Ronald D. Paul

Selective action of alkylating agents against cells participating in suppression of antibody responses

The effect of several alkylating agents on the induction and expression of specific suppressor cell activity induced by supraoptimal immunization (SOI) with (4 x 10(9) ) SRBC was studied. Splenocytes taken 8-28 days after SOI and transferred to normal syngeneic recipients together with optimal dose of (3 x 10(8) ) SRBC caused 70-90% suppression. By contrast splenocytes harvested 2 days after SOI did not exert a significant suppressive effect. Treatment of donor mice with 30 mg/kg BCNU, CCNU, or MeCCNU, 8 mg/kg Melphalan or as much as 200 mg/kg Cy 2 days before SOI uniformly had no effect on the subsequent development of suppressor cells. By contrast, different drugs had diverse effects when injected after SOI: both BCNU and Cy injected 2 days post SOI alleviated suppression, whereas CCNU, MeCCNU and Melphalan injected 2 days post SOI were without effect. Another diversity between BCNU and Cy was noticed when the administration of drugs was delayed further. While Cy alleviated suppression 12 days post SOI, BCNU was ineffective at this time. Dose response and time course studies revealed that the effect of Cy was most severe when injected 2 days post SOI and gradually diminished with the passage of time after SOI. These results have been discussed in the light of the current concepts of multiple cell participation in the induction and expression of suppressor cell function.

IMPROVED ALLOGRAFT SURVIVAL USING HIGHLY ENRICHED POPULATIONS OF RAT ISLETS

A method is described for the purification of islets before the cells are placed in tissue culture, thus permitting the transplantation of islets cultured for three days against major histocompatibility barriers without adjuvant immunosuppression. Mixed lymphocyte culture reactions were carried out with three rat strain combinations and the in vitro responses were correlated with the in vivo survival of islet allografts. These results showed that islet allograft acceptance is independent of the degree of histoincompatibility between different rat strains.

Cytotoxic Effector Mechanisms Involved in the Immunity Against Mammary Tumors

There is considerable evidence that the initiation and growth of tumor masses can be modulated by the immune system. This influence not only applies to the primary tumor burdens, but is also apparent in the establishment and development of metastases. Several immunologic components are involved in these processes, however, the relevance of the cell-mediated compartment has been amply documented. The interactions among regulatory, helper, and effector cells are undeniably important, however, in many instances, the direct actions of several types of immune cells with lytic potential represent an important aspect of tumor immunology. Using animal models of breast cancer and, on a smaller scale, clinical experimentation, much information has been generated in several laboratories, providing insight into the relative contributions of various types of cytotoxic effectors operative against mammary tumors. Our understanding of the function of such cells in the context of in vivo tumor development is a first step in devising logical protocols for immu-notherapeutic approaches.

Emergence of a B lymphocyte population with ADCC effector function in mammary tumor bearing mice.

Differential expression of antibody dependent cellular cytotoxicity (ADCC) effectors was studied in normal Balb/cCrgl mice and those bearing a chemically induced 7, 12 dimethylbenzanthracene mammary adenocarcinoma. Depletion of macrophages from normal mouse splenocytes by Sephadex G‐10 columns resulted in elimination of ADCC. Further separation of the normal G‐10 nonadherent splenocytes on nylon wool columns did not result in any population with significant cytotoxicity. However, Balb/c mice bearing mammary tumors showed enhanced levels of ADCC which were not eliminated by macrophage removal. Lymphocytes from tumor bearers further separated on nylon wool yielded nonadherent and adherent populations both capable of effecting significant ADCC. Treatment of the nylon nonadherent cells of both normal and tumor bearing mice with anti‐asialo GM1 (AGM1) and complement decreased the ADCC responses. The same treatment only marginally affected cytotoxic levels of nylon adherent cells from tumor bearers, indicating that these effectors are primarily of non‐NK lineage. In addition, G‐10 nonadherent, nylon adherent cells from tumor bearers separated on a fluorescence activated cell sorter based on the presence of surface immunoglobulins (slg) revealed that both the slg‐ and slg+ (98% pure) sorted cells were capable of functioning in ADCC. To determine whether in the tumor mice the 2% of slg‐ cells present in the slg+ sorted population were the ADCC effectors, mixing experiments were done in which up to 10% of slg‐ cells from tumor bearers were added to nylon adherent cells from normal mice. No significant increases in ADCC levels were found over that of normal mice. These experiments indicate that the 2% slg‐ cells were not the ADCC effectors nor were they inducing normal B cells to exert this type of cytotoxic reaction in vitro. To further substantiate the B cell lineage of the slg+ ADCC effectors, surface immunoglobulins were removed with protease treatment. After a 36 hr incubation, 92% of the cells had regenerated their slg. The results presented in this paper demonstrate that various splenic lymphoreticular populations from tumor bearers possess an enhanced cytolytic activity against antibody coated target cells. Among these is a unique nylon adherent slg+ cell that is capable of functioning as an ADCC effector.

Treatment of a Murine Leukemia with Chlorambucil Bound Monoclonal Antibodies

Mice bearing a murine T cell leukemia (P388) were treated with a battery of monoclonal antibodies specific for normal T cell differentiation antigens. Therapy was evaluated during the early course of disease by DNA cell cycle analysis and recovery of leukemia cells from tumor bearing mice. This form of therapy was shown to be ineffective. However, treatment with chlorambucil bound monoclonal antibodies was shown to be effective in reducing the number of leukemia cells undergoing DNA synthesis and mitosis, in addition to diminishing the tumor burden as compared to control mice. The potential benefits of this form of therapy are discussed.

Immunomodulation by Corynebacterium Parvum

Corynebacteriumparvum which has recently been reclassified as a Propionibacterium (1) was initially noted for its ability to stimulate the reticuloendothelial system (RES) as manifested by increased clearance of particulate material from the circulation and increased spleen and liver weights (2–5). Since then, the ability of this organism to modify various immune functions has been extensively studied.

Mammary Tumor Virus Infections and Immunity

Mammary tumorigenesis appears to be influenced by several factors including genetic background, hormones and the immune response. In animal models there is a correlation between infection with mouse mammary tumor virus (MMTV) and an increased incidence of spontaneous mammary tumors. Exogenous MMTV is transmitted from high mammary tumor incidence strains to certain low incidence strains through the milk (1) with the result of increased numbers of tumors in the latter. There is evidence suggesting that MMTV infection results in integration of this retrovirus into the mammary glandular tissues (2). In recent years the possibility has been explored, that expression of an endogenous counterpart of MMTV may be involved in the etiology of mammary tumors. The observation by Hageman et al. (3) of MMTV virions in mammary tumors arising in some low incidence mouse strains that had different characteristics from the classic exogenous milk transmitted virus, supports a role of endogenous MMTV in tumorigenesis.