Selective and non‐selective lymphocytotoxicity in human melanoma: Observation on the effect of long‐term culture and fetal bovine serum on target‐cell sensitivity to lymphocytes

Abstract

In vitro cell mediated cytotoxicity (CMC) assays have been conducted in a human melanoma system with a 3H-proline retention technique. Melanoma target cells from long-term cultures ("cell lines") are found to exhibit increased susceptibility for lymphocyte cytotoxicity in comparison to the same target cells from short-term culture. The higher sensitivity of the "cell line" derived target cells is seen with lymphocytes, irrespective of diagnosis of the donor. In parallel experiments with the target cells grown in medium supplemented with fetal calf serum (FCS) and AB+ human serum (from a normal male doner), the melanoma target cells grown with FCS do not show any enhanced cytotoxicity, suggesting no causal relationship of such enhanced sensitivity of "cell line"-derived target cells to "heterologous melanoma antigens" that might have been acquired by the target cells following the use of FCS in tissue culture. In controlled assays of in vitro CMC, lymphocytes from melanoma patients (14/44) exhibited selective cytotoxicity (destruction of only one target-cell type) against the melanoma target cells, whereas only 3/97 control lymphocytes (other malignancies and normal donors) showed such melanoma-selective cytotoxicity. This difference is statistically significant at p less than 0.001. Non-selective cytotoxicity (destruction of two or more unrelated target cell types) was seen with lymphocytes from 9/44 melanoma patients, 13/51 patients with other malignancies and 8/46 normal donors. No correlation of selective cytotoxicity could be established with donors' age, sex, stage of disease, therapy or history of blood transfusion. Such a correlation may emerge as our series becomes larger. Despite the lack of any correlation between selective cytotoxicity and disease status, our study reaffirms the existence of selective cytotoxicity by melanoma patients' lymphocytes against melanoma target cells.

Cellular cytotoxicity in vitro in transitional cell carcinoma of the human urinary bladder: 51Cr‐release assay

“…The question of the existence of a disease-related cytotoxicity exerted by blood lymphocytes from cancer patients has recently caused considerable controversy (Baldwin and Embleton, 1977;Oldham et al, 1975;Takasugi et al, 1974;Oldham et al, 1973;Hellstrom et a/., 1971). Many workers have failed to establish this cytotoxicity and have reported the occurrence of either non-reactivity l, or more often, selective or non-selective cytotoxicity with no obvious relation to the patients' disease (Kiuchi and Takasugi, 1976;Peter et al, 1976;Mukherji et al, 1975;Heppner et al, 1973;Takasugi et al, 1973). These conflicting results are not surprising in view of: (1) the many technological problems inherent in the assay procedures commonly used (de Vries et al, 1975;Oldham et al, 1975;Mukherji et al, 1975;Heppner et al, 1975) target cell lysis; and (3) variations in specific and " non-specific " responses in individual patients during different phases of disease and in relation to therapy.…”

Antigens on melanoma cells detected by leukocyte dependent antibody assays of human melanoma antisera

“…A related vexed question in in vitro diagnostic tests is the reliability of the use of long-term cell cultures for CMC and humoral assays against melanoma antigen. Several reports have shown that cells from long-term cultures appear more susceptible to CMC Mukherji et al, 1975) and that the expression of tumour-associated antigens fluctuates according to the stage of the cell cycle (Cornain et al, 1975; Lewis and Sheikh, 1975).…”

Immunotherapy of Melanoma

“…Here more, a variety of in vivo and in vitro assays the recurrence rate may be high and we lack of cellular and/or humoral immunity have the diagnostic technology to separate out suggested that such patients do, in fact, have such patients who are cured from those who identifiable itnmunity against tumor are not. The potential usefulness of adjuvant associated, or even tumor specific transtherapy has been suggested but cliemo-plantation type antigens ( (0) somewhat both with diffuse metastases and with extensive prior treatment (Nathanson 1974, Mukherji et al 1975a, 1975b). However, attempts to prospectively predict prognosis in high-risk patients, such as those mentioned above, using nonspecific recall antigens have not been successful in this disease (Pritchard et al 1976).…”

“…That is to say, a significant proportion of melanoma patients, even those with early disease, where massive excess of antigen is not present, do not demonstrate detectable cellular immunity to tumor specific or tumor associated membrane extracts (Mukherji et al 1975b, Lui et al 1975). In some of these patients, there is no question that inhibitory factors, either humoral ("blocking" factors) or cellular ("suppressor" eells are responsible for the failure to detect specific tumor-directed immunity (Old 1977).…”

“…In other patients we can only speculate as to whetlier tumor membrane antigens are "hidden" from the appropriate immunologic recognition system, or whether such specific antigens do not, in fact, exist (Old 1977). Regardless of the theoretical explanation for these phenomena, however, the unfortunate result is that, to date, none of the in vivo or in vitro methods for detecting cellular immunity have been sufficiently specific or sensitive to be utilized in a prospective survey for either diagnosis or for the planning of a rational immonotherapy program (Mukherji et al 1975a, 1975b, Peters et al 1975. Thus, we have been left with the necessity of treating patients with a variety of specific, or nonspecific, immunotherapy programs without having precise information in a given patient as to optimal immunogen, dose, route, or' method (Al-Sarraf & Costanzi 1978, Bertogolio et al 1977, DeVries & Rumke 1976, Dufour & Morton 1977, Embleton et al 1978, Golub et al 1977, Holmes et al 1978, Iehiki et al 1978, Serrou et al 1978.…”