Protocol THS 94-002 
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little progress has occurred in developing therapeutic strategies that specifically target oncogenes 
and their products. Initially, research In this area was focused on dominant oncogenes, as these 
were the first to be characterized. DNA-mediated gene transfer studies showed acquisition of the 
malignant phenotype by normal cells following the transfer of DNA from malignant human tumors. 
Activated oncogenes of the ras family were Identified by this technique with transfection of human 
DNA into mouse NIH 3T3 cells. More recently a class of tumor suppressor genes have been • 
identified. Mutation or deletion of both copies of a tumor suppressor gene Is required to eliminate its 
function and cause the cell acquire characteristics of the malignant phenotype. 
Tumor Suppressor Gene Mutations in Lung Cancer 
The p53 gene Is the most frequently mutated gene yet identified in human cancers. It is mutated in 
over 50% of human NSCLC[3]. The p53 gene encodes a 375-amino-acld phosphoprotein that can 
form complexes with viral proteins such as large-T antigen and E1B[4]. Missense mutations are 
common for the p53 gene and are essential for the transforming ability of the oncogene. The 
wildtype p53 gene may directly suppress uncontrolled cell growth or indirectly activate genes that 
suppress this growth. Thus, absence or inactivation of wildtype p53 may contribute to 
transformation. However, some studies indicate that the presence of mutant p53 may be necessary 
for full expression of the transforming potential of the gene. Mutations of p53 are common in a wide 
spectrum of tumors[5-8]; they occur in both NSCLC and SCLC cell lines and fresh tumors[8,9]. 
An option for specific targeting of tumor suppressor genes is replacement of a deleted or mutated 
tumor suppressor gene. Progress in the understanding of the critical genes involved in tumor 
development and in technology for altering gene expression logically led to our studies of 
techniques for achieving these options. 
The working hypothesis that we developed is that reversal of a single altered genetic event in 
the cancer cell can potentially reverse critical features of the malignant phenotype of that cell. 
This finding has Important therapeutic implications. Cancer cells have multiple genetic alterations. 
Therapy directed toward oncogenes will be practical only If therapeutic effects occur with targeting 
of one or two genes. It is unlikely that any therapy targeting oncogenes or their products will be 
absolutely specific for cancer cells. If other genes can compensate for loss of normal function by a 
specific oncogene mediated by an antisense construct, the harmful effects of the therapy will be 
reduced. Studies from our laboratory indicate that reversal of a single genetic alteration has 
profound effects on the growth and tumorigenicity of lung cancer cells[10,1l]. Additional support for 
this concept comes from a recent study by Soriano and coworkers[12] in which transgenic mice 
were created that lacked a functional c-src proto-oncogene. The resulting developmental defect In 
the mice was osteopetrosis. The ubiquity of c-src, its high degree of conservation among species, 
and Its role in mitosis suggest that inactivation would be lethal, but this was not the case; viable 
mice were recovered. A possible explanation is that other closely related nonreceptor tyrosine 
kinases such as yes and fyn can compensate for loss of c-src. introduction of a single copy of a 
wildtype tumor suppressor gene into normal cells would be unlikely to have adverse effects if it 
occurred during therapy directed at replacing inactivated tumor suppressor genes in cancer cells. 
Recombinant 
Data on transfection of an antisense K-ras expression vector indicated that Inhibition of expression 
of a single oncogene reduced the growth rate of cancer cells and tumorigenicity in nu/nu mice. 
However, transfected cells retained viability, as did cells with no endogenous K-ras mutation that 
were also transfected with the construct. The wlp53 appears dominant over the mutant gene and will 
select against proliferation when transfected into cells with the mutant gene[11,13]. Normal 
expression of the transfected wtp53 does not affect the growth of cells with endogenous wtp53. 
Thus, such constructs might be taken up by normal cells without adverse effects. This protocol will 
study regional delivery of wtp53 to lung cancer cells In patients with unresectable obstructing 
endobronchial cancers, unresectable local cancers, and malignant pleural effusions not treatable by 
conventional therapies. The efficiency of delivery and gene expression will be evaluated both in lung 
cancer cells and in normal cells in vivo . This Is of Importance for the design of constructs that may 
be useful therapeutically. The effects of these constructs on clinical progression of the cancer will be 
tudied 
Research, Volume 19 
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