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J Schlom

Publications and source records attributed to J Schlom.

At least 91 records · Page 5Linked to original sources
PubMed1997

In vitro generation of human cytotoxic T lymphocytes specific for peptides derived from prostate-specific antigen.

BACKGROUND: Protein antigens are presented to cytotoxic T lymphocytes as small peptides (approximately 9-10 amino acids long) bound to class I molecules of the major histocompatibility complex. The identification of tumor-associated antigens and specific peptide epitopes (i.e., antigenic determinants) may be useful in the development of anticancer vaccines. The generation of a cytotoxic T-cell response to one peptide epitope (amino acids 146-154) of human prostate-specific antigen (PSA) has been reported. PURPOSE: Our aim was to identify novel PSA peptides capable of eliciting specific cytotoxic T-cell responses. METHODS: Candidate peptides were identified on the basis of the following criteria: 1) they contained consensus amino acid motifs for binding to HLA-A2, which is the most common type of class I molecule; 2) they lacked strong homology with PSA-related kallikrein proteins; and 3) they were capable of stabilizing HLA-A2 class I molecules on the surface of human T2 (transport deletion mutant) cells, which are defective in antigen presentation. T-cell lines capable of killing (i.e., lysing) T2 target cells that had been pulsed with specific PSA peptides were generated from three different males (two disease-free individuals and one patient with prostate cancer) by incubating peripheral blood mononuclear cells with the peptides and interleukin 2. Specific cell lysis was monitored by the release of radioactivity from target cells that had been labeled with [111In]oxyquinoline. RESULTS: Two novel PSA peptides capable of eliciting cytotoxic T-cell responses were identified; these peptides were designated PSA-1 (amino acids 41-150) and PSA-3 (amino acids 154-163). Four different cytotoxic T-cell lines were generated in response to these peptides-three against PSA-3 and one against PSA-1. Specific lysis of peptide-pulsed T2 cells by the T-cell lines was blocked by the addition of a monoclonal antibody directed against class I molecules. The T-cell lines were also capable of lysing PSA-positive, HLA-A2-positive LNCaP cells (human prostate carcinoma cells). The specificity of LNCaP cell lysis was shown by the following: 1) the inability of added human K562 (chronic myelogenous leukemia) cells to inhibit it, 2) the ability of added anti-HLA-A2 antibodies to block it, and 3) the inability of the T-cell lines to induce substantial lysis of PSA-negative, HLA-A2-positive human cancer cells. IMPLICATIONS: Our studies form a rational basis for the use of PSA peptides or recombinant vectors encoding PSA in the development of anticancer vaccine immunotherapy protocols for patients with prostate cancer.

Cell Line↗
PubMed1997

Intraperitoneal radioimmunotherapy of ovarian cancer with 177Lu-CC49: a phase I/II study.

BACKGROUND: Twenty-seven ovarian cancer patients who failed chemotherapy entered a phase I/II trial of intraperitoneal 177Lu-CC49 antibody. METHODS: Patients had disease confined to the abdominal cavity +/- retroperitoneal lymph nodes, adequate organ function, and no previous radiation. RESULTS: The most common side effects were delayed, transient arthralgia (10/27) and marrow suppression with 1.665 GBq/m2 (45 mCi/m2), which was considered the maximum tolerated dose. One of thirteen patients with gross disease had >50% tumor reduction after therapy, whereas most others with gross disease progressed (one went off study with stable disease at 11 weeks). Seven of nine patients with <1-cm nodules progressed in < or =21 months, and two of nine remain without evidence of disease at 4 to 5 months. Of patients with microscopic or occult disease, one relapsed at 10 months and four of five remain without evidence of disease at >6 to 35 months. CONCLUSIONS: Marrow suppression was the dose-limiting toxic effect of intraperitoneal immunotherapy with 177Lu-CC49. Antitumor effects were noted against chemotherapy-resistant ovarian cancer, even at lower dose levels, and resulted in prolonged disease-free survival of most patients with microscopic disease. This form of treatment deserves further study.

Adult↗
PubMed1997

A CDR-grafted (humanized) domain-deleted antitumor antibody.

While several murine monoclonal antibodies (MAbs) directed against carcinoma associated antigens have shown excellent tumor targeting properties in clinical trials, the use of radiolabeled MAbs for both diagnostic and therapeutic applications has been hindered by two factors: (a) the induction of host anti-immunoglobulin (Ig) responses and (b) slow plasma clearance of unbound radiolabeled MAb, resulting in bone marrow toxicity for therapeutic application, and long intervals between MAb administration and tumor detection for diagnostic applications. This report describes the development of the first recombinant Ig with properties designed to reduce or eliminate both of the above problems: a complementarity determining region (CDR)-grafted humanized (Hu) MAb with a CH2 domain deletion (delta CH2). The MAb chosen for engineering was CC49, which is directed against a pancarcinoma antigen designated TAG-72 that is expressed on the majority of colorectal, gastric, breast, ovarian, prostate, pancreatic and lung carcinomas. When characterized for antigen binding in solid phase competition radioimmunoassays, the HuCC49 delta CH2 MAb completely inhibited the binding of murine (mu) CC49 and HuCC49 for TAG-72. The relative affinity constants (Ka) of MAbs HuCC49 delta CH2, HuCC49 and muCC49 were 5.1 x 10(-9), 2.1 x 10(-9) and 2.3 x 10(-9), respectively. The plasma clearance of 131I-HuCC49 delta CH2 was significantly faster than that of intact 125I-HuCC49 after either i.v. or i.p. administration in athymic mice (p(2)0.05). Biodistribution studies in athymic mice bearing human colon carcinoma xenografts after i.v. or i.p. administration of 131I-HuCC49 delta CH2 and 125I-HuCC49 demonstrated the efficient tumor localization and substantially lower percent of the injected dose (%ID/g) of the HuCC49 delta CH2 in normal tissues. This is reflected in the significantly higher radiolocalization indices (%ID/g in tumor divided by %ID/g in normal tissue) observed with the HuCC49 delta CH2 for most normal tissues tested (p(2)0.05). The differential between the rate of plasma clearance of HuCC49 delta CH2 and HuCC49 was even more pronounced in SCID mice, which have been shown to be an appropriate model to study the metabolism of human IgG. These studies thus describe the development of a recombinant Ig molecule which, for the first time, combines 1) the properties of more rapid blood clearance than an intact humanized Ig molecule--without loss of antigen binding affinity--and 2) reduced potential for eliciting a human anti-murine antibody (HAMA) response in patients. These studies also demonstrate the potential utility of HuCC49 delta CH2 for i.p. as well as i.v. radioimmunodiagnosis and radioimmunotherapy in patients with TAG-72 positive tumors.

Animals↗
PubMed1997

Correlation of toxicity with treatment parameters for 131I-CC49 radioimmunotherapy in three phase II clinical trials.

Analyses were performed on 40 patients with TAG-72 expressing metastatic cancer who were entered into three phase II clinical trials. The dose selected was the maximum tolerated dose in phase I studies. Patients all had unresectable metastatic colon or prostate cancer and had recovered from prior therapies. Patients in trials #1 and #2 received 75 mCi/m2 131I-CC49 antibody whereas those in trial #3 received a total of 75 mCi/m2 with equal amounts of 131I-CC49 and 131I-COL-1. The three trials have resulted in a reproducible degree of reversible marrow suppression; 72.5% of patients experienced moderate or severe toxicity. Comparisons were made between demographic, clinical and pharmacokinetical variables and the grade of WBC toxicity, platelet toxicity and the sum of the two as total toxicity. Whole body radiation dose had a statistically significant relationship with platelet toxicity (r = 0.38, p = 0.015) and total toxicity (r = 0.34, p = 0.035). The bone marrow radiation dose is significantly related to all toxicity indicators with correlation coefficients with WBC and platelet toxicities of 0.47 (p = 0.002) and 0.34 (p = 0.033), respectively. Plasma half-life had the strongest correlation with WBC toxicity and combined toxicities. Multivariate models were developed to help describe the simultaneous effect of these variables on toxicity. The results show that the MTD dose was safely given to patients who varied in age, disease burden and degree of marrow compromise. This supports the contention that a fixed dose of radiolabeled antibody per body mass or m2 can be given to a diverse group of non-lymphoma patients with a predictable toxicity range.

Adult↗
PubMed1997

Therapeutic antitumor response after immunization with an admixture of recombinant vaccinia viruses expressing a modified MUC1 gene and the murine T-cell costimulatory molecule B7.

Tumor-associated antigens have considerable promise not only as diagnostic or prognostic markers but also as targets for active or passive immunotherapy. DF3/MUC1 is a tumor-associated antigen that is overexpressed with an abnormal glycosylation pattern in breast, ovarian, lung, and pancreatic cancers. The major extracellular portion of MUC1 is composed of tandem repeat units of 20 amino acids. Recombinant vaccinia viruses encoding mucin molecules have been constructed by several groups. However, these recombinants have met with limited success in protecting animals from MUC1-expressing tumors because of the vaccinia genome being subject to high-frequency homologous recombination, therefore being unstable in expression of the tandem repeats. In light of these studies, two concurrent strategies were used to improve immune responses to MUC1: a recombinant vaccinia virus was constructed containing a modified "mini" MUC1 gene containing only 10 tandem repeat sequences to minimize vaccinia-mediated rearrangement (designated rV-MUC1); and an admixture was used containing rV-MUC1 and a recombinant vaccinia virus containing the gene for the murine T-cell costimulatory molecule B7-1 (rV-B7). The rV-MUC1 gene product maintained a consistent molecular weight throughout several passages, indicating stability of the inserted gene. Mice inoculated with rV-MUC1 demonstrated MUC1-specific cytolytic responses that were further enhanced by admixture with rV-B7. In a MUC1-expressing pulmonary metastases prevention model, mice inoculated two times with rV-MUC1 were protected from the establishment of metastases. No additive effect on antitumor immunity (> 90% with rV-MUC1 alone) was observed in mice primed with an admixture of rV-MUC1 and rV-B7 and boosted with rV-MUC1. When rV-MUC1 was used to treat established MUC1 positive metastases, however, three administrations of rV-MUC1 were not sufficient to confer antitumor effects. In contrast, when tumor-bearing mice were primed with an admixture of rV-MUC1 and rV-B7, followed by two boosts with rV-MUC1, there was a significant reduction in pulmonary metastases (p = < 0.0001), which correlated to 100% survival. Coexpression of the B7 molecule, although not necessary for the induction of an immune response of sufficient magnitude to prevent MUC1 tumors, was thus essential in a treatment setting.

Animals↗
PubMed1997

Adoptive immunotherapy as an in vivo model to explore antitumor mechanisms induced by a recombinant anticancer vaccine.

We have described previously the construction, generation, and in vivo biologic consequences of a recombinant vaccinia virus containing the human CEA gene (rV-CEA) in an experimental murine colon carcinoma model. Immunization of C57BL/6 mice with rV-CEA led to antigen-specific inhibition of tumor growth in both prophylactic and therapeutic settings. Although such antitumor effects were correlated with the induction of CEA-specific T-cell responses, their exact contribution in the tumor rejection mechanism remained unclear. In this study, we examined the mechanism of action of rV-CEA, with emphasis on definition of the immune cells important for such antitumor effects. To that end, a cellular adoptive transfer model was established in vivo, which allowed specific functional analysis of donor-derived immune cells in naive, sublethally irradiated, tumor-bearing recipients. Splenocytes from rV-CEA-immunized donors expressed strong antitumor activity in such tumor-bearing recipients, whereas nonimmune donor cells did not. Depletion of immune T cells before cellular transfer abolished the antitumor response. Moreover, depletion of CD8+ T cells before transfer resulted in the loss of antitumor activity, despite the presence of CD4+ T cells. In contrast, antitumor activity was demonstrable with CD8-containing, CD4-depleted effectors, although it was not as effective as with both T-cell subpopulations combined. Finally, in beta 2-microglobulin/CD8+ T-cell-deficient mice, rV-CEA immunization exerted only partial antitumor protection, compared with the immune-competent controls. Overall, we demonstrated that (a) antitumor activity induced by rV-CEA was essentially mediated by CD8+ effectors; and (b) the combination of both CD8+ and CD4+ lymphocytes led to maximal antitumor therapeutic effects, suggesting an important helper or immunoregulatory contribution of the CD4+ subset. Thus, adoptive cellular transfer strategies may have implications for both the study of recombinant anticancer vaccines and the development of potential clinical applications for cancer immunotherapy.

Adenocarcinoma↗
PubMed1997

Immunization with a syngeneic tumor infected with recombinant vaccinia virus expressing granulocyte-macrophage colony-stimulating factor (GM-CSF) induces tumor regression and long-lasting systemic immunity.

A recombinant vaccinia virus encoding the gene for granulocyte-macrophage colony-stimulating factor (rV-GM-CSF) was used to infect the poorly immunogenic murine colon adenocarcinoma cell line, MC-38. Infection of MC-38 tumor cells with rV-GM-CSF completely suppressed the growth of the MC-38 primary tumors, whereas progressively growing tumors were formed in mice injected with MC-38 cells infected with wild type V-Wyeth. Irradiation of the recipient B6 mice before implantation of rV-GM-CSF-infected tumor cells resulted in the development of progressively growing tumors. Moreover, in vivo T-cell depletion studies revealed that growth suppression of the rV-GM-CSF-infected tumor cells was dependent on the presence of both CD4+ and CD8+ T-cell subsets. Subsequent studies established that this immunity was long-lasting and antigen specific, as demonstrated by the protection of rV-GM-CSF-immunized mice from MC-38 tumor challenge but not from challenge with another syngeneic tumor cell type. No such effects were observed when MC-38 tumor cells were infected with recombinant vaccinia viruses expressing interleukin (IL)-2 or IL-6. The results demonstrate that paracrine release of biologically active murine GM-CSF by tumor cells infected with rV-GM-CSF enhances the intrinsic immunogenicity of a poorly immunogenic murine tumor. Presumably the augmentation of tumor immunogenicity induces an antigen-specific T-cell-dependent antitumor response that prevents the formation of primary tumors and protects mice from tumor challenge. Thus in this experimental model, GM-CSF functions as a highly effective vaccine adjuvant.

Adenocarcinoma↗
PubMed1997

Phenotypic stability of a cytotoxic T-cell line directed against an immunodominant epitope of human carcinoembryonic antigen.

CTL lines have now been generated against defined peptides of a range of human tumor-associated antigens (TAAs). One of the potential uses of these epitope-specific CTLs is in adoptive transfer immunotherapy. This is a modality, however, that will require long-term in vitro culture of CTLs. To date, little has been reported concerning the phenotypic stability of human epitope-specific CTLs as a consequence of long-term in vitro propagation via peptide stimulation. We report here the serial phenotypic characterization of a CTL line directed against an immunodominant epitope (YLSGANLNL, designated CAP-1) of human carcinoembryonic antigen (CEA). This CTL line was derived from peripheral blood mononuclear cells of a patient with metastatic carcinoma who had been treated with a recombinant CEA-vaccinia vaccine in a Phase I trial; the CTLs were analyzed through 20 in vitro cycle passages of stimulation with CAP-1 peptide and interleukin 2 in the presence of autologous antigen-presenting cells. The CTL line was shown to be phenotypically stable in terms of high levels of cytokine (IFN-gamma, tumor necrosis factor, and granulocyte-macrophage colony-stimulating factor) production, expression of homing-adhesion molecules, ability to lyse peptide-pulsed targets, and ability to lyse human carcinoma cells endogenously expressing CEA in a MHC-restricted manner. Vbeta T-cell receptor gene usage was also analyzed. These studies thus present a rationale for the use of long-term cultured epitope-specific human CTLs, directed against a human TAA for potential adoptive transfer immunotherapy protocols.

Antigens, CD↗
PubMed1997

Effect of recombinant alpha-interferon on pharmacokinetics, biodistribution, toxicity, and efficacy of 131I-labeled monoclonal antibody CC49 in breast cancer: a phase II trial.

Preclinical studies have demonstrated that recombinant IFN-alpha (rIFN-alpha) can enhance the tumor associated glycoprotein 72 (TAG-72) on tumors. To determine whether rIFN-alpha could enhance TAG-72 expression in vivo in patients, 15 women with breast cancer were randomized to receive daily injections of rIFN-alpha (3 x 10(6) units/m2 for 14 days) beginning on day 1 (group 1 = 7 patients) or on day 6 (group 2 = 8 patients). On day 3, all patients received a 10-20-mCi tracer dose of 131I-CC49, a high-affinity murine monoclonal antibody reactive against TAG-72, followed by a therapy dose of 60-75 mCi/m2 of 131I-CC49 on day 6. Whole body and single-photon emission computed tomography scans along with whole blood pharmacokinetics were performed following tracer and treatment phases. Hematological toxicity was considerable; reversible grade 3-4 neutropenia and thrombocytopenia was observed in 12 of 15 patients. Twelve of 14 patients tested developed human antimouse antibodies 3-6 weeks after treatment. For group 1 patients, whole blood residence time increased significantly between that predicted from the tracer doses and therapy doses (42.6 +/- 4.7 versus 51.5 +/- 4.8 h, respectively; P < 0.01). The calculated radiation absorbed dose to red marrow from therapy compared to tracer activity was also significantly higher for this group (1.25 +/- 0.35 versus 1. 07 +/- 0.26 cGy/mCi; P < 0.05). Treatment with rIFN-alpha was found to enhance TAG-72 expression in tumors from patients receiving rIFN-alpha (group 1) by 46 +/- 19% (P < 0.05) compared to only 1.3 +/- 0.95% in patients not initially receiving IFN (group 2). The uptake of CC49 in tumors was also significantly increased in rIFN-alpha-treated patients. One partial and two minor tumor responses were seen. In summary, rIFN-alpha treatment altered the pharmacokinetics and tumor uptake of 131I-CC49 in patients at the expense of increased toxicity.

Adult↗
PubMed1996

Comparison of linear and branched peptide forms (MAPs) in the induction of T helper responses to point-mutated ras immunogens.

The utility of multiple antigenic peptides (MAPs) for the induction of antibody and cellular immune responses in animal models has been demonstrated for a variety of peptide epitopes involved in human disease. However, little is known about immune responses to MAPs constructed with antigenic tumor epitopes, nor has peptide specificity in branched forms been addressed. A potentially important advantage of the MAP system over linear peptide immunogens for clinical applications is elimination of the need for a protein carrier with its associated toxicity and immunogenicity. Here, we examined cellular immune responses following in vivo administration of MAPs incorporating a 13-mer T helper epitope from point-mutated ras p21 (ras V12) and compared the potency of the responses to that of the linear peptide. The Gly --> Val mutation in position 12, which is associated with a range of human carcinomas, represents a useful system for evaluating the specificity of the immune response. In initial studies with the point-mutated linear peptide epitope, optimal in vitro proliferation responses were obtained following sc administration of the peptide in a squalane-containing adjuvant formulation. Comparative immunization studies using point-mutated MAPs bearing two, four, or eight branches were administered either in saline or in adjuvant. These studies showed that adjuvant was required for the induction of cellular immune responses using both linear and all three forms of branched peptides. Moreover, there was no apparent advantage of using any of the MAPs vs linear peptide when equivalent mass amounts were administered, i.e., the intensity of the immune response was no greater using any of the branched structures compared to the linear form. Specificity of the in vivo responses for both the linear and the MAP immunogens was demonstrated by the higher stimulation indices observed in vitro in the presence of the mutant ras V12 vs the normal ras G12 linear peptide. No apparent cellular immune response to the MAP core structure itself was observed. However, a nonspecific response to the two-branched MAP2G12 structure was observed in some assays, the nature of which is unknown at this time. This work represents the first reported investigation of a cellular immune response using MAP immunogens incorporating a tumor-specific peptide epitope and demonstrates that linear peptides are as efficient as three different MAP structures in the generation of specific T cell responses.

Adjuvants, Immunologic↗
PubMed1996

Correlation between tumor-associated glycoprotein 72 mucin levels in tumor and serum of colorectal patients as measured by the quantitative CA 72-4 immunoassay.

Colorectal tissue biopsies were obtained from 110 patients diagnosed with primary colorectal carcinoma (tumor and normal colonic mucosa samples), 20 patients diagnosed with benign colorectal disease, and 31 healthy donors. The level of expression of tumor-associated glycoprotein 72 (TAG-72) was quantitatively measured in each sample using a double-determinant RIA with monoclonal antibodies B72.3 and CC49 and detecting the sialyl-Tn epitope; this assay was termed CA 72-4. Statistical analysis revealed a significant (approximately 10-fold) increase of TAG-72 expression in the colon tumor biopsies when compared with the expression in normal colonic mucosa from the same patients. A regression analysis revealed a significant correlation (r = 0.459; P < 0.001) between TAG-72 levels measured in biopsies from the tumor lesions and those found in the corresponding normal colonic mucosa. Furthermore, regression analysis showed a significant positive correlation between TAG-72 levels in the tumors and sera of the same patients (r = 0.491; P < 0.001). TAG-72 levels in normal colonic mucosa from healthy donors and patients diagnosed with colorectal cancer were compared. TAG-72 expression was 5-fold higher in the normal mucosa from the colorectal carcinoma patients. No relationship between TAG-72 tumor tissue content and stage of disease was found. Moreover, the correlation between TAG-72 distribution and degree of tumor differentiation observed (P < 0.05) was not any more evident when mucinous carcinomas were excluded. Finally, the results provide further evidence that TAG-72 may be considered an important early marker for colorectal cancer and/or other dysplastic colonic diseases. The statistical correlation between TAG-72 levels in tumors and circulating TAG-72 indicates that patients with elevated levels of serum TAG-72, as measured by the CA 72-4 assay, would be most suited for diagnostic and/or therapeutic intervention with the anti-TAG-72 monoclonal antibodies B72.3 or CC49 or vaccine trials using the sialyl-Tn epitope.

Adult↗
PubMed1996

Construction of phosphorylatable monoclonal antibody to a tumor-associated antigen.

A phosphorylation site was introduced into chimeric monoclonal antibody B72.3 (MAb-chB72.3) by site-specific mutation of the coding sequence. The phosphorylation site for the cAMP-dependent protein kinase was positioned at the carboxyl terminus of the heavy-chain constant region of MAb-chB72.3. The resultant modified MAb-chB72.3-P was expressed in 293 cells and purified. The MAb-chB72.3-P protein was phosphorylated by the catalytic subunit of cAMP-dependent protein kinase with [gamma-32P]ATP to high radiospecific activity. The 32P-labeled MAb-chB72.3-P protein bound to cells expressing the tumor-associated glycoprotein 72 antigen. The introduction of phosphorylation sites into MAbs provides a new type of MAb for the diagnosis and treatment of cancers.

Amino Acid Sequence↗
PubMed1996

Improved immunotherapy of a recombinant carcinoembryonic antigen vaccinia vaccine when given in combination with interleukin-2.

Interleukin-2 (IL-2) has been an effective immune modulator in several active-specific immunotherapy experimental protocols using either viral or oncolysate-based vaccines. In this report, data indicate that IL-2 administration can appreciably augment the therapeutic effect of a single immunization of a recombinant vaccinia virus-carcinoembryonic antigen (rV-CEA) vaccine using a CEA-expressing syngeneic experimental murine model system. A single rV-CEA immunization of C57BL/6 mice bearing palpable CEA-positive colon adenocarcinoma tumors results in complete tumor regression in approximately 20% of the mice. The addition of a course of low-dose IL-2 results in complete tumor regression in 60-70% of the mice. Moreover, the combination of rV-CEA and IL-2 induces systemic immunity, which protects those tumor-free mice from subsequent rechallenge with the CEA-expressing tumor cells. No such tumor regression or protection was observed in those mice immunized with the wild-type vaccinia vaccine (V-Wyeth) alone or with IL-2 administration alone. Cellular immune assays revealed that the addition of IL-2 to rV-CEA immunization significantly increased the CEA-specific T-cell proliferative responses as well as the cytolytic T-cell responses when compared with rV-CEA immunization alone. The enhanced CEA-specific immune response, coupled with the improved experimental therapeutic outcome following IL-2 administration, suggests that treatment with that cytokine may effectively substitute for multiple rV-CEA immunizations in active-specific immunotherapy clinical protocols directed at CEA-expressing tumors.

Adjuvants, Immunologic↗
PubMed1996

Identification of overlapping epitopes in mutant ras oncogene peptides that activate CD4+ and CD8+ T cell responses.

Mutant ras p21 proteins contain sequences which distinguish them from normal endogenous ras and, thus, may represent unique epitopes for T cell recognition of antigen bearing tumor cells. Here, we examined the capacity of a mutant K-ras 9-mer peptide to induce in vivo CD8+ cytotoxic T lymphocytes (CTL). The peptide chosen reflected positions 4-12 of the point-mutated sequence of the K-ras oncogene encoding the Gly to Val substitution at codon 12. The overall rationale for selecting this particular 9-mer sequence was threefold: the mutant peptide contained a putative major histocompatibility complex (MHC) class I consensus anchor motif for murine H-2Kd; specific binding to MHC class I may then create an immunogenic complex for the induction of anti-ras CD8+ CTL; and finally, the mutant sequence overlapped with a newly characterized anti-ras CD4+ T helper type 1 epitope, which may have implications for the coordination and activation of both anti-ras immune mechanisms against the same target cell antigenic determinant. A functional interaction with H-2Kd was demonstrated with the mutant ras4-12(V12) peptide, but not the normal ras4-12(G12) peptide, which specifically inhibited an H-2Kd-restricted, anti-nucleoprotein NP147-155 CTL response in a dose-dependent fashion. An anti-ras CD8+ T cell line was then established from immune splenocytes of BALB/c (H-2d) mice injected with ras4-12 (V12) in adjuvant, which mediated peptide-specific lysis of syngeneic P815 tumor targets. Cytotoxicity was restricted by H-2Kd and strongly specific for the mutant ras peptide. Importantly, these anti-ras CTL specifically lysed a syngeneic tumor line (i.e. A20 lymphoma) transduced with the corresponding point-mutated ras oncogene, suggesting T cell receptor recognition of endogenously derived antigen. Overall, these data demonstrated that mutant ras p21 at codon 12(Gly-->Val) contained a peptide sequence which exhibited specific functional binding to a murine MHC class I molecule; the ability of the mutant, but not the normal sequence to bind selectively to murine MHC class I likely reflected the generation of a C-terminal anchor residue; and the ras4-12(V12) peptide was immunogenic for the production of antigen-specific CD8+ CTL, which lysed in vitro a syngeneic tumor cell line harboring the mutant K-ras oncogene.

Amino Acid Sequence↗
PubMed1996

Strategies for the development of recombinant vaccines for the immunotherapy of breast cancer.

The development of recombinant vaccines for specific immunotherapy of carcinoma represents a novel approach for the treatment of breast cancer and other tumor types. This article reviews the various parameters that should be considered in the development of recombinant vaccines. Several breast cancer associated antigens are also discussed which may provide potential target molecules. The human carcinoembryonic antigen (CEA), which is expressed on approximately 50% of breast cancers, represents one such target for immunotherapy. To enhance the immunogenicity of this antigen, a recombinant CEA-vaccinia vaccine, designated rV-CEA, was produced. To study the effects of this vaccine in an animal model, a murine colon carcinoma cell line was transduced with CEA and transplanted into immunocompetent mice for protection and therapy studies. Pre-clinical toxicity studies were also conducted in non-human primates. The results of these studies showed the rV-CEA vaccine to be immunogenic and safe in both rodents and primates, and to elicit good anti-tumor responses in the rodent model. In a Phase I clinical trial in metastatic breast, lung, and colorectal cancer patients involving three immunizations of rV-CEA, at three dose levels, enhancement of T-cell and antibody responses to vaccinia virus proteins were observed with no toxicity. Specific T-cell responses were studied via stimulation of peripheral blood lymphocytes with specific peptide epitopes from the CEA molecule. These studies demonstrated clear cut differences in establishment of T-cell lines pre- versus post-immunization. The T-cell lines were shown to be CD8+ and/or CD4+/CD8+, to lyse EBV transformed B-cells transduced with the CEA gene, and to lyse CEA positive carcinoma cells in a HLA restricted manner. Thus, in a Phase I clinical trial the rV-CEA vaccine has been shown to stimulate a CTL response specific for CEA defined epitopes in cancer patients.

Animals↗
PubMed1996

Phase I trial of iodine 131-labeled COL-1 in patients with gastrointestinal malignancies: influence of serum carcinoembryonic antigen and tumor bulk on pharmacokinetics.

PURPOSE: COL-1 is a high-affinity murine monoclonal antibody (MAb) specific for carcinoembryonic antigen (CEA). A phase I trial was conducted in which a uniform quantity of antibody labeled with escalating doses of iodine 131 (131I) was administered to patients with advanced gastrointestinal (GI) malignancies to evaluate tolerance and pharmacokinetics. PATIENTS AND METHODS: Eighteen patients with advanced, assessable GI malignancies (16 colon, one pancreas, and one gastric) previously treated with conventional chemotherapy (but no pelvic radiation) received 20 mg of COL-1 labeled with 131I, with doses from 10 mCi/m2 to 75 mCi/m2. In this cohort, the baseline serum CEA level ranged from 6 to 2,739 ng/mL (mean +/- SD, 500 +/- 639). RESULTS: Nuclear imaging detected at least one tumor site in all 18 patients; 82% of all tumor involved organs were positive and 58% of all lesions > or = 1.0 cm were detected. Immune complexes were detected in 89% of patients 5 minutes after completion of infusion, and levels correlated with CEA levels (r = .71). Elevated CEA (> 500 ng/mL) and tumor bulk (total tumor area > 150 cm2) correlated directly with clearance of serum radioactivity and inversely with serum half-life and cumulative serum radioactivity parameters. Nonhematologic toxicity was mild and non-dose-limiting. Hematologic toxicity, particularly thrombocytopenia, was both dose-related and dose-limiting. The maximal-tolerated dose is 65 mCi/m2. The correlation between dose (millicuries per square meter) and thrombocytopenia was made stronger, by accounting for either variation in pharmacokinetics, or variation in serum CEA and tumor bulk. CONCLUSION: 131I-COL-1 is well tolerated, except for hematologic toxicity. These data suggest that patients with highly elevated circulating CEA levels and/or increased tumor bulk may clear 131I-labeled COL-1 more rapidly from the circulation and experience less myelosuppression.

Antibodies, Monoclonal↗
PubMed1996

Systemic administration of recombinant interferon alfa in carcinoma patients upregulates the expression of the carcinoma-associated antigens tumor-associated glycoprotein-72 and carcinoembryonic antigen.

PURPOSE: The ability of interferons (IFNs) to enhance tumor-associated antigen expression may be an important approach to enhance the efficacy of some monoclonal antibody (MAb)-based protocols for tumor diagnosis and/or therapy. The present study was designed to determine whether systemic IFN alpha-2a administration (via the intramuscular [IM] route) could upregulate the expression of tumor-associated glycoprotein-72 (TAG-72) and/or carcinoembryonic antigen (CEA) at histologically confirmed sites of carcinoma. PATIENTS AND METHODS: Eighteen patients diagnosed with gastrointestinal (GI) carcinoma received systemic IFN alpha-2a according to four dose schedules. In cohorts I and II, patients received two injections of 3 or 6 x 10(6) U IFN alpha-2a per injection, respectively. Patients in cohorts III and IV received the same doses of IFN alpha-2a, 3 and 6 x 10(6) U, respectively, but three injections were given. Tumor and normal colonic mucosa biopsies were obtained from each patient by endoscopy before IFN alpha-2a and after IFN alpha-2a at surgery. The levels of TAG-72 and CEA expression were measured by (1) immunohistochemistry and reported as percent antigen-positive tumor cells, as well as the relative staining intensity, and (2) a quantitative radioimmunoassay. RESULTS: TAG-72 and CEA levels were consistently increased in tumor biopsies taken from patients in cohorts III and IV. For example, of 10 patients treated in cohorts III and IV, eight had enhanced TAG-72 expression when measured either as percentage TAG-72-positive tumor cells or as an increased MAb staining intensity following IFN alpha-2a. CEA expression in tumor biopsies from seven of 10 patients in cohorts III and IV was also elevated following IFN alpha-2a treatment. Quantitative analysis of TAG-72 and CEA levels in tumor biopsies confirmed higher tumor antigen levels following IFN alpha-2a administration. No such increases in TAG-72 or CEA levels were observed in tumor samples taken from patients in cohorts I and II. CEA or TAG-72 expression in samples of histologically confirmed normal colonic mucosa showed little or no change after IFN alpha-2a treatment. CONCLUSION: Systemic IFN alpha-2a administration can upregulate TAG-72 and CEA expression at distal tumor sites, which may play an important role in immunodiagnosis and therapy.

Adenocarcinoma↗