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therapy in this group of patients will be measured by determining length of patient survival and 
reduction in measurable tumor mass. There is no curative therapy for this stage of disease and 
thus the outcome is predictable enough to allow for an assessment of the results of gene 
therapy. The measurements that will be used are described in Section 7.0. 
2.2.2 Anticipated effect of protocol treatment 
It is anticipated that the administration of the adenovirus wildtype p53 will decrease the rate of 
proliferation of these cells and induce apoptosis of infected malignant cells. This would 
reduce the growth rate or cause regression of primary and/or nodal disease and therefore 
relieve symptoms and potentially prolong the patient's survival. 
2.2.3 Alternative therapies 
Patients with unresectable squamous cell carcinoma of the head and neck that have failed or 
are unable to receive external beam radiotherapy will be considered for this protocol. Existing 
therapies for this condition offers only the potential for short-term palliation. Most patients 
have recurred despite external beam radiotherapy. Patients receiving this treatment have a 
median survival of approximately 6 months. Patients failing brachytherapy would also be 
eligible to receive gene therapy. In those patients that may still have potentially resectable 
tumor, which has failed radiation therapy (alone or in combination with prior surgery or 
chemotherapy), in which tumor can be surgically excised following adenovirus p53 gene 
therapy and then additional gene therapy can be delivered to the surgical bed after the tumor 
volume has been drastically reduced to microscopic disease will also be eligible. These 
patients, although technically resectable have the same prognosis as unresectable patients. 
Patients with unresectable local-regional tumors who have failed surgery or radiation therapy 
have a poor prognosis. Chemotherapy is only palliative and the median survival remains less 
than 6 months. The administration of the adenovirus constructs would not preclude the 
patient from receiving other palliative therapy if the tumor progresses. 
2.3 Structure and characteristics of the biological system 
2.3.1 Restoration of expression of wtp53gene product 
2.3.1 .1 Preliminary studies with plasmid DNA 
The p53 gene is the most commonly altered gene yet described in human cancers. 
To study this gene, a cell culture model system of cell lines varying in p53 expression 
was established. The H322a adenocarcinoma cell line expresses the mutant p53 
protein as shown by the presence of high levels of endogenous p53 mRNA and 
phosphorylated protein. We showed that the H322a cell line has a G:T transversion at 
codon 248 (Arg to Leu) with absence of the wildtype allele. The H322a cell line has a 
homozygous p53 deletion. The H460a and H226b cell lines are homozygous for the 
wildtype p53. Expression vectors for sense (S -p53) and antisense p53 (AS -p53) 
cDNA with a 6-actin promoter were constructed to study the effect of wtp53 
expressed in lung cancer cells with mutant or deleted p53 and the effects of reducing 
wildtype and mutant p53 expression 11 . 
Stable transfectants of p53 mutant cells (H322a) or deleted p53 (H358) expressing S- 
p53 could not be rescued. Failure to isolate colonies expressing sense p53 RNA in 
cells with homozygous mutant or deleted alleles shows that wtp53 can suppress in 
vitro cancer cell growth in cells expressing a mutant p53 or having a homozygous p53 
deletion. 
In general, transfection with antisense p53 (AS -p53) reduced colony formation (10- 
fold) by cells with endogenous mutant p53. This indicates that expression of mutant 
p53 contributes to the transformed phenotype. As expected, cells with wto#* 
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