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W M Kast

Publications and source records attributed to W M Kast.

At least 73 records · Page 4Linked to original sources
PubMed1996

Peptide-based cancer vaccines.

The application of molecularly defined vaccines composed of a limited number of tumor-specific T cell epitopes has resulted in protective antitumor T cell immunity in several mouse tumor models. The first encouraging results with such vaccines have been obtained in human beings. The development of the next generation of rationally designed vaccines that are both effective and safe for application in a clinical setting requires comparison of different modes of delivery of tumor-associated T cell epitopes in multiple epitope constructs.

Animals↗
PubMed1996

Immunotherapy of cancer by peptide-based vaccines for the induction of tumor-specific T cell immunity.

Recent progress in defining the molecular nature of antigens and in finding ways to manipulate T cell-mediated immune responses may provide new modalities for cancer treatment. In this report, we review preclinical studies as well as the first clinical trials with vaccination strategies aiming at the induction of anti-tumor immunity. In particular, we focus on the development of a vaccine against human papillomavirus-induced cervical carcinoma.

Animals↗
PubMed1996

The current status of therapeutic HPV vaccine.

Cervical carcinoma is the second most common cause of cancer-related deaths in women worldwide. Recurrences occur in 15% of the patients after optimal treatment of low-risk early-stage disease. Treatment results of recurrent disease are relatively poor and for this reason new therapeutic strategies are warranted. Viral infection with human papillomavirus seems to have an essential part in the aetiology of cervical carcinoma. Evidence for the assumption that cervical carcinoma, among other malignancies such as melanomas, renal malignancies and Kaposi sarcoma, are immunogenic is provided by the fact that these malignancies grow more rapidly in the presence of systemic immunosuppression. Spontaneous regression for these tumour types is also described and immunohistochemical studies show extensive infiltrates in the tumour, consisting of immunocompetent cells. It is thus postulated that cellular immunity, and mainly the T-cell system plays an important role in the antitumour defence in cervical carcinoma. This review describes the rationale for the use of immunotherapy as treatment for cervical carcinoma as well as the results of recent developments in tumour immunology and its implications for the clinical use of immunotherapeutical approaches.

Animals↗
PubMed1996

Decreased expression of signal-transducing zeta chain in peripheral T cells and natural killer cells in patients with cervical cancer.

An impaired immune response is frequently observed in patients and experimental animals with advanced cancer. We and others have shown alterations in CD3-associated signal-transducing zeta molecules in tumor-infiltrating T cells and peripheral blood lymphocytes (PBLs) of patients with advanced cancer. By using flow cytometric analysis of permeabilized cells with a monoclonal antibody (TIA-2) that reacts with the cytoplasmic domain of the zeta chain, here we demonstrate a marked decrease (P < 0.01) in the expression of the signal-transducing CD3 zeta chain of PBLs in patients with cervical cancer (n = 22) as compared to PBLs from healthy donors (n = 21). In addition, PBLs isolated from patients (n = 23) with cervical intraepithelial neoplasia (CIN), to a lesser but significant (P < 0. 01) extent, expressed reduced CD3 zeta levels as compared to those from healthy donors. This decreased expression of zeta chains was also observed on CD16(+) natural killer cells in PBLs from patients with cervical cancer. Surface expression of CD3 epsilon on PBLs was also decreased in cervical cancer patients as compared to healthy donors, but not on PBLs from patients with CIN. CD3 zeta chain expression significantly (r = 0.53, P < 0.01) correlated with the ability of the PBLs to produce tumor necrosis factor in response to anti-CD3 stimulation. These findings suggest that alterations of signal-transducing zeta molecules commonly occur in patients with cervical cancer and to a lesser extent with CIN, and that they are associated with reduced cellular functions such as production of tumor necrosis factor.

Antibodies, Monoclonal↗
PubMed1995

Antigen capture and major histocompatibility class II compartments of freshly isolated and cultured human blood dendritic cells.

Dendritic cells (DC) represent potent antigen-presenting cells for the induction of T cell-dependent immune responses. Previous work on antigen uptake and presentation by human DC is based largely on studies of blood DC that have been cultured for various periods of time before analysis. These cultured cells may therefore have undergone a maturation process from precursors that have different capacities for antigen capture and presentation. We have now used immunoelectron microscopy and antigen presentation assays to compare freshly isolated DC (f-DC) and cultured DC (c-DC). f-DC display a round appearance, whereas c-DC display characteristic long processes. c-DC express much more cell surface major histocompatibility complex (MHC) class II than f-DC. The uptake of colloidal gold-labeled bovine serum albumin (BSA), however, is greater in f-DC, as is the presentation of 65-kD heat shock protein to T cell clones. The most striking discovery is that the majority of MHC class II molecules in both f-DC and c-DC occur in intracellular vacuoles with a complex shape (multivesicular and multilaminar). These MHC class II enriched compartments (MIIC) represent the site to which BSA is transported within 30 min. Although MIIC appear as more dense structures with less MHC class II molecules in f-DC than c-DC, the marker characteristics are very similar. The MIIC in both types of DC are acidic, contain invariant chain, and express the recently described HLA-DM molecule that can contribute to antigen presentation. CD19+ peripheral blood B cells have fewer MIIC and surface MHC class II expression than DCs, while monocytes had low levels of MIIC and surface MHC class II. These results demonstrate in dendritic cells the elaborate development of MIIC expressing several of the components that are required for efficient antigen presentation.

Animals↗
PubMed1995

Eradication of large human B cell tumors in nude mice with unconjugated CD20 monoclonal antibodies and interleukin 2.

Since antibody-dependent cellular cytotoxicity is considered an important mechanism by which mAbs may exert their antitumor effects, it seems likely that these antitumor effects can be enhanced by the activation of the appropriate effector cell populations. We have used nude mice xenografted with human Daudi tumor cells as a model to compare the antilymphoma effects of unconjugated CD19 (CLB-CD19) and CD20 (BCA-B20) mAbs (IgG2a subclass) alone or in combination with recombinant human interleukin 2 (rhIL-2) or recombinant mouse granulocyte-macrophage-colony-stimulating factor (rmGM-CSF). Treatment of established tumors with BCA-B20 or rhIL-2 or rmGM-CSF as a single agent, all resulted in highly significant decreases of tumor growth rates, but did not increase the number of complete regressions. The combination of CLB-CD19 or BCA-B20 mAbs with rhIL-2 or rmGM-CSF resulted in larger decreases of growth rates than either of the agents alone. Complete eradication of large Daudi tumors could be achieved when treatment with BCA-B20 mAbs was combined with rhIL-2, but not with the combination of CLB-CD19 mAbs and rhIL-2 nor with the combination of BCA-B20 mAbs and rmGM-CSF. Cured animals kept for 2-3 months after complete regression of the tumors were still tumor free. Regression of tumors was correlated with the infiltration of lymphocytes as well as macrophages into the tumor. This is the first report to show that unconjugated CD20 mAbs are to be preferred over unconjugated CD19 mAbs, and interleukin 2 over GM-CSF in the combinational treatment of large B cell tumors.

Animals↗
PubMed1995

Human CTL epitopes encoded by human papillomavirus type 16 E6 and E7 identified through in vivo and in vitro immunogenicity studies of HLA-A*0201-binding peptides.

Human papillomavirus type 16 (HPV16) is strongly associated with cervical carcinogenesis. The HPV16 E6 and E7 oncoproteins are constitutively expressed in the majority of cervical tumor cells and are, therefore, attractive targets for CTL-mediated immunotherapy. In mice, the outgrowth of a lethal dose of HPV16-induced tumor cells has been prevented by vaccination with a CTL epitope encoded by HPV16 E7, indicating the feasibility of peptide immunization to obtain antitumor CTL responses. In the present study, the immunogenicity of 9 HLA-A*0201-binding peptides encoded by HPV16 E6 and E7 was analyzed in vivo in HLA-A*0201Kb transgenic mice and in vitro in CTL cultures induced from PBMC of HLA-A*0201+ healthy donors. Four peptides with a good binding affinity were immunogenic in HLA-A*0201Kb transgenic mice, and three of them were also highly immunogenic in CTL induction experiments with PBMC of HLA-A*0201+ healthy donors. Human CTL clones specific for these three peptides were capable of lysing the HPV16 E7-containing HLA-A*0201+ cervical carcinoma cell line CaSki. These E7-derived peptides (11-20, YMLDLQPETT; 82-90, LLMGTLGIV; 86-93, TLGIVCPI), therefore, are likely to represent naturally processed human CTL epitopes of HPV16. Additionally, these three HPV16-encoded peptides have the highest affinity of binding to the HLA-A*0201 molecule. In this study, peptides with a lower binding affinity were less immunogenic. Therefore, our data illustrate that the HLA-binding affinity of a peptide has a major impact on its immunogenicity. In conclusion, we have identified immunogenic peptides encoded by HPV16 E6 and E7 that could be used in vaccines for the prevention and treatment of cervical carcinoma.

Amino Acid Sequence↗
PubMed1995

MHC class II compartments and the kinetics of antigen presentation in activated mouse spleen dendritic cells.

MHC class II (MHC-II) molecules bind fragments of exogenous Ags in an intracellular endocytotic compartment. In view of divergent data on the MHC-II distribution in different cell lines, it was of interest to localize MHC-II molecules in a natural and the most potent APC type, the dendritic cell (DC). By using immunogold labeling of ultrathin cryosections of cultured mouse spleen DC, we found that MHC-II molecules were present abundantly at the plasma membrane and in intracellular compartments containing internal membrane vesicles and/or membrane sheets. The majority of these compartments was situated late in the endocytotic route, as demonstrated by the late appearance (after a lag of 30 min) of internalized exogenous tracer. These compartments contained the lysosomal enzymes cathepsin D and beta-hexosaminidase, but lacked the late endosomal marker cation-dependent mannose-6-phosphate receptor. We conclude that most of the intracellular MHC-II molecules in cultured spleen DC reside in a compartment with (pre)lysosomal characteristics, resembling the so-called MHC-II-enriched compartments (MIIC), originally described in B cells. We also investigated whether the presence of MHC-II molecules in endocytotic compartments was related to the kinetics of Ag processing and presentation by these cells. Pulse-chase endocytosis experiments with hen egg lysozyme (HEL) as a model Ag showed that activated spleen DC were able to efficiently process and present this Ag to an HEL-specific T hybridoma cell line. However, presentation started only after a lag of 2 h and was maximal after 6 h. The difference in time between the arrival of Ag in proteolytic endocytotic compartments, in particular MIIC, and effective Ag presentation is discussed in the context of DC maturation.

Animals↗
PubMed1995

Induction and characterization of cytotoxic T-lymphocytes recognizing a mutated p21ras peptide presented by HLA-A*0201.

The ras oncogene is frequently found to be activated in human cancer through point mutations at codons 12, 13 or 61. We explored whether these altered p21ras protein sequences contain peptide sequences that can activate naive CD8+ cytotoxic T lymphocytes (CTL). Several wild-type and mutated p21ras peptides were identified that carry a binding motif for human leukocyte antigen (HLA)-A*0201. Two peptides were found to bind strongly to this allele. CD8+ CTL bulk cultures specifically reacting with one of these peptides could be induced, using processing-defective T2 cells loaded with peptide CLLDILDTAGL as stimulators. The peptide is derived from p21ras, position 51-61, and carries a 61 Gln-->Leu mutation. In contrast, a 9-mer peptide CLLDILDTA corresponding to amino acid sequence 51-59 of wild-type p21ras did not yield reactive CTL cultures. T-cell clones with low affinity for the 11-mer peptide were isolated from CLLDILDTAGL-reactive bulk cultures. These T cells did not lyse melanoma cells transfected with 61-Leu N-ras, although lysis was found when these transfectants were pulsed with the 11-mer peptide. Possibly, T cells of higher affinity may be required to demonstrate processed peptide on the cell surface. The combined experiments suggest that a peptide derived from mutated p21ras can be recognized by HLA class I-restricted CTL, whereas an analogous wild-type p21ras peptide may not be immunogenic.

Alleles↗
PubMed1995

CTL specific for the tyrosinase autoantigen can be induced from healthy donor blood to lyse melanoma cells.

CTL that lyse melanoma cells were previously isolated from several melanoma patients. Such CTL recognize autologous proteins, indicating the occurrence of autoreactive T cells in melanoma patients. We have now raised CTL, using responding T lymphocytes from healthy donor blood, that lysed not only cells incubated with an HLA-A*0201-binding tyrosinase peptide but also melanoma cells endogenously processing and presenting the epitope. Our results suggest that autoreactive CTL precursors are present in healthy donor blood and can be activated in vitro with synthetic peptides presented on appropriate APCs. Therefore, tissue-specific, autoantigen-derived peptides might be useful in immunotherapy of both poorly and nonimmunogenic tumors.

Amino Acid Sequence↗
PubMed1995

An adenovirus type 5 early region 1B-encoded CTL epitope-mediating tumor eradication by CTL clones is down-modulated by an activated ras oncogene.

Mouse embryo cells (C57BL/6, H-2b) transformed by the E1A and E1B genes of adenovirus type 5 (Ad5E1 MEC) are highly immunogenic. Previously, CTL were cloned from mice immunized with Ad5E1 MEC. These CTL clones were capable of tumor eradication in nude mice, and were directed against the Ad5E1A-encoded decapeptide SGPSNTPPEI, presented by the H-2Db MHC molecule. We have now generated Ad5E1 MEC containing a mutated Ad5E1A-encoded epitope. The mutant Ad5E1 MEC induce a strong CTL response when injected into immunocompetent mice. CTL clones generated against mutant Ad5E1-transformed tumor cells recognize an Ad5E1B-encoded epitope (VNIRNCCYI) in the context of H-2Db. Because this epitope is also present on wild-type Ad5E1 MEC, it is concluded that Ad5E1-transformed tumor cells express at least two CTL epitopes. Interestingly, the lysis of Ad5E1 MEC by the Ad5E1B-specific, but not by the Ad5E1A-specific, CTL clones was strongly diminished by the action of the activated ras oncogene. CTL directed against the Ad5E1B-encoded epitope were, like Ad5E1A-specific CTL, able to eradicate large established Ad5E1-induced tumors in B6 nude mice, demonstrating that CTL activity directed against different CTL epitopes expressed by the same tumor can be exploited for immunotherapy of cancer.

Adenovirus E1 Proteins↗
PubMed1995

Major differences in transporter associated with antigen presentation (TAP)-dependent translocation of MHC class I-presentable peptides and the effect of flanking sequences.

The MHC-encoded transporter associated with Ag presentation (TAP) translocates peptides from the cytosol to the ER lumen, where association with MHC class I molecules occurs. The MHC class I/peptide complex is subsequently transported to the cell surface for presentation to CD8+T cells. We studied TAP-dependent translocation of defined MHC class I presentable murine peptides by competition for translocation of a radiolabeled model peptide, to address whether efficient peptide presentation by MHC class I molecules is preceded by equal efficient peptide translocation by TAP. Surprisingly, we observed that four immunodominant viral peptides of 16 peptides tested were very inefficiently transported by TAP. Inefficient translocation could be overcome by substitution of a proline residue present at position 3 in the peptides. Furthermore, addition of natural flanking amino acids directly surrounding a poorly transported peptide could considerably improve translocation by TAP. Our data suggest that some peptides are efficiently transported by TAP in their optimal size for MHC class I binding, whereas other peptides are transported as larger peptide fragments that need further trimming in the ER for MHC class I binding.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

Cytotoxic T lymphocytes raised against a subdominant epitope offered as a synthetic peptide eradicate human papillomavirus type 16-induced tumors.

Previously, we have shown that immunization with human papillomavirus (HPV) type 16-derived cytotoxic T lymphocyte (CTL) epitope E7 49-57 (RAHYNIVTF) renders C57BL/6 mice insensitive to tumors formed by HPV16-transformed cells. In this study, we provide evidence that E7 49-57 is expressed as a subdominant CTL epitope on HPV16-transformed C57BL/6 cells. Using acid peptide elution, it is shown that HPV16-transformed cells express another CTL epitope, besides E7 49-57, which appears to be dominant. We demonstrate that a CTL line raised against the subdominant CTL epitope, offered as synthetic peptide E7 49-57, eradicates established HPV16-induced tumors in mice. Our data show that synthetic peptide-induced CTL can be applied successfully in vivo against (virus-induced) tumor, and emphasize that subdominant CTL epitopes are useful targets for immunotherapy. Furthermore, it is illustrated for the first time that HPV16-specific CTL interfere directly with HPV16-induced tumors.

Animals↗
PubMed1995

Detailed motifs for peptide binding to HLA-A*0201 derived from large random sets of peptides using a cellular binding assay.

Extensive sets of in total about 2000 synthetic peptides were investigated for their binding affinities to HLA-A*0201. Comparisons of the amino acid compositions of binding to nonbinding sets of peptides provided new information concerning the rules for 9-, 10-, and 11-mer peptide binding at the amino acid level. Preferred primary anchors were shown to depend on peptide length, longer peptides being more demanding in this respect. A clear preference exists for certain amino acids at several nonanchor positions. In addition, the presence of particular amino acids at those positions almost completely precludes peptide binding. We found no evidence for preferred anchor pairs. From these results new and detailed HLA-A*0201 peptide-binding motifs for 9-, 10-, and 11-mer peptide binding were deduced. The motifs are in accordance with earlier reports but include new findings, including C as a C-terminal anchor, the importance of D at positions 4 for binding, and the deleterious effect of R at position 5 (in 9-mers). The motifs are presented in such a way that they can be used to predict peptide binding to HLA-A*0201 by computer analysis (see accompanying paper [56]).

Alleles↗
PubMed1995

A computer program for predicting possible cytotoxic T lymphocyte epitopes based on HLA class I peptide-binding motifs.

Vaccination with peptides recognized by antigen-specific CTLs can prevent lethal virus infections and tumor growth. In order to avoid the synthesis and testing of the numerous overlapping peptide of long AA sequences of proteins of interest, we developed a computer program which utilizes the rules, "motifs" which govern how peptides bind to HLA class I molecules, to derive a predicted binding score for each overlapping peptide. Correlations between the predicted and actual binding results to HLA-A*0201 for 100 peptides selected from six early and two late protein sequences of human papillomavirus type 1a revealed an acceptable level (61%) of concordance. The program is very flexible with regard to the input of protein sequences and motif definitions and is able to handle various motif and peptide lengths.

Amino Acid Sequence↗
PubMed1995

An HLA class I peptide-binding assay based on competition for binding to class I molecules on intact human B cells. Identification of conserved HIV-1 polymerase peptides binding to HLA-A*0301.

A peptide-binding assay employing the HLA class I molecules on intact human B cells is described. The peptide antigens are stripped from the HLA class I molecules by mild acid treatment, after which the cells are incubated with a FL-labeled reference peptide together with different concentrations of the peptide of interest. The effectiveness by which the latter peptide competes for binding to the HLA class I molecules is assayed by measuring the amount of HLA-bound FL-labeled reference peptide with FACscan analysis. The assay is easy to perform because there is no need to purify HLA class I molecules, or to transfect cells with HLA class I molecules, and no radioactive label is used. Moreover, large panels of HLA-typed human B-cell lines are available as tools for peptide binding to a vast array of HLA molecules. The binding assay was optimized and validated with peptides of known binding capacity to either HLA-A*0201 or HLA-A*0301. The kinetics of peptide binding in this assay were shown to be comparable to that in assays employing soluble HLA class I molecules. Application of the assay in the search for potential HLA-A*0301 restricted CTL epitopes, derived from HIV-1 polymerase, resulted in the identification of five high-affinity binding peptides.

Amino Acid Sequence↗
PubMed1995

Bone marrow-derived dendritic cells pulsed with synthetic tumour peptides elicit protective and therapeutic antitumour immunity.

Dendritic cells, the most potent 'professional' antigen-presenting cells, hold promise for improving the immunotherapy of cancer. In three different well-characterized tumour models, naive mice injected with bone marrow-derived dendritic cells prepulsed with tumour-associated peptides previously characterized as being recognized by class I major histocompatibility complex-restricted cytotoxic T lymphocytes, developed a specific T-lymphocyte response and were protected against a subsequent lethal tumour challenge. Moreover, in the C3 sarcoma and the 3LL lung carcinoma murine models, treatment of animals bearing established macroscopic tumours (up to 1 cm2 in size) with tumour peptide-pulsed dendritic cells resulted in sustained tumour regression and tumour-free status in more than 80% of cases. These results support the clinical use of tumour peptide-pulsed dendritic cells as components in developing effective cancer vaccines and therapies.

Animals↗