2.0 OBJECTIVES 
2.1 To determine the period of time over which irradiated, gene-modified 
fibroblasts elaborate IL-4 in vitro . 
2.2 To define the local and systemic toxicity associated with vaccines composed 
of tumor cells admixed with fibroblasts that have been transduced with the 
gene encoding IL-4. 
2.3 To evaluate the local immune response induced by these vaccines. 
2.4 To evaluate the clinical efficacy of these vaccines. 
3.0 EXPERIMENTAL DESIGN 
3.1 Obtaining Tissue for Use in this Study: 
There are a number of obstacles to the timely preparation of tumor vaccines using 
gene -modified cells from patients with melanoma, breast cancer, renal cell 
carcinoma, and colon cancer. With the exception of patients with metastatic 
melanoma, relatively few patients develop accessible subcutaneous metastases. 
Rather, pulmonary metastatic disease is a far more common site of dissemination. 
These lesions can now be resected thoracoscopically with little morbidity and a 
small theoretical risk of mortality (13). This minimally invasive surgical 
technique makes pulmonary metastasectomy a reasonable approach to obtain tumor 
tissue for these studies. Some patients who have previously undergone curative 
resection of their primary malignancy have had their tumor stored in the PCI 
tissue bank. About 200 of these patients are currently followed in the PCI 
outpatient facility. In the event that they develop metastatic disease, they 
would be eligible for this protocol without the need for tumor harvesting. 
Subcutaneous metastases and thoracoscopically resected lung metastases will be 
transported in sterile fashion to the laboratory where they will be processed 
(appendix 19.1). A small representative piece of tissue will be saved for 
histopathologic exam. The remaining tissue will be digested as previously 
described and a single cell suspension made (appendix 19.1). This heterogeneous 
population of cells will be stored in complete medium containing 90% human AB 
serum and 10% DMS0 at -180°C. 
Rapidly proliferating cells are a prerequisite for retroviral gene transduction. 
Unfortunately, our ability to generate tumor lines from these resected metastases 
is limited. In order to obtain large numbers of proliferating cells that can be 
transduced and available for use in a short period of time, we propose to use 
cultured, autologous fibroblasts grown from skin resected at the time of 
metastasectomy (19.2). An elipse of skin measuring 4cm by 1cm will be resected 
from the anterior abdominal wall. This will be performed in sterile fashion in 
the operating room using either general or local anesthesia depending on the 
approach to metastasectomy. All wounds will be closed primarily with either 
sutures or staples. The skin will be transferred in sterile fashion to the 
laboratory where it will be cleared of fat, divided into lmm-2mm diameter pieces 
and plated in T25 flasks containing sterile culture medium and 10% human AB 
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