In Vivo Marking of Spontaneous or Vaccine-Induced Fibrosarcomas in the Domestic House Cat, Using an Adenoviral Vector Containing a Bifunctional Fusion Protein, GAL-TEK

Abstract

We evaluated the ability of a replication-deficient, recombinant adenoviral vector to transfer the bifunctional gene GAL-TEK, which expresses a marking/therapeutic gene product, to naturally occurring cat fibrosarcomas in situ. GAL-TEK contains an in-frame fusion of the bacterial LacZ gene for histochemical marking of tumors with β-galactosidase (β-Gal) and the HSV tk gene for enzyme-prodrug activation of the prodrug ganciclovir (GCV) to induce selective tumor cell killing. GAL-TEK bifunctional marking and cell killing activities were tested in vitro after adenoviral vector infection of HT1080 human fibrosarcoma cells. The tk activity of GAL-TEK is shown to be almost as potent as HSV tk to catalyze conversion of GCV to GCV nucleotides and promote selective cell killing. Using 8 cats with recurring 2.5-cm² fibrosarcomas that either arose spontaneously or were induced by vaccine, we determined experimentally the administration routes and times required for optimum GAL-TEK gene transfer by β-Gal histological staining and reverse transcriptase polymerase chain reaction to the multiple compartments of the growing fibrosarcomas consonant with minimizing collateral infection of neighboring tissues and other unwanted side effects. The vaccine-induced cat fibrosarcoma served as the model for gene transfer into naturally occurring cancer cells. We utilized a recombinant adenoviral vector carrying the fusion protein GAL-TEK to evaluate gene transfer into fibrosarcomas in situ. Using two different intratumor injection protocols and titers ranging from 10⁴ to 10⁸ pfu/ml, we were able to show, by reverse transcriptase polymerase chain reaction and β-galactosidase histochemical staining, good transfer of GAL-TEK to the different compartments of the growing tumor at all levels of pfu examined. However as titers increased so did collateral infection of neighboring tissues, especially when utilizing a protocol that involved circumscribing the tumor with additional injections of recombinant adenovirus.