000;6:2821-8.

15.Teknos TN, Cox CC, Yoo SK, Wolf GT, Fischer SG. Serum vascular endothelial growth factor correlates with outcome in the V.A. laryngeal cooperative study Group Protocol #268. (Unpublished data)

16.Watanabe T, Seno M, Sasada R, Igarashi K. Molecular characterization of recombinant human acidic fibroblast growth factor produced by E. coli: comparative studies with human basic fibroblast growth factor. Mol Endocrinol. 1990;4:869-79.

17.Patstone G, Maher P. Copper and calcium binding motifs in the extracellular domains of fibroblast growth factor receptors. J Biol Chem. 1996;271:3343-6.

18.Ziche M, Jones J, Gullino PM. Role of prostaglandin E and copper in angiogenesis. J Natl Cancer Inst. 1982;69:475-82.

19.Brem SS, Zagzag D, Tsanaclis AMC, Gately S. Inhibition of angiogenesis and tumor growth in the brain. Am J Pathol. 1990;137:1121-42.

20. Brem S, Tsanaclis AMC, Zagzag D. Anticopper treatment inhibits pseudopodial protrusion and the invasive spread of 9L gliosarcoma cells in the rat brain. Neurosurgery. 1990;26:391-6.

21.Brewer GJ, Dick RD, Yuzbasiyan-Gurkin V, Tankanow R. Initial therapy of patients with Wilson's disease with tetrathiomolybdate. Arch Neurology. 1991;48:42-7.

22.Merajver SD, Irani J, van Golen K, Brewer GJ. Copper depletion as an anti-angiogenic strategy in HER2-neu transgenic mice. Proc AACR Special Conference on Angiogenesis and Cancer Research. 1998, B11.

23.Cox C, Teknos TN, Barrios M, Brewer GJ, Dick RD, Merajver SD. The role of copper suppression as an anti-angiogenic strategy in head and neck squamous cell carcinoma. The Laryngoscope. In press.

24.Cox CC, Teknos TN, Merajver SD, Brewer GJ. Tetrathiomolybdate as a chemopreventative agent in squamous cell carcinoma. (Unpublished data.)

25.Teknos TN, Cox CC, Gupta A, et al. Copper suppression in the treatment of human squamous cell carcinoma. (Unpublished data.)


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Please note: The articles listed in the Cancer Center's News Archive are here for historical purposes. The information and links may no longer be up-to-date.
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