[Magnetic-resonance-guided radiotherapy : The beginning of a new era in radiation oncology?] / Magnetresonanzgeführte Strahlentherapie : Beginn einer neuen Ära in der Radioonkologie?

CLINICAL ISSUE: Image-guided radiotherapy (IGRT) using X­rays and cone-beam computed tomography (CT) has fostered precision radiotherapy. However, inter- and intrafractional variations of target volume position and organs at risk still limit target volume dose and sparing of radiosensitive organs at risk. METHODOLOGICAL INNOVATIONS: Hybrid machines directly combining linear accelerators and magnetic resonance (MR) imaging allow for live imaging during radiotherapy. PERFORMANCE: Besides highly improved soft tissue contrast, MR-linacs enable online, on-table adaptive radiotherapy. Thus, adaptation of the treatment plan to the anatomy of the day, dose escalation and superior sparing of organs at risk become possible. ACHIEVEMENTS: This article summarizes the underlying intention for the development of MR-guided radiotherapy, technical innovations and challenges as well as the current state-of-the-art. Potential clinical benefits and future developments are discussed. PRACTICAL RECOMMENDATIONS: Increasing availability of MR imaging at linear accelerators calls for the ability to review and interpret MR images. Therefore, close collaborations of diagnostic radiologists and radiation oncologists are mandatory to foster this fascinating technique.

Stereotactic ultrahypofractionated MR-guided radiotherapy for localized prostate cancer - Acute toxicity and patient-reported outcomes in the prospective, multicenter SMILE phase II trial.

Background: Due to superior image quality and daily adaptive planning, MR-guided stereotactic body radiation therapy (MRgSBRT) has the potential to further widen the therapeutic window in radiotherapy of localized prostate cancer. This study reports on acute toxicity rates and patient-reported outcomes after MR-guided adaptive ultrahypofractionated radiotherapy for localized prostate cancer within the prospective, multicenter phase II SMILE trial. Materials and methods: A total of 69 patients with localized prostate cancer underwent MRgSBRT with daily online plan adaptation. Inclusion criteria comprised a tumor stage ≤ T3a, serum PSA value ≤ 20 ng/ml, ISUP Grade group ≤ 4. A dose of 37.5 Gy was prescribed to the PTV in five fractions on alternating days with an optional simultaneous boost of 40 Gy to the dominant intraprostatic lesion defined by multiparametric MRI. Acute genitourinary (GU-) and gastrointestinal (GI-) toxicity, as defined by CTCAE v. 5.0 and RTOG as well as patient-reported outcomes according to EORTC QLQ-C30 and -PR25 scores were analyzed at completion of radiotherapy, 6 and 12 weeks after radiotherapy and compared to baseline symptoms. Results: There were no toxicity-related treatment discontinuations. At the 12-week follow-up visit, no grade 3 + toxicities were reported according to CTCAE. Up until the 12-week visit, in total 16 patients (23 %) experienced a grade 2 GU or GI toxicity. Toxicity rates peaked at the end of radiation therapy and subsided within the 12-week follow-up period. At the 12-week follow-up visit, no residual grade 2 GU toxicities were reported and 1 patient (1 %) had residual grade 2 enteritic symptoms. With exception to a significant improvement in the emotional functioning score following MRgSBRT, no clinically meaningful changes in the global health status nor in relevant subscores were reported. Conclusion: Daily online-adaptive MRgSBRT for localized prostate cancer resulted in an excellent overall toxicity profile without any major negative impact on quality of life.

MR-guided adaptive versus ITV-based stereotactic body radiotherapy for hepatic metastases (MAESTRO): a randomized controlled phase II trial.

BACKGROUND: Stereotactic body radiotherapy (SBRT) is an established local treatment method for patients with hepatic oligometastasis or oligoprogression. Liver metastases often occur in close proximity to radiosensitive organs at risk (OARs). This limits the possibility to apply sufficiently high doses needed for optimal local control. Online MR-guided radiotherapy (oMRgRT) is expected to hold potential to improve hepatic SBRT by offering superior soft-tissue contrast for enhanced target identification as well as the benefit of gating and daily real-time adaptive treatment. The MAESTRO trial therefore aims to assess the potential advantages of adaptive, gated MR-guided SBRT compared to conventional SBRT at a standard linac using an ITV (internal target volume) approach. METHODS: This trial is conducted as a prospective, randomized, three-armed phase II study in 82 patients with hepatic metastases (solid malignant tumor, 1-3 hepatic metastases confirmed by magnetic resonance imaging (MRI), maximum diameter of each metastasis ≤ 5 cm (in case of 3 metastases: sum of diameters ≤ 12 cm), age ≥ 18 years, Karnofsky Performance Score ≥ 60%). If a biologically effective dose (BED) ≥ 100 Gy (α/ß = 10 Gy) is feasible based on ITV-based planning, patients will be randomized to either MRgRT or ITV-based SBRT. If a lesion cannot be treated with a BED ≥ 100 Gy, the patient will be treated with MRgRT at the highest possible dose. Primary endpoint is the non-inferiority of MRgRT at the MRIdian Linac® system compared to ITV-based SBRT regarding hepatobiliary and gastrointestinal toxicity CTCAE III or higher. Secondary outcomes investigated are local, locoregional (intrahepatic) and distant tumor control, progression-free survival, overall survival, possible increase of BED using MRgRT if the BED is limited with ITV-based SBRT, treatment-related toxicity, quality of life, dosimetric parameters of radiotherapy plans as well as morphological and functional changes in MRI. Potential prognostic biomarkers will also be evaluated. DISCUSSION: MRgRT is known to be both highly cost- and labor-intensive. The MAESTRO trial aims to provide randomized, higher-level evidence for the dosimetric and possible consecutive clinical benefit of MR-guided, on-table adaptive and gated SBRT for dose escalation in critically located hepatic metastases adjacent to radiosensitive OARs. TRIAL REGISTRATION: The study has been prospectively registered on August 30th, 2021: Clinicaltrials.gov, "Magnetic Resonance-guided Adaptive Stereotactic Body Radiotherapy for Hepatic Metastases (MAESTRO)", NCT05027711.

Measuring self-efficacy and outcome expectancy in evidence-based practice: A systematic review on psychometric properties.

Background: Evidence-based practice has developed over the last 30 years as a tool for the best possible nursing care. Nevertheless, many nurses do not regularly participate in the evidence-based practice process. Barriers to participation include nurses' self-perceived ability in successfully fulfilling evidence-based practice-related tasks (self-efficacy) and their expectations of the positive outcomes of such tasks (outcome expectancy). To evaluate progress and provide feedback to professionals, monitoring the levels of self-efficacy and outcome expectancy with validated instruments is desirable. A comprehensive overview of the psychometric properties of such instruments is lacking. Objectives: To determine the psychometric properties of instruments designed to measure nurses' self-efficacy and outcome expectancy in evidence-based practice. Design and method: This systematic review was performed on studies reporting psychometric properties of instruments that measure self-efficacy and outcome expectancy in EBP. MEDLINE, EMBASE and CINAHL databases were searched up to March 2020. Studies that reported psychometric properties on eligible scales and studied nurses or other healthcare professionals were included. Psychometric properties included content validity, construct validity, reliability, and responsiveness. The COSMIN risk of bias checklist and criteria for good measurement properties were applied independently by two reviewers. This review is registered with PROSPERO (CRD42020183069). Results: Eleven scales measuring self-efficacy or a similar construct and one scale measuring outcome expectancy were identified. The vast majority of the research focused on nurses. Internal consistency and structural validity were the most frequently reported properties, though the recommended confirmative factor analysis to verify the structural validity was rarely performed correctly. In addition, most studies that reported on construct validity did not hypothesise on the expected strength or direction of an effect before the data analysis. Responsiveness was not typically reported or was incorrectly studied. The included articles showed a high quality of evidence for four scales on structural validity and internal consistency. The Self-Efficacy in Evidence-Based Practice Activities scale showed the best content validity and was accompanied by an Outcome Expectations of Evidence-Based Practice scale. Both scales met the COSMIN standards for construct validity with high-quality evidence. Conclusions: In light of the evidence, the Self-Efficacy in Evidence-Based Practice Activities scale is considered promising, and along with the accompanying Outcome Expectations of Evidence-Based Practice scale, appears capable of accurately measuring both self-efficacy and outcome expectancy. The use of these scales is recommended, and further research should be conducted on the responsiveness of the scales.

Equivalence of human and mouse CD4 in enhancing antigen responses by a mouse class II-restricted T cell hybridoma.

We have examined the ability of hCD4 to interact functionally with mouse class II MHC molecules using the mouse T cell hybridoma BI-141, specific for beef insulin. We have previously shown that expression of mouse CD4 results in a marked enhancement of IL-2 release by BI-141 cells in response to beef insulin or, in a cross-reactive response, to pork insulin, on the appropriate mouse APCs. We now demonstrate that expression of hCD4 results in an equivalent stimulation of antigen responses by this mouse T cell hybridoma. The specificity of this effect was demonstrated by mAb and gp120 blocking studies. These data provide the first direct evidence for function of hCD4 and in an exclusively mouse system.

The alpha 4 integrin chain is a ligand for alpha 4 beta 7 and alpha 4 beta 1.

The heterodimeric alpha 4 integrins alpha 4 beta 7 lymphocyte Peyer's patch adhesion molecule ([LPAM]-1) and alpha 4 beta 1 (very late antigen-4) are cell surface adhesion molecules involved in lymphocyte trafficking and lymphocyte-cell and matrix interactions. Known cellular ligands include vascular cell adhesion molecule (VCAM)-1, which binds to alpha 4 beta 1 and alpha 4 beta 7, and the mucosal addressin cell adhesion molecule (MAdCAM)-1, which binds to alpha 4 beta 7. Here we show that the alpha 4 chain of these integrins can itself serve as a ligand. The alpha 4 chain, immunoaffinity purified and immobilized on glass slides, binds thymocytes and T lymphocytes. Binding exhibits divalent cation requirements and temperature sensitivity which are characteristic of integrin-mediated interactions, and is specifically inhibited by anti-alpha 4 integrin antibodies, which exert their effect at the cell surface. Cells expressing exclusively alpha 4 beta 7 (TK-1) or alpha 4 beta 1 (L1-2) both bound avidly, whereas alpha 4-negative cells did not. A soluble 34-kD alpha 4 chain fragment retained binding activity, and it inhibited lymphocyte adhesion to alpha 4 ligands. It has been shown that alpha 4 integrin binding to fibronectin involves an leucine-aspartic acid-valine (LDV) motif in the HepII/IIICS region of fibronectin (CS-1 peptide), and homologous sequences are important in binding to VCAM-1 and MAdCAM-1. Three conserved LDV motifs occur in the extracellular sequence of alpha 4. A synthetic LDV-containing alpha 4-derived oligopeptide supports alpha 4-integrin-dependent lymphocyte adhesion and blocks binding to the 34-kD alpha 4 chain fragment. Our results suggest that alpha 4 beta 7 and alpha 4 beta 1 integrins may be able to bind to the alpha 4 subunit on adjacent cells, providing a novel mechanism for alpha 4 integrin-mediated and activation-regulated lymphocyte interactions during immune responses.

Effects of mutagens on the immunogenicity of murine tumor cells: immunological and biochemical evidence for altered cell surface antigens.

Eb lymphoma cells were subjected to treatment in vitro with the alkylating mutagen N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and then cloned by limiting dilution. When tested in vivo for tumorigenicity in groups of syngeneic DBA/2 mice, 6 from 18 clones were found to be strongly reduced (tum- phenotype). The other clones showed only moderate or no change in tumorigenicity compared to the untreated control. All clones were able to grow in 400-rad-irradiated mice. Mice in which MNNG clones had regressed were able to generate tumor-specific cytolytic T-lymphocytes in vitro. Limiting dilution analysis indicated that 3 of 4 MNNG clones analyzed in detail displayed additional antigenic determinants that were detected by cytolytic T-lymphocytes. These data thus provided evidence for increased immunogenicity of some of the MNNG clones. Membrane proteins of MNNG clones and original Eb cells were compared biochemically after metabolic labeling with [35S]methionine, TX114 solubilization, and electrophoretic separation. Two-dimensional gel maps revealed a general quantitative decrease in the expression of membrane proteins in MNNG clones. In addition, several proteins were only found in MNNG clones but not in untreated cells. Two membrane proteins of molecular weight 22,000 and 38,000 were greatly increased in expression in all MNNG clones but could be detected at a low level in the original Eb cells. MNNG is known to be a strong mutagenic agent, but it can also interfere with DNA methylation and cause transcriptional activation of genes. We suggest that amplified cell surface structures may be the consequence of such transcriptional activation and could be involved in altered immunogenicity.

Dynamic expression changes in vivo of adhesion and costimulatory molecules determine load and pattern of lymphoma liver metastasis.

Although intradermal primary tumor growth and spontaneous liver metastasis of ESbL-lacZ lymphoma in syngeneic DBA/2 mice are progressive and malignant, they are characterized by a transient plateau period with a constant tumor diameter and a low number of metastasized cells in the liver. This period, which was shown to be immune dependent, was followed by a second expansion phase characterized by a preferential localization of tumor cells in the periportal areas of liver lobules (mosaic phenotype). To elucidate possible mechanisms leading to the plateau period as well as for the mosaic-like metastasis pattern, we investigated, using flow cytometry analysis, alterations in costimulatory and adhesion molecule expression in liver sinusoidal cells as well as in tumor cells isolated directly ex vivo throughout the kinetics of metastasis. In tumor and sinusoidal cells, we found up-regulation in the expression of MHC class II and B7 molecules during the plateau period. These molecules, which facilitate cell-mediated immune responses, were again down-regulated during the final exponential tumor growth and metastasis. In the final expansion phase, in which the mosaic phenotype of liver metastasis is seen, we detected a significant increase of leukocyte function-associated antigen-1/intercellular adhesion molecule-1 expression in both tumor and sinusoidal cells, suggesting tumor cell-sinusoidal cell interactions. vascular cell adhesion molecule-1/very late activated antigen-4 did not show any modification during the whole metastatic process. In vivo application of monoclonal antibodies directed to leukocyte function-associated antigen-1 and intercellular adhesion molecule-1 appeared to block the spread of metastasis, while no effect was seen with monoclonal antibodies directed to vascular cell adhesion molecule-1 and very late activated antigen-4. This study reveals in situ expression changes of cell surface molecules in tumor and host cells during metastasis. The changes seen during the plateau phase and during the second expansion phase differ, suggesting associations with mechanisms of immune control and tumor immune evasion, respectively.

A clinically relevant HIV-1 subunit vaccine protects rhesus macaques from in vivo passaged simian-human immunodeficiency virus infection.

OBJECTIVES: To investigate whether immunization with recombinant HIV-1 envelope protein derived from a clinical isolate could protect macaques from infection with an in vivo passaged chimeric simian-human immunodeficiency virus (SHIV). DESIGN AND METHODS: A total of 16 animals were studied from which three groups of four animals were immunized with vaccine formulations of the CC-chemokine receptor-5-binding recombinant gp120 of HIV-1W6.1D. Four weeks after the last immunization, all 16 animals were intravenously challenged with in vivo passaged SHIV derived from the same HIV-1 group B clinical isolate (W6.1D) as the vaccines. RESULTS: Vaccine protection from infection was demonstrated in 10 out of 12 macaques immunized with recombinant gp120. Complete protection from infection was achieved with all of the animals that received the SBAS2-W6.1D formulation, a potent inducer of both T-cell and humoral immune responses. Partial protection was achieved with SBAS1-W6.1D, a formulation based on immunomodulators known to induce T-cell responses in humans. In vaccinated animals that were infected, virus load was reduced and infection was delayed. CONCLUSIONS: In a relatively large number of primates, vaccine efficacy was demonstrated with a clinically relevant HIV-1 vaccine. These results reveal that it is possible to induce sterilizing immunity sufficient to protect from infection with SHIV which was passaged multiple times in vivo. Our findings have implications for current HIV-1 clinical vaccine trials and ongoing efforts to develop safe prophylactic AIDS vaccines.

Synthetic biomimetic supra molecular Biovector (SMBV) particles for nasal vaccine delivery.

For the optimal delivery of antigens to mucosal tissues, especially as nasal sprays, protein antigen alone is often not sufficient. A clear need for nasal delivery systems has therefore evolved, especially for Influenza A vaccines. Such technologies will be even more essential for new modern vaccines based on recombinant antigens. Here we describe synthetic biomimetic supra molecular Biovector (SMBV) which have proven in preclinical and clinical evaluation to be suitable candidates for the delivery of nasal vaccines. They also demonstrate the potential to work with multiple antigens and furthermore allow combination with adjuvants. These Biovectors can associate with internal or lipid layer membrane proteins and peptides due to their charged polysaccharide core. The mimicry with viruses is also provided through their size of 60-80 nm, which allows sterilization by filtration. This makes them an ideal tool for the development of modern nasal vaccines, as they have shown to be able to induce the desired types of humoral immunity (serosal and mucosal immunity, IgA and IgG antibodies) as well as cellular immunity (CD4 and CD8 responses).

Synergistic role of type I interferons in the induction of protective cytotoxic T lymphocytes.

Differentiation of cytolytic T cells can be supported by type I and type II interferons (IFN). To characterize the role of type I interferons further we tested the role of recombinant IFN-alpha and IFN-beta on the induction of a weak immune response, against a low immunogenic tumor, which has been shown to be increased by IFN. Both type I interferons IFN-alpha and IFN-beta were able to support the differentiation of cytolytic T lymphocytes (CTL). In case of IFN-alpha no correlation with the antiviral activity could be seen by comparison of IFN-alpha1 and IFN-alpha4. The maximal in vitro effects were achieved with very low concentrations in the range of 1-100 IU/ml. IFN-alpha showed the strongest effects, if added in the early phase of the mixed leukocyte culture, whereas IFN-beta was most effective when given at the last day the culture. In combination, both IFNs gave additional/synergistic effects, whereby addition of IFN-alpha at day 0 and IFN-beta at day 4 led to maximal specific CTL responses. In vivo augmentation of the anti-tumor immune response by both types of IFNs supported the in vitro findings and also the synergistic effect of both types of IFNs could be demonstrated. Therefore we propose that IFN-alpha is relevant in the induction of CTL responses, i.e., the conversion of precursor T cell into mature cells and growth promotion whereby IFN-beta might trigger the lytic machinery of the cells and promote differentiation. This synergistic efficacy is also operative in tumor rejection.

Proteomic prediction of disease outcome in cancer : clinical framework and current status.

Better than gene sequencing or quantitative amplification, proteomics tools allow the study of tumor phenotype. Indeed, most current prognostic tests in cancer (carcinoembryonary antigen [CEA], prostate-specific antigen [PSA], CA 19-1, CA 125, alpha-fetoprotein [AFP], etc.) are based on the detection and quantification of single proteins in body fluids. However, a common characteristic of these tests is their relatively low predictive value, so that they are usually complemented with other procedures such as biopsy and/or endoscopy. Recently, improved analytical and bioinformatics tools have driven the attention on pattern recognition approaches rather then single-marker tests for prognostic forecasting. It is expected that predicting metastasization on the basis of tumoral protein patterns will soon be a reality. However, currently available technologies either limit the number of proteins that can be analyzed simultaneously or they are expensive, difficult, and time-consuming. Moreover, the tools adapted for expression proteomics might not be the same as those for prognostic studies that require investigation of protein function over time. We believe that clinical proteomics research designed within a precise clinical and pathology framework should be strongly supported, since many prognostic factors are determined not by the tumor itself, but by the patient, the treatment and the environment.

Immunological and molecular characterization of an aggressive murine lymphoma variant: modulation in vitro and in vivo.

The highly metastatic murine ESb-L lymphoma was analyzed with respect to its possible origin and phenotype modulation. By determining the methylation status of the CD8 gene an early thymic origin of the ESb-L lymphoma cells is suggested. It revealed that the precursors of ESb-L cells had at least one CD8 allele expressed during their development. ESb-L tumor cells were found to express ICAM-1, ICAM-2, VLA-4 and Mel14 as adhesion molecules and homing receptors and CD25, CD69 and CD124 (HSA) as T-cell related activation markers. PCR analysis revealed that ESb-L tumor cells express a Th2-like cytokine pattern with mRNAs for IL-4, IL-5, IL-6, IL-10 and IL-13, but not for IL-2 and IFNgamma. In addition mRNA for TNFalpha, LT, IFNalpha and the chemokines MIP1alpha and MIP1beta was found. The expression of the adhesion molecules ICAM-1, ICAM-2, VLA-4 and of the T-cell activation marker CD25 on ESb-L tumor cells could be upregulated by incubating the cells with 10 ng/ml TNFalpha. For CD25 this effect was confirmed also at the mRNA level. Using the lacZ transduced T-cell lymphoma ESb-L-CI we were able to re-isolate live tumor cells from the primary site or from a metastasized liver and to investigate their phenotype ex vivo. MIP1alpha mRNA expression was strongly reduced in ex vivo isolated tumor cells as compared to in vitro grown cells indicating the modulatory role of the tumor microenvironment. The presented data suggest possible roles of TNFalpha and/or other microenvironmental factors modulating the expression of molecules involved in cell migration and adhesion thereby influencing cancer metastasis.

Adenovirus-mediated gene transfer of pathogen-associated molecular patterns for cancer immunotherapy.

The delivery of stimulatory signals to dendritic cells (DCs) in the tumor microenvironment could be an effective means to break tumor-induced tolerance. The work presented here evaluates the immunostimulatory properties of pathogen-associated molecular patterns (PAMPs), microbial molecules which bind Toll-like receptors and deliver activating signals to immune cells, when expressed in tumor cells using adenoviral (Ad) vectors. In vitro, transduction of A549 tumor cells with Ad vectors expressing either flagellin from Listeria monocytogenes or P40 protein from Klebsiella pneumoniae induced the maturation of human monocyte-derived DCs in co-cultures. In mixed lymphocyte reactions (MLRs), Ad-flagellin and Ad-P40 transduction of tumor cells stimulated lymphocyte proliferation and the secretion of IFN-gamma. In vivo, these vectors were used either as stand-alone immunoadjuvants injected intratumorally or as vaccine adjuvants combined with a tumor antigen-expressing vector. When Ad-PAMPs were administered intratumorally to mice bearing subcutaneous syngeneic B16F0-CAR (cocksackie-adenovirus receptor) melanomas, tumor progression was transiently inhibited by Ad-P40. In a therapeutic vaccine setting, the combination of Ad-MUC1 and Ad-PAMP vectors injected subcutaneously delayed the growth of implanted RenCa-MUC1 tumors and improved tumor rejection when compared with vaccination with Ad-MUC1 alone. These results suggest that Ad-PAMPs could be effective immunoadjuvants for cancer immunotherapy.

Adhesion versus coreceptor function of CD4 and CD8: role of the cytoplasmic tail in coreceptor activity.

CD4 and CD8 play an important role in T-cell recognition and activation; however, their mechanisms of action are not well understood. We compare the effects of expressing CD4 and CD8 alpha either individually or together in a class II-restricted T-cell hybridoma. We also compare the effects of expressing truncated forms of CD4 or CD8 alpha that do not have a cytoplasmic tail and thus do not associate with the T-cell-specific tyrosine kinase p56lck, which has been implicated in T-cell activation. We demonstrate that, although CD4 and CD8 alpha can specifically enhance interleukin 2 secretion, maximal potentiation occurs with expression of CD4, which, unlike CD8, can bind to the same major histocompatibility complex protein as the T-cell receptor. Our data further indicate that the cytoplasmic tail and/or the associated p56lck are primarily significant for interleukin 2 secretion by the hybridomas we have examined when CD4 or CD8 can bind to the same major histocompatibility complex ligand as the T-cell receptor.

Immunoresistant metastatic tumor variants can re-express their tumor antigen after treatment with DNA methylation-inhibiting agents.

Immuno-escaping variants which arise during metastasis of ESb lymphoma cells in syngeneic DBA/2 mice have been shown to exhibit selective resistance to lysis by ESb-specific cytotoxic T-lymphocytes (CTL). The immuno-resistant variants present no changes in the expression of H-2Kd molecules which appear to be the restricting elements for ESb-specific CTL. We now show that treatment of clonal immuno-resistant ESb variant cells with MNNG or 5'azacytidine can restore the sensitivity to tumor-specific CTL lysis in a high percentage of cloned progenitor cells. The acquisition of susceptibility to lysis by these clones is most likely due to re-expression of ESb-type tumor antigens because such cells regain the capacity to compete with original 51Cr-labelled ESb cells for lysis by ESb-specific CTL, and regain the capacity to induce ESb-specific CTL in vivo. Our data suggest that the immuno-resistant variants are not cellular mutants but rather gene regulatory variants. This could explain: their high frequency of occurrence during metastasis; the relative stability of the variant phenotype; and the reversibility observed after the use of DNA-demethylating and gene-activating drugs like 5'-azacytidine or MNNG.

New antigens presented on tumor cells can cause immune rejection without influencing the frequency of tumor-specific cytolytic T cells.

The specific immune response against syngeneic tumors by T cells is dependent on the existence of tumor-associated transplantation antigens (TATA). In the case of the chemically induced DBA/2-derived lymphoma Eb and its highly metastatic variant ESb the immunogenicity of these antigens is not sufficient to prevent tumor growth. Therefore we tested in two systems the influence of additional antigens as possible helper determinants for the generation of tumor-specific immune responses. In the Eb tumor system additional antigens were induced by mutagenization. The frequency of cytotoxic T lymphocytes (CTL) in response to mutagenized Eb cells was higher than that in response to untreated Eb cells. Fine specificity analysis revealed there there was no increase in the CTL response against the original TATA, but an activation of additional CTL clones responding to mutagen-induced antigens. In the ESb tumor system we tested the effect of additional recognition of minor histocompatibility antigens on the frequency of TATA-specific CTL. Transplantation of ESb tumor cells into B10.D2 mice, which are H-2-identical but differ in minor antigens, results in strong tumor rejection responses. In a limiting dilution mixed-leukocyte-tumor microculture system it was found that the minor antigens are recognized at the clonal level as independent antigens. The overall frequency of anti-tumor CTL in ESb-immunized B10.D2 mice was about 1/3000. Among these, the frequency of TATA-specific CTL was 1/16,709 and thus not significantly different from that of syngeneic DBA/2 mice. Thus neither minor antigens nor mutagen-induced antigens acted in the Eb/ESb tumor system as helper determinants and did not increase the frequency of tumor-specific CTLs.

An immunological role for the CD8 beta-chain.

Mature T cells can be functionally divided into two categories distinguished by surface expression of either CD4 or CD8, which in turn corresponds to restriction by and binding to class II or class I major histocompatibility complex proteins, respectively. CD8 can be expressed as a homodimer of the alpha-chain, or as a heterodimer of alpha- and beta-chains on human and mouse T cells, although most peripheral T cells seem to express CD8 alpha beta heterodimers exclusively (reviewed in ref. 9). Functional characterization of CD8 has focused primarily on the effect of the alpha-chain, which enhances or reconstitutes T-cell responses in homodimeric form and may play a specific role in thymic selection. In contrast, no role has been ascribed to CD8 beta or alpha beta heterodimers specifically. Here we show that CD8 alpha beta transfectants produce more interleukin-2 than CD8 alpha transfectants in response to specific stimuli. Increased interleukin-2 is also observed in cells expressing hybrid CD8 beta-alpha molecules (extracellular CD8 beta plus CD8 alpha transmembrane and cytoplasmic regions) on their surface. These results indicate that external portions of CD8 beta could be critical and that they may act independently of CD8 alpha in mediating their augmentation effect.

Modification of tumor cells by a low dose of Newcastle disease virus. II. Augmented tumor-specific T cell response as a result of CD4+ and CD8+ immune T cell cooperation.

Augmented tumor-specific T cell responses were observed against the high metastatic murine lymphoma variant ESb when using as immunogen ESb tumor cells that had been modified by infection with a low dose of Newcastle disease virus (NDV). Such virus-modified inactivated tumor cells (ESb-NDV) were potent tumor vaccines when applied postoperatively for active specific immunotherapy of ESb metastases. We demonstrate here that immune spleen cells from mice immunized with ESb-NDV contain enhanced immune capacity in both the CD4+, CD8- and the CD4-, CD8+ T cell compartments to mount a secondary-tumor-specific cytotoxic T cell response in comparison with immune cells from mice immunized with ESb. ESb-NDV immune CD4+, CD8- helper T cells also produced more interleukin 2 after antigen stimulation than the corresponding ESb immune cells. There was no participation of either CD4+ or CD8+ virus-specific cells in the augmented response. The specificity of the T cells for the tumor-associated antigen remained unchanged. Thus, there is the paradox that the virus-mediated augmentation of the tumor-specific T cell response in this system involves increased T helper activity but does not involve the recognition of viral epitopes as potential new helper determinants.