The effect of purification on the immunogenicity of tumor-specific transplantation antigens

Abstract

Immunization with the tumor-specific transplantation antigens (TSTA) of experimental, chemically induced sarcomas engenders specific host resistance to challenge with viable, homotypic neoplastic cells. The strength of tumor resistance depends upon the physical state of the TSTA used for immunization. Treatment with 10(5)-10(6) irradiated tumor cells, a 2-log dose range, induces complete rejection of neoplastic challenges, while immunization within a 1-log dose range with crude 3 M KCl or with 2.5% butanol extracts containing TSTA evokes a weak state of resistance characterized by decreased outgrowth of tumor challenges, but not neoplastic regression. The reduced immunogenicity may be due to either contamination with substances that antagonize host resistance, for example by induction of suppressor cells, or an intrinsic limitation by virtue of the molecular properties of extracted compared with cell-surface TSTA. MCA-F and MCA-D, two noncross-reactive fibrosarcomas induced in C3H/HeJ mice with 3-methylcholanthrene, were employed to compare the relative immunogenic activity of intact tumor cells, 2.5% butanol extracts, and materials sequentially purified by preparative isoelectric focusing (pIEF), preparative isotachophoresis (pITP), and high performance gel permeation chromatography (HPGPC). Immunoprotective TSTA activity purified 50,000-fold by this protocol extended the effective dose range by four to five logs: 15 pg to 1.5 micrograms MCA-F or 1 pg to 10 ng MCA-D antigen-induced specific host resistance. However, despite the appreciable purification of TSTA, immunization with extracted materials only delayed neoplastic outgrowth. They induced neither immediate rejection nor only temporary progression of transplanted tumor cells. Thus, purified TSTA preparations by themselves lack the immunogenic properties of intact cells that result in maximal induction of tumor resistance.

Tumor rejection antigens of chemically induced sarcomas of inbred mice.

“…This has proven to be extremely difficult, and only in the case of BALB/c Meth A sarcoma have antibodies to surface antigens with TRArelated characteristics been repeatedly generated (12,15,16). For this reason, attempts to isolate TRAs have relied on transplantation tests to monitor antigen purification, and DuBois et al (17)(18)(19) and other workers (20)(21)(22)(23) have characterized cellular fractions with TRA activity. In the present report, we describe the purification and characterization of TRAs from two antigenically distinct BALB/c sarcomas and compare their properties with antigens isolated by other investigators.…”

Noncytolytic extraction of cell surface antigens using butanol

“…The TSTA present in crude butanol extracts (CBE) displayed a 20-fold higher specific activity than did 3M KC1 extracts, and the activity was recovered with high yield following preparative isoelectric focusing (27). The butanol-extracted antigens have proved stable during isolation, and the TSTA from three different MCA-induced tumors have since been purified 33,000-fold from CBE by high performance gel permeation chromatography and 2.2 x 106-fold by preparative isotachophoresis (49)(50).…”