Killer Activity Of Lymphocytes Research Articles

Pre-clinical toxicity and immunogenicity evaluation of a MUC1–MBP/BCG anti-tumor vaccine

Mucin 1 (MUC1), as an oncogene, plays a key role in the progression and tumorigenesis of many human adenocarcinomas and is an attractive target in tumor immunotherapy. Our previous study showed that the MUC1–MBP/BCG anti-tumor vaccine induced a MUC1-specific Th1-dominant immune response, simulated MUC1-specific cytotoxic T lymphocyte killing activity, and could significantly inhibit MUC1-expression B16 cells' growth in mice. To help move the vaccine into a Phase I clinical trial, in the current study, a pre-clinical toxicity and immunogenicity evaluation of the vaccine was conducted. The evaluation was comprised of a single-dose acute toxicity study in mice, repeat-dose chronic toxicity and immunogenicity studies in rats, and pilot toxicity and immunogenicity studies in cynomolgus monkeys. The results showed that treatment with the MUC1–MBP/BCG anti-tumor vaccine did not cause any organ toxicity, except for arthritis or local nodules induced by BCG in several rats. Furthermore, the vaccine significantly increased the levels of IFN-γ in rats, indicating that Th1 cells were activated. In addition, the results showed that the MUC1–MBP/BCG anti-tumor vaccine induced a MUC1-specific IgG antibody response both in rats and cynomolgus monkeys. Collectively, these data are beneficial to move the MUC1–MBP/BCG anti-tumor vaccine into a Phase I clinical trial.

MUC1 and maltose‑binding protein recombinant fusion protein combined with Bacillus Calmette‑Guerin induces MUC1‑specific and nonspecific anti‑tumor immunity in mice.

Human mucin1 (MUC1) is a target for immunotherapy. The major problem associated with MUC1‑based cancer vaccines is the weakness of the immunogenicity of MUC1. The present study aimed to develop an efficient cancer vaccine through generating a recombinant fusion protein consisting of MUC1 and maltose‑binding protein (MBP) by inserting seven tandem repeats encoding the human MUC1 gene into the pMAL‑c2 expression vector. Bacillus Calmette‑Guerin (BCG) was used as an adjuvant. MUC1 was found to predominantly induce Thelper type2 (Th2) cell responses. MUC1/BCG and MUC1‑MBP were found to generate Thelper (Th) type 1 and 2 responses, while MUC1‑MBP/BCG induced a Th1 immune profile and stimulated MUC1‑specific cytotoxic Tlymphocyte killing activity. MUC1‑MBP, as well as MBP and BCG alone were found to induce natural killer (NK) cell activity, with MUC1‑MBP/BCG observed to synergistically induce NK cell activity. Furthermore, MUC1‑MBP/BCG significantly inhibited MUC1+ B16 cell growth in mice. These findings show that MBP augments the immunogenicity of MUC1 and that BCG enhances the efficacy of the MUC1‑MBP vaccine. Thus, MUC1‑MBP/BCG may have potential as a cancer vaccine for clinical application.

Interleukin 10 decreases MICA expression on melanoma cell surface

Natural-killer group 2, member D (NKG2D) binds to a variety of ligands, including the major histocompatibility complex (MHC) class I chain-related proteins (MIC) and UL16-binding proteins (ULBP). It is regarded as a co-activating receptor on NK cells, having an important role in the cell-mediated immune response to tumours. We studied the influence of interleukin (IL)-10 on the regulation of MIC and ULBP expression on melanoma cells, and its effect on the cytotoxic function of NK cells in vitro. Here, we show that, in the presence of IL-10, FMS mel and BL mel cell lines decreased MICA and ULBP2 surface expression, whereas MHC class I did not change substantially on the cell surface. MICA mRNA levels decreased in IL-10-treated FMS and IL-10-transduced BL cell lines. Interestingly, we observed that MICB surface expression and its mRNA levels increased upon IL-10 treatment in a melanoma cell line. These changes in NKG2D ligands surface expression patterns owing to IL-10 treatment resulted in an effect on lysis susceptibility mediated by lymphocyte-activated killer cells, as tumour cell lines that displayed a higher decrease of MICA on their surface had lower levels of lysis. In addition, expression of CD107a was downregulated on the surface of NK cells following stimulation with IL-10-treated FMS cells. Our results suggest a novel function for IL-10 in the modulation of NKG2D ligand expression and in the control of cytotoxicity mediated by NKG2D/NKG2D ligand axis.

Immunotherapy in high-risk, chemotherapy-resistant patients with metastatic solid tumors and hematologic malignancies using intentionally mismatched donor lymphocytes activated with rIL-2: A phase II study.

2510 Background: Feasibility and safety of immunotherapy mediated by intentionally mismatched rIL-2 activated killer lymphocytes (IMAK) with no prior stem cells engraftment was investigated in pati...

The expression of 4-1BBL in different gastric cancer cell lines

Objective To study the expression of 4-1BBL and its biological function in different human gastric cancer cell lines. Methods The expression of 4-1BBL in different human gastric cancer cell lines was detected by using immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR). The killing activity of lymphocytes against gastric cancer cell lines was measured by MTT, and interleukin (IL)-2, IL-10 and tumor necrosis factor (TNF)-ct were determined in the supernatant by en-zyme-linked immunosorbent assay (ELISA). Results The RT-PCR revealed that the 4-1BBL mRNA was expressed in MKN-28 (46. 63% ), SGC-7901 (37. 13% ), BGC-823(20. 43% ) and MGC-803 (14. 14% ) in the sequence from high to low levels. The 4-1BBL protein was expressed in MKN-28, SC, C-7901, BGC-823 and MGC-803 in the sequence from high to low levels. MTF analysis indicated that the killing activity was found from high to low in MKN-28, SGC-7901, BGC-823, MGC-803. ELISA analysis showed that IL-2 and TNF-α were found in MKN-28, SGC-7901, BGC-823 and MGC-803, and IL-10 was found in MGC-803, BGC-823, SGC-7901 and MKN-28 from high to low. Conclusion The four different gastric cancer cell lines all express 4-1BBL at the mRNA and protein level. The expression level of 4-1 BBL is related to dif-ferentiation of gastric cancer cell lines. The killing activity of lymphocytes against gastric cancer cell lines is increases with the increased 4-1BBL expression. Moreover, 4-1BBL can regulate tumor immunity by pro-moting the production of TNF-α, IL-2 and IL-10. Key words: Gastric carcinoma; Tumor immunity; 4-1BBL

Clinical and immunological evaluation of zoledronate-activated Vγ9γδ T-cell-based immunotherapy for patients with multiple myeloma

To evaluate the potential anti-tumor activity of zoledronate-activated Vgamma9gammadelta T cells in vivo, we initiated a pilot study involving administration of zoledronate-activated Vgamma9gammadelta T lymphocyte-activated killer (LAK) cells to patients with multiple myeloma. Subjects (n = 6) received four intravenous infusions at 2-week intervals of zoledronate-activated Vgamma9gammadelta T LAK cells generated from the culture of peripheral blood mononuclear cells (PBMCs) in the presence of zoledronate and interleukin-2. If the M-protein level in the patient's serum remained at baseline following four intravenous infusions, the patient underwent four more treatments at 4-week intervals. Subjects (n = 6) received a median of 0.99 x 10(9) Vgamma9gammadelta T LAK cells per infusion. No serious treatment-related adverse effects were observed during the study period. The percentage of Vgamma9gammadelta T cells in PBMCs and absolute numbers of Vgamma9gammadelta T cells in peripheral blood, particularly those of CD45RA(-)CD27(-) effector memory (TEM) Vgamma9gammadelta T-cell subsets increased in all the patients. Percentages of Vgamma9gammadelta T cells and TEM Vgamma9gammadelta T cells in bone marrow also increased in all the patients so far tested. M-protein levels in the serum remained at baseline in four of six patients and increased in two of six patients. Soluble major histocompatibility complex class I chain-related antigen A was detected only in the serum of patients whose M-protein level increased. Administration of zoledronate-activated Vgamma9gammadelta T LAK cells is a safe and promising immunotherapy approach for treatment of patients with multiple myeloma.

Immune killing activity of lymphocytes on Hela cells expressing interleukin-12 in vitro

The killing effects of lymphocytes on Hela cells expressing interleukin-12 (IL-12) in vitro were explored. By using gene transfection technique, full length IL-12 gene was transfected into Hela cells. The expression of IL-12 in Hela cells was detected quantitatively by ELISA; Changes in killing effects of lymphocytes on Hela cells expressing IL-12 were observed by MTT. It was found that Hela cells could express IL-12 between 24 h and 72 h after transfection. Killing activity of lymphocytes on Hela cells expressing IL-12 was significantly enhanced. It was concluded by cell transfection technique, Hela cells could express IL-12 and were more easily killed by lymphocytes.

Characterization of Cord Blood Natural Killer and Lymphokine Activated Killer Lymphocytes Following Ex Vivo Cellular Engineering

Cord blood (CB) natural killer (NK) and lymphokine-activated killer (LAK) cytotoxic cells are poorly characterized but might be used to treat minimal residual and/or recurrent malignant disease. Currently, there is no mechanism to use CB for adoptive cancer cellular immunotherapy after CB transplantation (CBT). Recognizing this as a deficiency, we hypothesized that CB aliquots could be engineered ex vivo for potential donor lymphocyte infusion after CBT. Cryopreserved CB aliquots were thawed, depleted of monocytes, and cultured in serum-free medium alone or serum-free medium with anti-CD3 and interleukins 2, 7, and 12 combined with antibody/cytokines for 48 hours. Immunophenotyping, cytotoxicity, and proliferation were evaluated. A significant expansion of CD3 + was seen, in addition to increases in lymphocyte subsets of CD8 +, CD8 +/CD25 +, and CD3 +/45RO + versus medium alone. A significant enhancement of CD3 proliferation ( P < .001), NK cytotoxicity, NK subset expansion, LAK cytotoxicity, and T-helper 1 subset expansion was also demonstrated. Significant enrichment was seen in NK CD16 +/CD56 +bright, CD16 +/CD56 +dim, CD56 +bright and CD56 +dim/KIR3DL1 +, CD56 +bright and CD56 +dim/KIR2DL1 +, CD56 +bright and CD56 +dim/KIR2DL2 + and CD94 +/NKG2a + subsets. These increases in CB NK subsets were in part secondary to augmentation of cell survival. Further, survival of NOD-SCID mice xenografted with human K562 cells and treated with CB cells expanded with antibody/cytokines was significantly higher than that in animals that received no treatment (phosphate buffered saline) and those that were treated with CB ex vivo expanded in medium alone ( P < .005, respectively). These data suggest that cryopreserved CB cells could be ex vivo engineered for potential use as adoptive cancer cellular immunotherapy for donor lymphocyte infusion after CBT.

Cytotoxic difference of T cells expanded with anti-CD3 monoclonal antibody in the presence and absence of anti-CD 28 monoclonal antibody.

Bulk T cells can be expanded by CD3 stimulation alone (CD3-Ts) or by CD3/CD28 dual stimulation (CD3/CD28-Ts) of peripheral blood mononuclear cells (PBMC). However, few reports have described the difference of features between CD3-Ts and CD3/CD28-Ts. PBMC were stimulated with anti-CD3 monoclonal antibody (mAb) alone or co-stimulated with anti-CD3/CD28 mAbs immobilized on plastic plates, in the presence of rhIL-2 for 4 days, subsequently cultured in the presence of rhIL-2 with no antibody then analyzed. The expansion rate was significantly lower for CD3-Ts (965 + 510-fold, n=5) than CD3/CD28-Ts (2263 + 856-fold, n=5) (p<0.05). The CD4/CD8 ratio, the percentage of CD28(+) cell, and the percentage of T cells with no ability to generate intracytoplasmic interleukin-4 (IL-4) or interferon-gamma (IFN-gamma) were all significantly higher, but, phenotypically, memory cells were lower in CD3/CD28-Ts than in CD3-Ts. The levels of activity of both natural killer (NK) and lymphocyte-activated killer (LAK) cells were lower in CD3/CD28-Ts than CD3-Ts. In comparison to CD3-Ts, CD3/CD28-Ts showed impaired migration toward RANTES. In conclusion, T cells expanded with anti-CD3 and anti-CD28 mAbs differ from those expanded with anti-CD3 alone with proliferation, cytotoxicity, chemotaxis, and phenotype. These differences may exert profound influences on the therapeutic potential of output cells.

Effect of salmonella-infected human monocytes on natural killer cell cytotoxicity. In vitro studies

Various chemicals, including some bacteria-derived components, modulate natural killer cell (NKC) activity. We have analyzed the effect of wild-type Ty2 and of mutant strain TYT1231 Salmonella typhi-infected monocytes (U937 cells and human autologous monocytes) on NKC cytotoxicity of peripheral blood mononuclear cell (PBMC) and highly purified NKC (HPNKC; CD16 +/56 +>95%; the rest corresponding to CD3 + T-cells). PBMC's co-culture with either S. typhi strain infected U937 cells (medium or non-infected U937 cells as controls) resulted in the induction of lymphocyte activated killer (LAK) cell activity showing cytotoxicity against target human NKC-resistant lymphoblastoid Daudi cell line. Comparable experiments using autologous monocytes gave similar results. Co-culture of HPNKC preparations with either S. typhi strain infected U937 cells resulted in increased LAK cell activity against target Daudi cells in each and everyone of the five samples tested; paired Student's t-test p<0.01 for both times (20 and 40 h) tested. Similar to the results observed in the experiments using PBMC, we did not find significant differences in the ability between medium and non-infected cells, or between wild-type S. typhi Ty2 and mutant strain TYT1231 infected U937 cells, to induce LAK activity in HPNKC preparations. PBMC co-incubation with either S. typhi strain infected U937 cells or autologous monocytes resulted in significant increases in IL-12, TNF-α, and IFN-γ secretion. In similar experiments using HPNKC samples instead, infected U937 cells significantly increased IL-12 and IFN-γ, but not TNF-α secretion. PBMC co-incubation with non-infected U937 cells, but not with non-infected monocytes, significantly increased supernatant IL-12 and TNF-α levels (no significant changes in IFN-γ were recorded). Secreted cytokines remained essentially unchanged after co-incubating HPNKC preparation with non-infected U-937 cells. Incubation of PBMC or HPNKC preparations with either S. typhi strain infected U937 cells failed to produce significant changes in the expression of NKC lineage (CD16 +/56 +) or activation (CD28 +, CD69 + and CD95 +) markers. The ability of infected monocytes to induce LAK activity, release NKC cytokines and upmodulate NKC's CD95 + marker expression was essentially the same for both infecting Salmonella strains used. These results suggest a role for NKC in the physiological defensive response against intracellularly infected monocytes representing, perhaps one of the earliest antimicrobial mechanisms of the innate immune system.

Synergistic suppressive effect of double transfection of tumor necrosis factor-alpha and interleukin 12 genes on tumorigenicity of Meth-A cells.

Tumor necrosis factor‐α(TNF‐α) and interleukin 12 (IL‐12), both potent antitumor cytokines, are known to be involved in the host's antitumor immune surveillance in tumor bearers, via different mechanisms. The former enhances the activities of dendritic cells, natural killer/lymphocyteactivated killer (NK/LAK) and cytotoxic T lymphocyte (CTL), while the latter induces Th1‐type cellular immunity and enhances the activities of natural killer T (NKT), NK/LAK and CTL. In the present study, in the expectation of synergistic actions of these cytokines in stimulating the host's immune responses, we investigated the feasibility of a cancer vaccine involving double transfection with both genes in a murine model. The expression of major histocompatibility complex (MHC) class I, class II and B7.1 on the surface of the double transfectants was enhanced as revealed by FACS analysis. A significant decrease in tumorigenicity was observed in mice inoculated with the double transfectants. Cytotoxicity assay revealed that the activities of NK/LAK and CTL from spleens of mice bearing the double transfectants were enhanced. The induction of tumor‐specific immunity was confirmed by rechallenge with parental Meth‐A cells in mice that had rejected the double transfectants. Thus, double transfection of TNF‐αand IL‐12 genes was considered to bring about synergistic suppressive effects on the tumorigenicity of transfectants through the activation of killer cells by produced cytokines and the enhancement of expression of MHC class I, II and B7.1 molecules.

Expression and functional activity of the very late activation antigen-4 molecule on human natural killer cells in different states of activation.

In the present study we describe the expression and functional activity of the alpha4beta1 heterodimer molecule on human natural killer (NK) cells. Flow cytometric analyses showed that fresh and activated NK cells expressed high levels of very late activation antigen-4 (VLA-4) molecules. These cells bound to fibronectin (FN) and to its 38 000-MW proteolytic fragment through the VLA-4 integrin that was blocked with HP2/1 anti-alpha4 monoclonal antibodies (mAbs) and with the FN peptide fragment CS1. No inhibitory effects were observed in the presence of anti-alpha5 mAb, FN peptide fragment CS2 or other irrelevant mAb. Fresh NK cells were unable to aggregate, despite their expression of VLA-4, and only activated (cultured and lymphocyte-activated killer cells) NK cells showed homotypic aggregation with HP1/7 and HP2/4 anti-alpha4 mAb related to cellular activation. These results underline new evidence of how NK cells in different states of activation maintain different constitutive levels of alpha4beta1 integrin activity, and highlight the possibility of a different functional regulation by the cells bearing VLA-4, in the expression of these epitopes and their ability to interact with their ligands.

Apoptosis Induced by Granzyme B-Glycosaminoglycan Complexes: Implications for Granule-Mediated Apoptosis In Vivo

Lymphocyte granule-mediated apoptosis occurs by perforin-mediated intracellular delivery of granule-associated serine proteases (granzymes). A granule-associated proteoglycan, namely serglycin, that contains chondroitin 4-sulfate (CS) glycosaminoglycans is present in the granules of cytotoxic cells. Serglycin acts as scaffold for packaging the positively charged granzymes and probably chaperones the proteases secreted extracellularly. To learn how the interaction of granzyme B (GrB) with serglycin might influence the apoptotic potential of this proteases, we have evaluated a model system where desalted CS is combined with isolated human granzyme. CS-GrB complexes were very stable, remaining undissociated in salt concentrations upwards to 500 mM (pH 7.4). On the basis of a capture enzyme immunoassay that accurately detects GrB, equivalent amounts of active free and CS-GrB, delivered by perforin or adenovirus, efficiently induced apoptosis in Jurkat cells and produced a similar time-dependent increase in caspase-3-like activity. CS-GrB processed isolated caspases-3 and -7 less efficiently than free granzyme. However, when added to cytosolic extracts, rates of processing were nearly equivalent for the two forms, suggesting cationic GrB may nonspecifically bind cytosolic proteins, leading to reduce proteolytic activity. Finally, GrB was found to be exocytosed from lymphocyte-activated killer cells as a neutral, high macromolecular weight complex, which possessed apoptotic activity. Collectively, the results indicate that neutral, high m.w. GrB has the capacity to induce cell death and will be useful to study the mechanism of cytotoxic cell-mediated apoptosis in vitro.

Antibody-IL-12 Fusion Proteins Are Effective in SCID Mouse Models of Prostate and Colon Carcinoma Metastases

IL-12 is a complex cytokine in both its structure and its range of biologic activities. Fusions of this heterodimeric molecule with an intact antitumor Ab were made to test the feasibility and efficacy of targeting IL-12 to tumors to elicit a local immune response. Fusion proteins composed of the human p35 and p40 subunits had IL-12 bioactivities that were nearly as potent on human immune cells as the rIL-12 standard, but were inactive on mouse cells. Hybrid IL-12 fusion proteins composed of mouse p35 and human p40, fused to Ab, were capable of inducing IFN-gamma, but were much less active on mouse spleen cells than a mouse IL-12 standard. Despite this relatively low activity, the hybrid fusion protein was as effective in a SCID mouse model as a fully active Ab-IL-2 fusion protein in eliminating established pulmonary metastases of CT26 colon carcinoma. Specific targeting of a human IL-12 fusion protein to metastatic prostate carcinoma xenografts was also shown to be effective in SCID mice transplanted with human lymphocyte-activated killer cells. These results demonstrate the importance of directing this potent cytokine to the tumor microenvironment and suggest an important alternative to systemic IL-12 administration or gene therapy for increasing its therapeutic index.

Monitoring of effector and target cell stimulation during conjugation by fluorescence polarization

The aim of the present study was to trace early intracellular changes induced in effector and target cells during their conjugation. This was performed by monitoring the intracellular fluorescein fluorescence polarization (IFFP), using the Cellscan apparatus. This apparatus permits the repetitive spectroscopic measurement of individual selected live cells within a population of many cells, while the location of each cell is known and preserved during the various cell manipulations and/or their suspending medium. Both natural killer (NK) and lymphocyte activated killer (LAK) cells were used as effector cells, while NK-sensitive K562 and NK-resistant Daudi cell lines were used as targets. In this study kinetic IFFP measurements were carried out for a period of approximately 4 h following cell attachment. Within minutes following effector-target conjugation, transient reduction of IFFP was observed consecutively, first in the effector and then in the target cells. A continuous reduction of IFFP occurring only in target cells was also found 50 min following conjugation. No reduction in IFFP was observed using NK- and LAK-resistant target cells. Good correlation was found between early stages of conjugation, as assessed by IFFP, and cytolytic efficiency as assessed by 51chromium release assay. When NK-resistant and LAK-resistant target cells were used, no reduction of IFFP was observed.

IL-15 Mediates Anti-tumor Effects after Cyclophosphamide Injection of Tumor-Bearing Mice and Enhances Adoptive Immunotherapy: The Potential Role of NK Cell Subpopulations

The daily administration of IL-15 to cyclophosphamide (CY)-injected mice bearing the 76-9 rhabdomyosarcoma was shown to prolong the period of remission induced by CY. In addition, IL-15 was shown to enhance the efficacy of adoptive immunotherapy. Cytotoxicity assays using spleens from normal and tumor-bearing mice indicated that IL-15 enhanced NK cell activity but there was no evidence for class I-restricted cytolytic T cell activity. To determine whether IL-15 was likely to induce different cytotoxic effectors at the tumor site compared with the spleen, tumors were removed after CY injection and cell suspensions were incubated with IL-15 in parallel with isolated spleen cells. Both populations were seen to expand to yield predominantly cells coexpressing NK1.1 and B220 antigens. However, tumor-associated NK cells were shown to differ from expanded spleen NK cells in terms of the proportions of LGL-1+cells and cells expressing early and late NK cell differentiation antigens. Both expanded populations expressed high NK cell cytotoxic activity but only the spleen cells expressed lymphocyte-activated killer cell activity. It was apparent that the expanded tumor-associated NK cells expressed low-level class I-restricted lytic activity. The potential of activated NK cells in the circulation to exert anti-tumor effects was shown by the adoptive transfer of expanded NK cells to tumor-bearing mice after CY injection when significant prolongation of life was seen in all cases. The data indicate that IL-15 may serve as a useful anti-cancer adjuvant by activating initially the NK cell arm of the immune network.

Development of Vaccine Strategies for the Treatment of B-Cell Malignancies

INTRODUCTION Early observations suggesting some cancer patients can mount a therapeutic immune response to their own tumors (reviewed in Ref. 1) have supported long-standing efforts to develop immunotherapy regimens for the adjunct treatment of human neoplastic disease. Immunotherapy protocols are classified as either passive or active specific, depending on how they are designed to deliver the desired immunity to the patient. Passive immunotherapy regimens are characterized by the passive administration of immune effector components such as lymphokines, antibodies, cytotoxic T cells, or lymphocyte-activated-killer (LAK) cells. By contrast, active specific immunotherapy (ASI) regimens actively induce effector mechanisms and tumor immunity in the patient by deliberate vaccination with a tumor cell or its antigenic components. The ASI approach is preferred because the resulting immunity is longer lasting, and a potential memory response may provide beneficial immune surveillance in the host against further outgrowth of the tumor.

Human tumour antigens recognized by T-cells: perspectives for new cancer vaccines

Conference Article| August 01 1995 Human tumour antigens recognized by T-cells: perspectives for new cancer vaccines B. Van den Eynde; B. Van den Eynde * 1Ludwig Institute for Cancer Research, Brussels Branch, 74 avenue Hippocrate, B-1200 Brussels, Belgium, and Cellular Genetics Unit, Université Catholique de Louvain, B-1200 Brussels, Belgium *To whom correspondence should be addressed Search for other works by this author on: This Site PubMed Google Scholar B. Gaugler; B. Gaugler 1Ludwig Institute for Cancer Research, Brussels Branch, 74 avenue Hippocrate, B-1200 Brussels, Belgium, and Cellular Genetics Unit, Université Catholique de Louvain, B-1200 Brussels, Belgium Search for other works by this author on: This Site PubMed Google Scholar P. van der Bruggen; P. van der Bruggen 1Ludwig Institute for Cancer Research, Brussels Branch, 74 avenue Hippocrate, B-1200 Brussels, Belgium, and Cellular Genetics Unit, Université Catholique de Louvain, B-1200 Brussels, Belgium Search for other works by this author on: This Site PubMed Google Scholar P. Coulie; P. Coulie 1Ludwig Institute for Cancer Research, Brussels Branch, 74 avenue Hippocrate, B-1200 Brussels, Belgium, and Cellular Genetics Unit, Université Catholique de Louvain, B-1200 Brussels, Belgium Search for other works by this author on: This Site PubMed Google Scholar V. Brichard; V. Brichard 1Ludwig Institute for Cancer Research, Brussels Branch, 74 avenue Hippocrate, B-1200 Brussels, Belgium, and Cellular Genetics Unit, Université Catholique de Louvain, B-1200 Brussels, Belgium Search for other works by this author on: This Site PubMed Google Scholar T. Boon T. Boon 1Ludwig Institute for Cancer Research, Brussels Branch, 74 avenue Hippocrate, B-1200 Brussels, Belgium, and Cellular Genetics Unit, Université Catholique de Louvain, B-1200 Brussels, Belgium Search for other works by this author on: This Site PubMed Google Scholar Biochem Soc Trans (1995) 23 (3): 681–686. https://doi.org/10.1042/bst0230681 Article history Received: March 03 1995 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation B. Van den Eynde, B. Gaugler, P. van der Bruggen, P. Coulie, V. Brichard, T. Boon; Human tumour antigens recognized by T-cells: perspectives for new cancer vaccines. Biochem Soc Trans 1 August 1995; 23 (3): 681–686. doi: https://doi.org/10.1042/bst0230681 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Society Transactions Search Advanced Search Keywords: CTL, cytolytic T-lymphocyte, DOPA, dihydroxyphenyl alanine, EBV, Epstein-Barr virus, LAK, lymphocyte-activated killer, MHC, major histocompatability complex, MLTC, mixed lymphocyte-tumour cultures, NK, natural killer, RT-PCR, reverse transcription-polymerase chain reaction, TIL, tumour-infiltrating lymphocyte, TNF, tumour necrosis factor This content is only available as a PDF. © 1995 Biochemical Society1995 Article PDF first page preview Close Modal You do not currently have access to this content.

Perforin and lymphocyte-mediated cytolysis.

We have discussed in the previous sections the recent progress made toward elucidating the regulatory mechanism of perforin gene transcription and the domain structure of the perforin molecule. It appears that the expression of perforin is, at least partially, controlled at the transcription level through the interaction between killer cell-specific cis- and trans- acting factors. One of such cognate pairs, NF-P motif (an EBS-homologous motif) and NF-P2 (a killer cell-specific DNA-binding protein), has been described. The regulatory mechanism of gene transcription, however, is likely to involve multiple factors which act in a coordinated fashion to bring about the most efficient expression of perforin limited strictly to activated killer lymphocytes. Through studies using synthetic peptides and recombinant perforins, it has been suggested that the N-terminal region of the perforin molecule is an important, though not the only, domain responsible for the lytic activity. Further studies are warranted to elucidate the role(s) of other potential amphiphilic structures located in the central portion of the perforin molecule in the overall pore-forming activity. The molecular basis underlying the resistance of killer lymphocytes to perforin-mediated lysis still remains an open question. Preliminary results, however, suggest that the surface protein(s) restricted to killer cells may account for their self-protection against perforin. Based on recent studies using perforin-deficient mice, the involvement of perforin in lymphocyte-mediated cytolysis both in vivo and in vitro has been confirmed. Two functional roles, a direct (lytic) and an indirect (endocytosis enhancer; conduit), both of which may contribute critically to the cell-killing event can be attributed to perforin. The fact that lymphocytes may also employ perforin-independent killing mechanism(s), e.g. Fas-dependent pathway, is beyond the scope of this review. There is, nevertheless, no doubt that these alternative cytolytic mechanisms may also play important roles in immune effector and/or regulatory responses associated with killer lymphocytes. Obviously, we are still a long way from concluding on the functional relevance of each individual cytolytic mechanism seen in different physiopathological situations. Suffice it to say, however, that a wealth of information on lymphocyte-mediated killing has already emerged through the multidisciplinary efforts conducted in our and other laboratories that promise to further dissect this complicated event in the years to come.

IL-12 directly enhances in vitro murine erythropoiesis in combination with IL-4 and stem cell factor.

It has been demonstrated recently that in vivo administration of murine IL-12 to mice enhances the activity of cytotoxic NK cells and lymphocyte-activated killer cells, and that it has antitumor and antimetastatic activity. However, one side effect observed in response to systemic IL-12 treatment is anemia. In the present study, we examined for the first time the ability of IL-12 to affect directly the growth of murine erythroid progenitor cells in vitro. Whereas IL-12 alone or in combination with Erythropoietin (Epo) showed no stimulatory effect on erythroid progenitors, IL-12 potently enhanced the number of erythroid burst-forming unit (BFU-E) colonies formed in response to Epo+IL-4 by 63% and Epo+stem cell factor by 80%. The stimulatory effect of IL-12 occurred in a concentration-dependent fashion, with maximum enhancing effect observed at 50 ng/ml. Furthermore, single cell experiments suggested that the stimulatory effect of IL-12 on erythroid colony formation was directly mediated. Thus, IL-12 can directly enhance murine erythropoiesis in vitro, suggesting that IL-12-induced anemia is mediated through an indirect mechanism.