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On the basis of histologic evaluation of many vaccine-associated sarcomas and immunohistochemical analysis of a few, the author has developed the following working hypothesis: persistent inflammation or wound healing, or both, associated with vaccine components leads to proliferation of fibroblasts and myofibroblasts that undergo neoplastic transformation through unknown mechanisms. Probable "unknown mechanisms" that are based on examples in other species, would include feline sarcoma virus (FeSV), oncogenes, and growth factors. At present, it appears that FeLV (and therefore FeSV) does not have a role; this statement is based on the results of 2 epidemiologic retrospective studies and the study by Ellis et al in which FeLV was not detected in vaccine-associated sarcomas by use of immunohistochemical analysis and polymerase chain reaction (PCR). The author's laboratory has focused on growth factors as likely players. Growth factors are essential for regulation of the cellular events involved in granulation tissue formation and wound healing. When growth factors are infused into cultures of fibroblasts, the physical attributes of the cells change to those of neoplastic cells. Many tumors in human beings have been associated with autocrine
stimulation (ie, the tumors have receptors for growth factors, which they themselves produce in an abnormal/autonomous manner). Finally, many oncogenes cause cancer by coding for and causing over expression of growth factors or their receptors (v-sis codes for platelet-derived growth factor (PDGF); v-erb codes for epidermal growth factor receptor).
Preliminary .studies at the author's laboratory involved immunohistochemical identification and localization of growth factors and their receptors in feline vaccine-associated lesions. It was found that vaccine-associated sarcomas had a mild to strong positive reaction for PDGF and its receptor, whereas nonvaccine-associated fibrosarcomas (NVFSA) had a negative or faint positive reaction. Interestingly, lymphocytes in vaccine-associated sarcomas were positive for PDGF, but lymphocytes in NVFSA and in normal lymph nodes or aggregated lymphoid follicles (Peyer's patches) had negative reaction. Macrophages in the area stained positive for PDGF receptor. The neoplastic cells in vaccine associated sarcomas that were closest to the lymphocytes had strongest staining for PDGF receptor. These findings led to the hypothesis that lymphocytes in vaccine-associated lesions may secrete PDGF to recruit macrophages and cause fibroblast proliferation.
Expression of c-jun was examined. This gene is a protooncogene that codes for a transcriptional protein AP-1 that has been proved to have a role in cellular proliferation and oncogenesis in vitro. The homologue of c-jun, v-jun, has been documented to be oncogenic in avian and murine postwounding sarcoma models. Vaccine-associated sarcomas were strongly positive for c-jun, whereas NVFSA were negative. Increased growth factor stimulation by lymphocytes and via autocrine stimulation may lead to induction and overexpression of c-jun in these tumors.
The author's current research, funded by the VAFSTF, is a continuation of the aforementioned investigations, but at a molecular level. An attempt will be made to confirm the overexpression of c-sis (which encodes for PDGF) and c-jun in the lymphocytes and tumor cells by use of molecular analysis. If there is overexpression, determination will be made as to whether it is attributable to structural gene alterations, gene amplification, or retroviral insertion. Also, the lymphocytes in vaccine-associated lesions will be defined as to lineage (T or B) and subset (CD4 vs CD8, and TH1 vs TH2). The first step, which has been taken, is the identification of T and B cells by use of immunohistochemistry. In preliminary studies, staining of 3 vaccine-associated sarcomas and 3 NVFSA for CD3, a T-cell marker, indicated no differences between vaccine- and nonvaccine-associated tumors (data not published). In all tumors, there was a mixture of positive and negative-reacting (presumably B) lymphocytes. However, this technique highlighted few to numerous T cells infiltrating the vaccine- and nonvaccine-associated tumors, a feature not noticeable on H&E-stained sections. This line of research will be continued, using additional markers for B cells. After that, reverse-transcriptase PCR will be used to evaluate levels of mRNA for various cytokines in these tumors. High activity of interferon-y and interleukins 2 anti 12 will indicate a dominant TH1 response, indicative of cell-mediated immunity, whereas high activity of interleukin 4 will indicate a dominant TH2 response, associated with humoral immunity. By identifying which lymphocytes are in these lesions and what they are producing (or overproducing), it may be possible to more specifically define their role in the pathogenesis of vaccine-associated neoplastic transformation. Also, insight may be gained into ways to prevent or treat tumors by modulating lymphocyte response and secretion.
From the Department of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104.
References
1 Kass PH, Barnes WG Jr, Spangler WL, et al. Epidemiologic evidence for a causal relation between vaccination and fibrosarcoma tumorigenesis in cats. J Am Vet Med Assoc1993;203:396-405.
2. Hendrick MJ, Shofer FS, Goldschmidt MH, et al. Comparison of fibrosarcomas that developed at vaccination sites and at nonvaccination sites in cats: 239 cases (1991-l992). J Am Vet Med Assoc1994;205:1425-1429.
3. Ellis JA, Jackson ML, Bartsch RC, et al. Use of immunohistochemistry and polymerase chain-reaction for detection of oncornaviruses in formalin-fixed, paraffin-embedded fibrosarcomas from cats. J Am Vet Med Assoc1996;209:767-771.
4. Marshall GM, Vanhamme L, Wong WY, et al. Wounding acts as a tumor promoter in chickens inoculated with avian sarcoma virus 17. Virology1992;188:373-377.
5. Shalaby F. Schuh AC, Breitman ML. Two distinct target cells for v-jun-mediated wound tumorigenesis. Oncogene 1994;9:2579-2588.
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