Debra J. Wilson

Phase I study of the adoptive immunotherapy of human cancer with lectin activated autologous mononuclear cells

In previous in vitro studies, the authors showed that phytohemagglutinin (PHA) stimulated peripheral blood lymphocytes (PBL) from cancer patients to generate cells that were lytic for fresh autologous tumor but not for lymphocytes or lymphoblasts. Thus, after IRB approval, a phase I clinical protocol was instituted in cancer patients who had failed all other therapy to determine the toxicity and effects, in vivo, of the infusion of large numbers of such PHA activated autologous PBL. Ten patients were treated on the protocol, six with sarcoma, one with melanoma, and three with colorectal cancer. Up to a total of 1.7 × 1011 PBL were obtained from 7 to 15 successive leukaphereses, the cells from each leukapheresis being incubated in vitro in medium containing PHA and human AB serum for 2 days and then reinfused following the next leukapheresis 2 days later. Toxicity encountered included fever and chills in 10/10 patients, headaches in 5/10, nausea and vomiting in 3/10, and requirement for erythrocyte transfusion in 8/10. No evidence for autoimmune disease, abnormal serum chemical or coagulation studies, or pulmonary emboli was found. 111Indium trafficking studies showed distribution of infused cells mainly to the spleen and liver, with some accumulation in the lungs and tumor especially after repeated infusions. In 9/10 patients, activated PBL were detected in the peripheral circulation by the sixth leukapheresis. Evidence for this was found by assaying the incorporation of tritiated thymidine (3H‐Tdr) into, and lysis of fresh tumor cells by, unstimulated PBL from successive leukaphereses. No tumor regression was seen in these patients with bulk disease. These studies demonstrated that large numbers of PHA‐activated PBL can be safely obtained and infused into humans, achieving an increase in the number of circulating activated cells with evidence of migration of cells to tumor, lungs, liver and spleen. Further studies of the use of activated lymphocyte infusion in conjunction with chemotherapy in humans are in progress.

NIH guidelines for research involving recombinant DNA molecules

The National Institutes of Health Guidelines for Research Involving Recombinant DNA Molecules (NIH Guidelines) specifies practices for constructing and handling: (1) recombinant DNA molecules, and (2) organisms and viruses containing recombinant DNA molecules.

Interleukin-2 and lymphokine activated killer (LAK) cells in the treatment of malignant glioma: clinical and experimental studies.

The phenomenon of glioma killing by lymphokine activated killer cells (LAK) was studied. We demonstrate that LAK cells generated by culturing the lymphokine interleukin-2 (IL-2) with peripheral blood lymphocytes from brain tumour patients destroys autologous glioma. The rat 9L glioma model was used to show that LAK killing was tumour-selective as glioma but not syngeneic normal brain tissue was destroyed. The susceptibility of both human and 9L rat glioma to LAK cell killing was markedly diminished by pretreating glioma cells with trypsin or chymotrypsin, but was unaffected by pretreatment with neuraminidase, glycosidases, sodium periodate or hydrocortisone. These results suggest that the cell surface determinant on glioma cells responsible for its tumour selective lysis by LAK is a protein sensitive to trypsin and chymotrypsin. The tumour-selective killing of glioma by LAK in vitro prompted the initiation of a Phase I study in which ten patients with malignant glioma have been treated with direct intracerebral injection of IL-2 or LAK without evidence of systemic or brain toxicity.

The differential inhibitory effects exerted by cyclosporine and hydrocortisone on the activation of human cytotoxic lymphocytes by recombinant interleukin-2 versus allospecific CTL.

Two immunosuppressive drugs, cyclosporine (CsA) and Hydrocortisone (Hy) were examined in parallel for their effect on the generation of cytolytic T lymphocytes (CTL). Peripheral blood lymphocytes (PBL) were stimulated with allogeneic cells to produce allospecific CTL, or with purified recombinant Interleukin 2 to activate lymphokine activated killer cells (LAK). CTL and LAK activity were measured in a 4 hr chromium release assay after 7 days of activation. Lysis by CTL was tested against stimulator PBL (not blasts) and LAK against fresh sarcoma tumor cells. At pharmacologic doses, CsA inhibited only CTL generation, and Hy inhibited only LAK. This greater understanding of the selective role, or roles, in vitro of CsA and Hy provides a basis by which to consider selective immune suppression—and, alternatively, the possibility of combining modalities for a more thorough immune suppression.

Human NK Resistant Tumor Cell Lysis is Effected by IL-2 Activated Killer Cells

In experimental animal systems, trarlsfer of cell populations immune to solid tumors has been successful in mediating the regression of such tumors. In human systems, much conflicting and confusing data has been reported attempting to define the analogous effector cells. Our approach to define human anti-tumor effectors cells was to avoid as many in vitro induced artifacts as possible, and to employ fresh (or cyropreserved, uncultured) surgical biopsy specimens of tumors as target cells, and to generate effectors in human serum with naturally occurring immune stimulants. We have found that normal or cancer patients1 PBL incubated for 2–3 days in preparations of IL-2, including purified recombinant IL-2 (RIL-2) acquires the unique capacity to kill human tumors in 4 hour chromium release assays (1–6). These lymphokine-activated killer cells (LAK) kill NK resistant fresh tumors with an efficiency and polyspecificity that has not been found reproducible in any other anti-tumor lytic system, and therefore provides yet another interesting and potentially biologically relevant system in which to examine the mechanism of cell mediated cytotoxicity. This report addresses some of the basic aspects of LAK killing, such as the killing efficiency of whole populations versus clones, kinetics of lysis as measured by the chromium release assay, correlation of chromium release with the expression of LAK effector cell markers, and morphology and phenotype of LAK effector cells, thereby setting the groundwork for further detailed studies of the mechanism of tumor killing by LAK.