Imaging of Prostate-Specific Membrane Antigen with Small-Molecule PET Radiotracers: From the Bench to Advanced Clinical Applications.

In recent years, small-molecule inhibitors of prostate-specific membrane antigen (PSMA) labeled with radionuclides that allow for positron emission tomography (PET) imaging have been extensively studied in many clinical contexts in men with prostate cancer (PCa). The high sensitivity and specificity of these agents for identifying sites of PCa has quickly led to their widespread adoption as a de facto clinical standard of care throughout much of the world. PSMA-targeted PET radiotracers have been particularly well-studied in preoperatively staging men with high-risk PCa, evaluating biochemical recurrence following definitive therapy, and guiding metastasis-directed therapy in patients suspected of having oligorecurrent/oligometastatic disease. Furthermore, the expression of PSMA on the tumor neovasculature of many nonprostate malignancies has enabled a burgeoning subfield concentrated on delineating the potential utility of PSMA-targeted PET agents for imaging other cancers. In this review, we highlight the preclinical development of key small molecules that are now being clinically utilized for PCa imaging, discuss the roles of PSMA-targeted agents in guiding patient management, and consider the role these compounds may play in imaging nonprostate cancers.

Increase of NKG2D ligands and sensitivity to NK cell-mediated cytotoxicity of tumor cells by heat shock and ionizing radiation.

In this study, we have investigated if current cancer therapeutic modalities including hyperthermia and ionizing radiation can increase the expression of NKG2D ligands in human cancer cell lines. The expressions of NKG2D ligands were induced by both heat shock and ionizing radiation in various cell lines including KM12, NCI-H23, HeLa and A375 cells with peaks at 2 h and 9 h after treatment, respectively, although inducibility of each NKG2D ligand was various depending on cell lines. During the induction of NKG2D ligands, heat shock protein 70 was induced by heat shock but not by ionizing radiation. These results were followed by increased susceptibilities to NK cell-mediated cytolysis after treatment with heat shock and ionizing radiation. These results suggest that heat shock and ionizing radiation induce NKG2D ligands and consequently might lead to increased NK cell-mediated cytotoxicity in various cancer cells.

Flow cytometric analysis of thymocyte subpopulations in mice after whole-body X-irradiation.

To determined the cellular kinetics of thymocyte subpopulations in DBA1 mice after whole-body 6.8 Gy X-irradiation, they were analyzed for the expression of several cell surface antigens using flow cytometry. The results show that i) The majority of thymocytes rapidly depleted by irradiation was CD4+8+ cells. ii) radioresistant CD4+8- and CD4-8+ survived 18-48 hr after X-irradiation were considered to be relatively mature type, since they expressed high levels of CD3 and LECAM-1. iii) CD3-positive cells were detected in CD4-8- cells at 72 hr after irradiation.

Identification of surface tegumental antigens of normal and irradiated schistosomula.

Sodium dodecyl sulphate- poly acrylamide gel electronphoresis (SDS-PAGE) fractionation of tegumental surface antigens (STEG-Ags.) of 7-day cultured normal and irradiated schistosomula showed no obvious qualitative differences. The observed polypeptide bands of both normal irradiated STEG-Ags. were almost identical and have similar corresponding molecular weights. The immunoblotting assay, using different types of mouse sera, revealed similarity between the bands of both normal and irradiated STEG-Ags. recognized by each type of mouse serum. No qualitative rather than quantitative differences have been observed. The quantitative differences were reflected in intensively staining of some bands from normal STEG-Ag. rather than their corresponding bands of the same molecular weights from irradiated STEG-Ag.

A low, adaptive dose of gamma-rays reduced the number and altered the spectrum of S1- mutants in human-hamster hybrid AL cells.

We examined the effects of a low, adaptive dose of 137Cs-gamma-irradiation (0.04 Gy) on the number and kinds of mutants induced in AL human-hamster hybrid cells by a later challenge dose of 4 Gy. The yield of S1- mutants was significantly less (by 53%) after exposure to both the adaptive and challenge doses compared to the challenge dose alone. The yield of hprt- mutants was similarly decreased. Incubation with cycloheximide (CX) or 3-aminobenzamide largely negated the decrease in mutant yield. The adaptive dose did not perturb the cell cycle, was not cytotoxic, and did not of itself increase the mutant yield above background. The adaptive dose did, however, alter the spectrum of S1- mutants from populations exposed only to the adaptive dose, as well as affecting the spectrum of S1- mutants generated by the challenge dose. The major change in both cases was a significant increase in the proportion of complex mutations compared to small mutations and simple deletions.

[An experimental in-vitro evaluation of membrane antigen expression and of interleukin production by monocytes exposed to magnetic resonance]. / Valutazione sperimentale in vitro dell'espressione di antigeni di membrana e della produzione di interleuchine da parte di linfomonociti esposti a Risonanza Magnetica.

The interactions of magnetic fields and radiofrequency with biological systems have been described by several authors. However, no definitive conclusions have been drawn yet as to the safety of the magnetic fields and radiofrequencies used in clinical examinations. The immune system is one of the most complex biological systems, in which a network of intracellular signals regulates the immune response. Interleukins are released by an activation process which involves, at least in part, intra- and/or extracellular calcium mobilization. The latter step can be influenced by the in vitro effect of this type of nonionizing radiations produced by an MR system on peripheral blood mononuclear cells. Our results show that the 2-hour exposure to magnetic fields (0.5 T) and radiofrequency caused an increase in the spontaneous release of IL2, IL4, IL10, TNF alpha and INF gamma, while the amount of the same cytokines induced by PHA stimulation was decreased. No differences were observed in the spontaneous or PHA-induced release of IL6 by exposure to magnetic fields (MRI). Furthermore, the expression of the CD 11b molecule was increased at the same time. These results may be useful for us to understand the interactions between magnetic fields and radiofrequencies and the immune system.

Ultraviolet B irradiation and cytomegalovirus infection synergize to induce the cell surface expression of 52-kD/Ro antigen.

Cultured human fibroblasts (MRC-5) have been previously demonstrated to express calreticulin, but not Ro autoantigen, on their surface after human cytomegalovirus (CMV) infection. The present study addresses the question of whether other stimuli, alone or in combination with CMV, can induce the surface expression of Ro autoantigens on human fibroblasts. Using a fixed-cell ELISA to detect autoantigen expression, a synergistic effect between ultraviolet B (UVB) exposure and CMV infection on the surface expression of 52-kD/Ro antigen, but not 60-kD/Ro or calreticulin, was observed. The enhanced expression of 52-kD/Ro antigen was significant and specific, compared with untreated cells, cells infected with CMV alone or irradiated with UVB only, and cells subjected to other treatments, such as low pH. Immunofluorescence studies confirmed these findings and indicated that cells expressed 52-kD/Ro protein on their surface at 24 h after a combined UVB and CMV treatment. These studies provide evidence that synergy between UVB irradiation and CMV infection may play a role in the induction of cell surface expression of the human autoantigen, 52-kD/Ro.

UV-irradiated melanocytic nevi simulating melanoma in situ.

A causative role of UV light in the development of melanocytic neoplasms has often been suggested. In order to investigate the short-term effects of UV light on melanocytic nevi, the morphological and immunohistochemical changes in nevi after a single UV irradiation are studied in 12 nevi from 10 patients and compared with the nonirradiated part of the same nevus. After irradiation more melanocytes above the dermal-epidermal junction are observed in seven nevi, simulating a melanoma in situ in three nevi. Moreover, a marked increase in the expression of HMB-45 is found after irradiation in all investigated nevi, indicating an activation of the melanocytes and active melanosome formation. The metabolic activity correlates with the ultrastructural findings, which show a large cytoplasm, hypertrophic Golgi apparatus, abundant mitochondria, and an increased number of melanosomes of different stages. One week after irradiation, no increase in the proliferative activity of the melanocytes is found. The morphological and immunohistochemical changes after one low dose of UV irradiation should be considered in the differential diagnosis of pigmented skin lesions. The UV-irradiated nevus should be added to the list of so-called simulators of malignant melanoma.

Molecular analysis of mutagenesis by high LET radiation.

Mutation induction by high linear energy transfer [LET] alpha particles and gamma-rays was scored in the human hamster hybrid [AL] cells. Southern blotting technique was used to analyse the molecular changes in the DNA from both the HGPRT- and S1- mutants. Dose dependent mutagenesis in the AL cells irradiated with the charged particles was higher by almost 20 fold at the S1 than the corresponding HGPRT locus. Southern analysis of the mutants induced by the high LET particles showed mostly multilocus deletion at both the HGPRT and S1 genes.

Dose-dependent visible effect on mice Langerhans cell membrane markers.

Ultraviolet B and psoralen plus UVA treatment induce antigenic and enzymatic changes in Langerhans cells (LC). The aim of this study was to investigate the visible (VIS) effect on mice LC surface markers. As visible source, a slide projector equipped with a 150-W tungsten lamp, emitting between 400 and 740 nm (maximum at 580 nm) was used. Mice (BALB/c and C3H) were divided into groups, each irradiated with visible single fixed doses (ranging from 10 to 1000 J/cm2). The mice backs were shaved before irradiation. Skin biopsies obtained immediately after irradiation were processed for immunofluorescence and electron microscopy. Immunofluorescent studies showed: 1) a complete depletion of LC membrane markers at a dose of 700 J/cm2; 2) no effect at visible doses ranging between 0 and 75 J/cm2; 3) a dose-dependent effect with doses between 100 and 700 J/cm2. Electron microscopy revealed no cellular damage of LC at the visible doses administered.

Effects of FK506 on surface antigen expression by regenerating thymocytes after sublethal irradiation in the rat.

To determine the effects of FK506 on the intrathymic maturation of T cells, we analyzed changes in surface antigen expression by regenerating thymocytes after sublethal irradiation. Two-color flow cytofluorometric analysis of the regenerating thymocytes revealed that FK506 had few effects on the maturation of rat thymocytes from CD4-8- to CD4+8+, and from TCR alpha beta- MHC class I- to TCR alpha beta low MHC class I+ (TCR alpha beta, alpha and beta chains of the T cell antigen receptor). However, defects in the maturation of CD4+8+ cells to CD4+8- cells and of TCR alpha beta low MHC class I+ cells to TCR alpha beta high MHC class I+ cells were observed after FK506 administration. Furthermore, three-color analysis also detected a maturational defect of CD4-8+ TCR alpha beta high cells. Immunohistochemical analysis using MoAbs specific for MHC and TCR alpha beta revealed marked atrophy of the thymus medulla, suggesting disturbances in thymocyte regeneration and maturation. These results suggest that the maturation of CD4-8- TCR alpha beta- MHC class I- cells to CD4+8+ TCR alpha beta low MHC class I+ cells may be FK506 resistant, whereas FK506 may interfere with the maturation of CD4+8+ TCR alpha beta low MHC class I+ thymocytes not only to CD4+8- TCR alpha beta high MHC class I+ cells but also to CD4-8+ TCR alpha beta high MHC class I+ cells. We also demonstrated the possibility of enrichment of CD4-8- cells after sublethal irradiation and the existence of small subpopulations of CD4+8- TCR alpha beta- and CD4-8+ TCR alpha beta- cells which might mature to CD4+8+ cells in rat thymus.

Effects of low-doses of X-rays on the expression of human cell surface CD2 antigen in CD2+ CHO cells.

Thioguanine resistant CHO cells (HPRT-) were stably cotransfected with pSV2-gpt and pi H3-CD2 vectors using the calcium phosphate coprecipitation technique. The effects of single low doses of ionizing radiation were studied in a CD2+ CHO clone. The CD2+ phenotype responsible for binding sheep erythrocytes and rosette formation, was not affected by X-rays doses in the range 2-6 cGy. However, after 10 cGy of X-irradiation, 50% of the cells lost the CD2+ phenotype. These results suggest that this CD2+ clone might be a very sensitive indicator of very low X-ray doses. The implications of the phenotypic changes, observed after very low doses of irradiation, are discussed.

Effects of irradiation on surface carbohydrates of larvae of Schistosoma mansoni.

Ultra-violet irradiated larvae of Schistosoma mansoni stimulate high levels of resistance to challenge infection in experimental animals. In the experiments presented here, the binding patterns of antisera specific for the cercarial glycocalyx, and of various lectins, demonstrate that u.v. irradiation causes a pronounced modification of the carbohydrate antigens expressed at the surface of cercariae and newly transformed schistosomula. These alterations were dependent on the irradiation dose, and on the batch of cercariae used in each experiment. Our results strongly suggest that the changes in carbohydrate antigens consequent upon u.v. irradiation may be important in generating the enhanced immunogenicity of irradiated cercariae.

The effects of hormonal and other stimuli on cell-surface Ro/SSA antigen expression by human keratinocytes in vitro: their possible role in the induction of cutaneous lupus lesions.

Ultraviolet B light (UVB) has previously been shown to induce the expression of the extractable nuclear antigens (e.g. Ro/SSA) on the surfaces of human keratinocytes in vitro. This study assessed whether injurious, metabolic, inflammatory, immunological or hormonal stimuli would also induce this expression or modulate that produced by UVB. No stimulus initiated expression alone, but 17-beta oestradiol doubled that found in response to UVB. These findings confirm the potential role of UVB in the initiation and potentiation of cutaneous lupus lesions and may help to explain the female preponderance of the disease.

Ultraviolet radiation (UVR) induces cell-surface Ro/SSA antigen expression by human keratinocytes in vitro: a possible mechanism for the UVR induction of cutaneous lupus lesions.

Antinuclear antibodies are useful markers of connective tissue disease. In this study, UVB but not UVA induced the expression of Ro/SSA antigen on keratinocyte surfaces in vitro. This expression was also found with the extractable nuclear antigens RnP and Sm, but not with single or double-stranded DNA. The expression was prevented by blocking protein synthesis, suggesting that it was an active process. The results suggest that UVB exposure may result in the expression of Ro/SSA antigen on the surfaces of basal keratinocytes in vivo. This antigen could then bind circulating antibody leading to the cutaneous lesions in neonatal and subacute cutaneous lupus erythematosus.

Platelet transfusions irradiated with ultraviolet-B light may have a role in reducing recipient alloimmunization.

The recipients of multiple platelet transfusions frequently develop alloantibodies directed against the human leucocyte antigens (HLA) present on both leucocytes and platelets. Such alloimmunization (AI) may result in refractoriness to further platelet transfusions. Contaminating leucocytes bearing Class II HLA and present in platelet concentrates (PC) are responsible for the formation of HLA antibodies and their removal by filtration reduces the rate of recipient AI. Ultraviolet irradiation (UVR) of PC at an appropriate dose inactivates the contaminating mononuclear leucocytes so that responses in vitro to mitogens and alloantigens are abrogated. It seems likely that UV-irradiation of donor dendritic cells (DC) is important in preventing in vitro responses to alloantigens and in vivo allosensitization. At the same time, satisfactory platelet function and structure is retained when measured by in vitro tests. In vivo assessments of platelet recovery and survival in healthy subjects and the ability to correct the bleeding time in thrombocytopenic patients are comparable to non-irradiated PC. Prospective studies are now in progress to determine if UVR will reduce recipient AI to HLA in multiply-transfused patients with leukaemia and lymphoma.

Immunomodulation by ultraviolet light: clinical studies and biological effects.

The interest of immunologists in ultraviolet (UV) irradiation stems from observations made in vitro and in vivo. In vitro, UV irradiation inhibits mitogen and mixed lymphocyte culture (MLC) responses and in vivo, it can induce cutaneous anergy, apparently via suppressor cells and serum factors. At present much interest is focused on the possible use of UV irradiation to permit transfusion without allosensitization and transplantation without either rejection or graft-versus-host disease (GVHD). Here, Derwood Pamphilon and colleagues discuss the current uses and potential of UV irradiation in transfusion and transplantation and relate these to experimental evidence on its effects at the cellular level.

Changes of CD2 receptor density determined by electron microscopy on blood lymphocytes following radiation treatment for breast cancer.

Changes of relative CD2 receptor on lymphocytes were examined in 11 women following radiation treatment for breast cancer by electron microscopy using antibody-coated gold particles. Proportions of blood lymphocytes with a high density of CD2 receptors were reduced, whereas those with no or a low density of such receptors were increased after radiation treatment.

Effects of irradiation and tunicamycin on the surface glycoproteins of Schistosoma mansoni.

The cercarial glycocalyx and schistosomulum surface contains a number of glycoproteins which are expressed in very variable amounts within a parasite population. Tunicamycin inhibits glycoprotein synthesis of schistosomula if the parasites are incubated for 24 hr with the drug (10 micrograms ml-1). An unexpected increase in lectin binding to the parasite surface was observed but no other changes were detected. Schistosomula treated in this way did not develop in the host past the lung stage. Ultraviolet irradiation (400 microW min cm-2) also inhibited glycoprotein synthesis. Synthesis of other proteins, and in particular heat shock proteins, were also inhibited. Sera from mice (NIH strain) infected with irradiated cercariae contained antibodies which bound to normal schistosomula with lower affinity than to irradiated parasites. This is evidence that irradiation modifies the surface and secreted glycoproteins of schistosomula, so they are processed in a different way to normal glycoproteins by the host's immune system. The effects of irradiation on heat shock protein synthesis may allow the parasite to release a variety of proteins and glycoproteins in abnormal conformations. This may explain the enhanced immunogenicity of irradiated cercariae.