Clinical testing of IL-2: in vivo administration of IL-2 induces IL-2 dependent non-MHC-restricted cytotoxicity (NRC).
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Antigens, Differentiation↗
Biomedical subjects
J A Hank
Publications and source records attributed to J A Hank.
Destruction of autologous human lymphocytes by interleukin 2-activated cytotoxic cells.
Human and murine lymphocyte populations differentiate into lymphokine activated killer (LAK) cells after in vitro or in vivo exposure to interleukin 2 (IL 2). LAK cells mediate destruction of neoplastic tissue in vitro and have been reported to spare normal tissue. However, systemic toxicity is observed in mice and patients receiving IL 2 infusions. Some aspects of this toxicity are similar to that seen in graft-vs-host disease, suggesting that IL 2 may cause an immune-mediated destruction of normal tissues. We have evaluated this issue by examining the destructive potential of fresh human lymphocytes cultured in media containing highly purified recombinant human IL 2. In the absence of any exogenous antigen or allogeneic stimulating cells, strong proliferative responses were induced after 6 days of exposure to IL 2. Lymphocytes harvested from these 6-day cultures were highly cytotoxic to K562 and Daudi target cells. These IL 2-activated cells were also cytotoxic against autologous and allogeneic normal lymphocyte target cells. This autologous lymphocyte destruction was detected in media containing autologous serum and was directly dependent on the concentration of IL 2 added to the cultures. These studies demonstrate that populations of IL 2-activated lymphocytes, containing LAK activity, can mediate low-level but significant destruction of normal lymphocytes in vitro.
Antigens, Surface↗
Presentation of Candida albicans and purified protein derivative soluble antigens by Epstein-Barr virus-transformed human lymphoblastoid B-cell lines.
Cells other than the macrophage can function as antigen-presenting cells (APCs). These class II-bearing accessory cells include dendritic cells, epidermal Langerhans cells, B cells, murine B-cell tumors, and human Epstein-Barr virus-transformed lymphoblastoid cell lines (EBV-LCL). We investigated the ability of EBV-LCL to present two soluble antigens, Candida albicans and purified protein derivative of tuberculin (PPD). The EBV-LCL derived from B cells of two different individuals can present both antigens to bulk cultures of autologous antigen-primed peripheral blood lymphocytes. The responses of PPD-reactive T-cell clones were weaker to PPD when presented by EBV-LCL than by PBL-APCs, with some clones responding only to PPD presented by PBL-APCs. This suggests that EBV-LCL are not equivalent to PBL monocytes in APC function, and that expression of class II major histocompatibility complex antigen is not sufficient in enabling antigen-presenting capability.
Antigen-Presenting Cells↗
Transplantation of HLA-haploidentical T-cell-depleted marrow for leukemia: autologous marrow recovery with specific immune sensitization to donor antigens.
Autologous marrow recovery without engraftment of donor marrow was observed after bone marrow transplantation (BMT) for two patients with acute lymphoblastic leukemia. Each had received marrow from a haploidentical mixed lymphocyte culture (MLC) reactive donor after pretransplant conditioning with total body irradiation and high-dose cyclophosphamide. To minimize graft-vs-host disease, the marrow was depleted of T cells in vitro by treatment with a monoclonal anti-T-cell antibody and complement. Two weeks after each transplant, reactive lymphocytes were noted transiently in the blood of each patient. Analysis of karyotype, HLA type, and in vitro MLC responsiveness proved the lymphocytes to be of host, not donor, origin. MLC studies showed rapid proliferative responses specifically to stimulating cells from the BMT donor, indicating in vivo sensitization to donor antigens. Return of hematopoietic function was markedly delayed, but it eventually normalized after several months, without evidence of chimerism. These studies confirm that some immune and hematopoietic stem cells of host origin survive the high-dose chemoradiotherapy used as transplant conditioning. Because these immune cells are specifically reactive to donor alloantigens, more potent suppression of host immunity may be needed to prevent nonengraftment of T-cell-depleted, HLA-mismatched bone marrow.
Adult↗
In vitro analysis of donor bone marrow following monoclonal antibody treatment for the prevention of acute graft versus host disease.
Graft-versus-host disease (GVHD) remains a major cause of morbidity and mortality following bone marrow transplantation. The in vitro removal of the GVHD-causing T-lymphocytes from donor marrow is one approach which could control this complication. Treatment of the donor bone marrow with lectins and erythrocyte-forming rosette depletion, anti-T-cell antisera or monoclonal antibodies are methods currently being tested to accomplish this. CT-2 is an immunoglobulin monoclonal antibody specific for the T-cell erythrocyte-forming rosette receptor. Bone marrow from 23 consecutive donors was treated in vitro with CT-2 and complement, prior to infusion, as a potential means of controlling GVHD. Surface marker analysis using erythrocyte-forming rosetting, and OKT-3 and OKT-11 monoclonal antibodies on paired samples of treated and untreated marrow demonstrated a mean depletion to 1% of the original number of T-cells. Proliferative responses to alloantigens and mitogens as well as cytotoxic and natural killer cell function were tested and found to be markedly reduced. Despite these effects on T-lymphocytes, viable hematopoietic stem cell colonies were retained. Clinical results following the in vitro T-lymphocyte depletion of donor bone marrow for the 8 histocompatible and 15 nonhistocompatible bone marrow transplantation are reported. Prompt engraftment with minimal GVHD, despite no posttransplant GVHD prophylaxis, was seen in seven of the matched patients. In the nonhistocompatible bone marrow transplantation, failure of engraftment occurred in 11 patients. Grades III-IV GVHD were seen in two of the four patients that engrafted despite good T-lymphocyte depletion. No predictive correlation could be found between the in vitro analysis of marrow following CT-2 treatment and clinical outcome.
Acute Disease↗
Clinical response of a patient with diffuse histiocytic lymphoma to adoptive chemoimmunotherapy using cyclophosphamide and alloactivated haploidentical lymphocytes. A case report and phase I trial.
Adoptive chemoimmunotherapy has cured experimentally induced tumors in animals, but its clinical use has been limited. Six patients were treated with refractory neoplasms in a Phase I study with cyclophosphamide (CPM) and alloactivated haploidentical lymphocytes. Patients received an immunosuppressive dose of CPM (800 mg/m2) followed by haploidentical lymphocytes primed in vitro with alloantigens in mixed lymphocyte culture (MLC). One week later patients received a second infusion of alloactivated lymphocytes expanded in T-cell growth factor (TCGF). The total number of cells given to each patient progressively increased, with a single patient receiving 35.5 X 10(9) cells. Transient febrile responses and delayed-type hypersensitivity reactions at the intravenous sites were the only toxicities noted. A complete clinical response lasting 12 weeks was seen in a single patient with diffuse histiocytic lymphoma. Our experience indicates that adoptive chemoimmunotherapy can be given to patients safely and merits further clinical testing.
Blood Transfusion↗
A human T cell clone that mediates the monocyte procoagulant response to specific sensitizing antigen.
A panel of human purified protein derivative of the tubercle bacillus (PPD)-reactive T cell clones was derived by cloning out of soft agar followed by cultivation on inactivated feeder cells in the presence of interleukin-2. 1 of 4 clones tested was able to mediate an increase in monocyte procoagulant activity (PCA) in response to PPD. All four clones had identical surface marker phenotypes (T4+, T8-) and proliferated in response to antigen. The reactive T cell clone possessed no PCA of its own, but upon being presented with PPD was able to instruct monocytes to increase their expression of PCA. Antigen presentation could be performed only by autologous monocytes; allogeneic monocytes from donors unrelated to the donor of the reactive clone could not present antigen to cells of the clone in a way that would initiate the procoagulant response. Cells of the reactive clone did not mediate increased monocyte PCA in response to Candida, even though peripheral blood mononuclear cells from the donor demonstrated increased PCA to both Candida and PPD. Thus, the PCA response to specific antigen can be mediated by a single clone of cells that shows specificity in the recognition of both antigen and antigen presenting cell.
Antigen-Presenting Cells↗
Relapse of host leukemic lymphoblasts following engraftment by an HLA-mismatched marrow transplant: mechanisms of escape from the "graft versus leukemia" effect.
Leukemic relapses and graft versus host disease (GvHD) remain major complications following allogeneic bone marrow transplantation for leukemia. We present clinical and laboratory details for an eight-year-old boy who received a T-cell-depleted HLA-mismatched marrow transplant as therapy for acute lymphoblastic leukemia (ALL) in second remission. Engraftment with donor marrow was prompt and without any acute GvHD. Nevertheless, the patient's original ALL recurred and proved fatal. The patient remained a chimera with persistent donor lymphocytes present at the time of posttransplant relapse and subsequent to treatment with unsuccessful reinduction chemotherapy. In vitro immune studies showed that these leukemic cells could be recognized and destroyed by the donor's lymphocytes. The relapse itself suggests, however, that the donor's lymphocytes did not effectively destroy the patient's histoincompatible ALL cells in vivo following establishment of the chimeric state. Potential mechanisms are presented to account for this presumed "escape" from the postulated "graft versus leukemia" effect.
Acute Disease↗
HLA typing and karyotyping of limited numbers of lymphocytes is facilitated by in vitro expansion with TCGF.
It is often difficult to obtain sufficient quantities of leukocytes from bone marrow (BM) transplant recipients for post-transplant immunological monitoring and evaluation of engraftment. We have developed a cell culture expansion technique that allows HLA-A,B, and DR typing as well as karyotyping on limited numbers (less than 10(7)) of lymphocytes. Lymphocytes from healthy control donors were allo-activated in vitro and further expanded by culturing in T-cell growth factor (TCGF). After 14 days of culture, these cells were sent for blind HLA-A,B and DR typing and for karyotyping. Surface marker studies showed that 90% of the T-cells expanded in growth factor are OK-la positive. HLA typing with these DR-bearing in vitro expanded T-cells showed excellent correlation with HLA typing on fresh unexpanded lymphocytes. Karyotyping of expanded T-cells showed normal male or female cells as expected. This technique may be especially valuable when testing pediatric or adult patients with very low lymphocyte counts.
Bone Marrow Transplantation↗
Transplantation of HLA-haploidentical T-cell-depleted marrow for leukemia: addition of cytosine arabinoside to the pretransplant conditioning prevents rejection.
A total of 41 patients with hematologic malignancies (other than acute leukemia in relapse) received allogeneic bone marrow transplants at the University of Wisconsin from 1 April 1980 through 31 March 1984. In an effort to minimize graft-versus-host disease, marrow was depleted of T-lymphocytes in vitro with monoclonal anti-T-cell antibody and complement prior to infusion for seven of 19 recipients of marrow from HLA-identical, MLC-nonreactive siblings, and for all 22 recipients of marrow from MLC-reactive HLA-haploidentical donors. The recipients of HLA-identical T-depleted marrow all showed excellent engraftment following standard pre-BMT conditioning with cyclophosphamide and total body irradiation. In contrast, five of five recipients of T-depleted haploidentical marrow failed to engraft following this same conditioning regimen. The addition of cytosine arabinoside to the pretransplant conditioning appeared to correct this problem, allowing engraftment in 14 of 17 subsequent patients. These clinical results, coupled with prior in vitro data, demonstrate the need to adequately suppress residual host-versus-graft immunity in order to prevent the rejection of T-cell-depleted HLA-haploidentical bone marrow.
Adolescent↗
The in vivo distribution of indium-111 labeled in vitro alloactivated syngeneic lymphocytes in a patient with relapsed acute myelomonocytic leukemia: implications for adoptive chemoimmunotherapy.
A single patient who had a leukemic relapse six months after receiving a syngeneic bone marrow transplant was given "adoptive chemoimmunotherapy." Lymphocytes from the patients identical twin were alloactivated and grown in T cell growth factor in vitro. After receiving chemotherapy to reduce the number of relapsed leukemia cells, he received 1.1 X 10(10) in vitro alloactivated twin lymphocytes via intravenous and intraperitoneal injections. Although progressive Candidal pneumonia was fatal and prevented analysis of either efficacy or delayed toxicity of this potential form of therapy for this patient, radioactive 111In labelling and scanning showed dissemination of these lymphocytes following either injection route, with no clinical evidence of immediate toxicity.
Adult↗
Recognition of autologous lymphoblastoid cells by cloned human T cells: inhibition of cytotoxicity by anti-HLA antibody but not by antibody to Epstein-Barr virus induced cell surface antigen.
Cytotoxic T cells (Tc) can be primed in vitro to specifically kill autologous Epstein-Barr virus (EBV) transformed lymphoblastoid cell line cells (LCL). The antigen recognized by Tc on the LCL cells is designated LYDMA (lymphocyte determined membrane antigen). Recognition of LYDMA may also require simultaneous recognition of HLA determinants. Despite its immunogenic potency, the genetic origin and molecular structure of LYDMA remains undetermined. Most LCL cells do not produce EBV virus, but all LCL cells appear to express an immunogenic, non-HLA class I, 47,000 Dalton determinant which is detected by several independently derived mouse monoclonal antibodies. This determinant is not readily detected on peripheral blood lymphocytes, nor on EBV-associated Burkitt's lymphoma cell lines. We examined the potential relationship between this 47,000 MW Epstein-Barr virus associated cell surface antigen (the EBVCS antigen) and LYDMA. Populations of Tc generated by in vitro activation with irradiated autologous LCL cells were potent at destroying EBVCS bearing autologous LCL cells. The addition of anti-EBVCS antibody to this cytotoxic interaction had no suppressive effect. Furthermore, single cell derived, LYDMA reactive, Tc clones able to specifically destroy autologous LCL were studied. Cytotoxicity could be inhibited by monoclonal antibody to HLA-Class I determinants, but not by monoclonal antibodies to EBVCS. Thus, these LYDMA reactive Tc clones do not recognize the same antigenic determinant as the monoclonal antibodies specific for EBVCS. It remains uncertain whether this 47,000 MW glycoprotein that carries the serologically detected EBVCS antigen may be directly recognized by human T cells.
Antibodies, Monoclonal↗
Cell mediated lysis of lymphocytes expressing bacterial antigens.
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Antigens, Bacterial↗
Heated lymphocytes express HLA-DR antigens despite their inability to stimulate in MLC.
We have utilized serological techniques and mixed lymphocyte culture (MLC) reactions to examine HLA-DR and HLA-D expression by heated (45 degrees C for 1 h) lymphocytes in order to study the functional relationship of these antigens. Heated lymphocytes do not stimulate proliferation of allogeneic lymphocytes in MLC, yet they express HLA-DR antigens. The fraction of peripheral blood lymphocytes (PBL) expressing DR is not altered by heating, nor is the staining intensity altered as detected by fluorescence microscopy. Alloantisera to "B cell alloantigens" recognize HLA-DR determinants on heated cells without any detectable change in either specificity or quantitative cytotoxic effects. Flow cytometry with monoclonal antibody demonstrates only minimal decrease in HLA-DR expression after heating. Thus stimulation in MLC requires more of the stimulating cell than the mere expression of HLA-DR.
B-Lymphocytes↗
HLA identical leukemia cells and T cell growth factor activate cytotoxic T cell recognition of minor locus histocompatibility antigens in vitro.
Lymphocytes from a healthy HLA-identical bone marrow transplant donor were tested for their ability to destroy her brother's acute myelogenous leukemia blasts in vitro. Primary mixed lymphocyte culture (MLC) and cell-mediated lysis (CML) responses between the patient's remission (pretransplant) and donor's lymphocytes were negative. Stimulation of donor lymphocytes for 7 d in vitro with irradiated leukemia cells, leukemia cells plus allogeneic irradiated lymphocytes, or a pool of irradiated lymphocytes from 10 donors, did not activate any cytotoxic cells able to destroy the HLA identical leukemic blasts. Further culturing for 7 additional d in T cell growth factor (TCGF) generated lymphocytes that induced effective cytotoxicity against the leukemic blasts, but not against autologous lymphocytes. Effective killing against the leukemia was observed only in cultures initially stimulated with the irradiated leukemia cells. These cytotoxic cells were maintained in TCGF and mediated persistent killing against the leukemic target cells. They were also able to destroy lymphocytes from the patient's mother and father, but not from an unrelated cell donor. This suggested specific recognition of non-HLA antigens inherited by the patient, that were foreign to the HLA identical bone marrow donor. These lymphocytes were cloned by a limiting dilution technique and one clone maintained cytotoxicity to the AML blasts and the father's lymphocytes, but not lymphocytes from the mother or an HLA-identical donor. This cytotoxicity was inhibited by a monoclonal anti-HLA antibody. Thus, in vitro sensitization of this sibling's lymphocytes with AML blasts followed by TCGF expansion, and cloning, enabled the detection of HLA-restricted cytotoxic cells that recognize minor locus histocompatibility antigens. This immune recognition may be relevant to the "graft vs. leukemia" effect that has been observed in leukemic animals and patients following histocompatible hematopoietic transplants.
Adult↗
Proliferative responses to a nonspecific factor produced by irradiated stimulating cells can simulate antigen-specific secondary responses in the primed lymphocyte test.
The primed lymphocyte test (PLT) was utilized to investigate the biologic relationship between alloantigens and soluble environmental antigens. Human lymphocytes primed in vitro to the soluble antigens Candida (CAN), Tetanus (TET), or Purified Protein Derivative of Tuberculin (PPD) gave a strong proliferative response when restimulated with the initial soluble antigen in the presence of irradiated peripheral blood lymphocytes (PBL) acting as antigen-presenting cells (APC). Surprisingly, the soluble antigen-primed cells as well as alloantigen-primed cells responded to other soluble antigens in the presence of APC. By testing primed cells with supernatants derived from irradiated lymphocytes plus soluble antigen, it was apparent that the responses observed were in part due to stimulation by a soluble factor produced by the irradiated "APC" in response to the soluble antigen rather than to recognition of widespread cross-reactivity by the primed cells. This "nonspecific" factor production could be diminished by increasing the dose of irradiation to the APC or by using a T lymphocyte depleted- (SRBC-E-) APC population. In addition, certain antigen-reactive T cell clones did not respond to the nonspecific factor to the same degree as the primed bulk cultures. Nevertheless, the recognition of nonspecific stimulation induced by factors produced by irradiated lymphocytes is critical in the interpretation of primed lymphocyte responses to alloantigens or soluble antigens.
Antigens, Fungal↗
Soluble bacterial antigen induces specific helper and cytotoxic responses by human lymphocytes in vitro.
Fresh human T lymphocytes respond proliferatively to soluble antigens, such as PPD, provided the cell donor has been immunized to those antigens in vivo. This proliferative response requires the addition of antigen-presenting cells bearing autologous HLA-D antigens. This report demonstrates that the proliferative response to PPD activates a collaborative response that aids in the activation of alloreactive cytotoxic T cells (Tc). Furthermore, the proliferative response to "soluble" PPD alone activates a population of Tc specifically able to destroy autologous target cells passively labeled with PPD. Allogeneic PPD-labeled targets are killed less effectively. This PPD-specific cytotoxicity is not inhibited by soluble PPD, but is competitively inhibited by PPD-labeled autologous cells. Thus, in vitro stimulation with "soluble" bacterial antigen activates both helper and cytotoxic recognition of antigen-altered self by a mechanism comparable to the recognition of viral or hapten-altered cells.
Antigens, Bacterial↗
Influence of the virulence of Mycobacterium tuberculosis on protection induced by bacille Calmette-Guérin in guinea pigs.
The protective efficacy of two bacille Calmette-Guérin (BCG) vaccines was examined in guinea pigs infected by the respiratory route with strains of Mycobacterium tuberculosis differing in virulence. Virulence was defined as the degree of tissue damage (weight) of primary lesions excised from lungs of unvaccinated guinea pigs killed 28-42 days after infection. Groups of animals vaccinated with BCG-Copenhagen (strain no. 1331), a vaccine of high potency, or those vaccinated with BCG-Prague (strain no. 725), an experimental vaccine of low potency, and groups given placebo were challenged six weeks later with one of three challenge strains differing in virulence. Protection was assessed from the difference in the number of tubercle bacilli recovered from excised primary lung lesions or from primary lesion-free lung lobes of vaccinated vs. unvaccinated animals. The virulence of the challenge strain influenced the efficacy of BCG vaccination; however, the results of other studies with a laboratory strain were in general replicated.
Animals↗