146 

 minutes which resulted in 58% inhibition of cell prolifer- 

 ation 4 8 hours later. Exposure to the same amount of ADGG- 

 Con a in the presence of ImM MDM resulted in a total of 

 6.7% inhibition. Almost complete protection from the toxic 

 effects of ADGG-Con A was achieved by blocking the binding 

 of the Con A portion of the conjugate to cell receptors 

 with saccharide ligand. 



The cell binding and cytotoxicity studies strongly 

 point to the targeting potential of ADGG-Con A for delivery 

 of amanitin to Con A receptor bearing cells. A numerical 



estimation of this targeting ability was obtained by ex- 



3 



posing H-7 CHO cells to H-DM-ADGG-Con A for 15 minutes at 



room temperature followed by incubation in culture media 

 at 37 °C for 1 hour. At the end of the hour, the culture 



media and three PBS washes of the cells were counted for 



3 

 H-DM-ADGG-Con A activity which eluted from the cells upon 



interaction with the glycoproteins and carbohydrates pres- 

 ent in the serum containing medium. The cells were 

 counted separately to determine the amount of Con A that 

 remained irreversibly associated with the cell and was 

 presumably internalized. A total of 60% of the activity 

 was found in the medium leaving 40% associated with the 



cell or 4.21 x 10 molecules/cell. Assuming an intracel- 



-9 

 lular volume of 3.09 x 10 ml/cell (Noonan and Burger, 



2 

 1973; volume calculated from a surface area of 2200y 



given for a 3T3 cell), the intracellular concentration of 



-9 



ADGG achieved is on the order of 2 x 10 M. This value 



