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R C Rees

Publications and source records attributed to R C Rees.

At least 37 records · Page 2Linked to original sources
PubMed1998

Cytogenetics of iris melanomas: disparity with other uveal tract melanomas.

The chromosomal alterations of iris melanomas are poorly characterized, only one report has been detailed. Cytogenetic analysis was performed on the tumors and heparinized blood samples of three patients with iris melanomas; in one case a primary tumor and its related seedling were examined. On analysis of lymphocytes, two of the patients were found to experience a low level fragility of chromosome 9, in the region of a cutaneous melanoma susceptibility gene. All iris melanoma lesions were karyotyped. Clonal abnormalities of chromosomes 3, 5, 6, 7, 8, 9, 12, 15, 17, 18, 19, and Y were found, and in one case a large number of marker chromosomes were observed. No specific chromosomal change was common to the iris melanomas, but two cases had different abnormalities of chromosomes 5 and 18. Variations between the primary tumor and its related seedling were the acquisition of an additional chromosome 15, and a polyploid form of the cell line in the seedling. This study suggests that the most common chromosomal changes of posterior uveal melanomas are less frequent in iris melanomas. Iris melanomas also appear to experience relatively high levels of chromosomal alterations, including the formation of marker chromosomes, which is perhaps reminiscent of cutaneous melanoma.

Chromosome Aberrations↗
PubMed1998

Fibronectin is chemotactic for CT 26 colon carcinoma cells: sub-lines selected for increased chemotaxis to fibronectin display decreased tumorigenicity and lung colonization.

CT 26 murine colon carcinoma cells demonstrated directional migration (chemotaxis) in response to fibronectin (FN). Sub-lines were derived by positive and negative selection to FN across Transwell filters of 8 microm pore size. The FL6 sub-line (positively selected) demonstrated a significantly increased chemotactic response (P<0.01) to FN compared with parental CT 26 cells, while the FU7 sub-line (negatively selected) showed a reduced chemotactic response to FN (P<0.01). Comparable levels of alpha4, alpha5, alphav and beta1 integrins, which mediate FN attachment, were expressed on positively and negatively selected sub-lines and parental CT 26 cells. Activation of integrins with Mn2+ suggested that the integrins expressed on FL6 cells were in the fully activated state; in contrast FU7 cells displayed only partially activated integrins. Cell attachment and integrin activation status of the sub-lines correlated with their chemotactic response to FN. In vivo FL6 cells showed a significantly reduced tumour growth rate s.c. and a reduction in the number of lung colonies formed following i.v. injection compared with parental CT 26 and FU7 cells. In contrast FU7 cells displayed a significant increase in s.c. tumour growth and the number of lung colonies when compared with the parental line and FL6 sub-line. The results indicate that interaction between integrin receptors expressed on cancer cells and FN plays a central role in the chemotactic response of CT 26 colon carcinoma cells, and that in this model cells selected for chemotaxis to FN displayed a reduced malignant potential.

Adenocarcinoma↗
PubMed1998

MAGE, BAGE and GAGE: tumour antigen expression in benign and malignant ovarian tissue.

To determine if ovarian cancer patients would be suitable for MAGE-peptide vaccine-based immunotherapy, the frequency of expression of the MAGE-1-4 genes in ovarian tumours was assessed using reverse transcription polymerase chain reaction (RT-PCR) and product verification with digoxigenin-labelled oligonucleotide probes specific for each MAGE gene. In addition, the frequency of expression of more recently discovered tumour antigens (BAGE, GAGE -1, -2 and GAGE -3, -6) was established using RT-PCR and ethidium bromide staining. In this study 1/16 normal ovarian tissue specimens and 11/25 benign lesions expressed MAGE-1. In non-malignant tissue there was preferential expression of MAGE-1 in premenopausal women. A total of 15/27 malignant specimens expressed MAGE-1, including 10/14 serous cystadenocarcinomas. Expression of other tumour antigens was infrequent. The finding of MAGE-1 expression in both benign and malignant tissue questions previous assumptions regarding the role of MAGE genes in carcinogenesis. In addition, preferential MAGE-1 gene expression in non-malignant premenopausal tissue suggests that the MAGE genes may be involved in cellular proliferation as opposed to carcinogenesis or possibly that MAGE gene expression is under cyclical hormonal control. Finally, this study indicates that serous cystadenocarcinomas may be suitable tumours for MAGE-1 peptide immunotherapy.

Antigens, Neoplasm↗
PubMed1998

Clinical applications of chromosome analysis, from fine needle aspiration biopsies, of posterior uveal melanomas.

PURPOSE: An accurate assessment of prognosis is essential to the clinical assessment of malignancy. In posterior uveal melanoma specific chromosome alterations have been shown to correlate significantly with prognosis; but the procedure is restricted to patients treated surgically, and in consequence has been limited mainly to large tumours. Fine needle aspiration biopsy (FNAB) may provide sufficient material to perform this technique, and allow its use in the in situ assessment of tumours, including small lesions. To determine the feasibility of this approach we have conducted a pilot study using enucleated tumours. METHODS: Ten cases of posterior uveal melanoma were studied. In each instance both a test FNAB and a standard tissue preparation were conducted, and the results compared. FNABS were obtained from enucleated tumours by aspirating cells using a 5 ml syringe with a .25 gauge needle; cells were injected into phosphate-buffered saline, spun down and established in vitro. Conventional short-term cultures were established from tumour tissue samples, which were minced prior to the establishment of cultures. Cytogenetic analysis was performed following standard protocols. RESULTS: Of the 10 cases examined, full chromosome analysis was obtainable from all standard tissue short-term cultures. Cytogenetics was successful from cultures of 6 FNAB, with 2 further FNAB producing partial analyses. No major clonal differences were determined between the two procedures. CONCLUSIONS: Cytogenetic analysis of FNAB appears to be entirely feasible for posterior uveal melanomas, and may permit an accurate in situ assessment of tumours, including small lesions.

Aged↗
PubMed1997

Chemokines induce migrational responses in human breast carcinoma cell lines.

Chemokines have been shown to chemoattract and activate different leukocyte populations. Here we report the in vitro effect of macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, regulated on activation, normal T-cells, expressed and secreted (RANTES), monocyte chemotactic protein-1 (MCP-1), interleukin-8 (IL-8), interferon inducible protein-10 (IP-10), neutrophil-activating peptide-2 (NAP-2), growth-related protein (GRO)-alpha and GRO-gamma, on the migration of 3 human breast carcinoma cell lines, MCF-7, T47D and ZR-75-1, using a microchemotaxis chamber to assess migration across fibronectin-coated polycarbonate membranes. MCF-7 cells responded chemotactically to all chemokines tested in a pattern which was dose and time dependent, although responses to the different chemokines were variable. ZR-75-1 responded to MIP-1beta and GRO-alpha, giving maximum migration indices of 3.7 and 5.3, respectively, and exhibited a migratory response to MIP-1alpha, IL-8 and MCP-1 although to a lower degree. T47D was unresponsive to the chemokines tested, but both MCF-7 and T47D cells bound radiolabelled ligands with binding constants (Kd) ranging from 0.6 to 2.2 nM and 0.6 to 2.1 nM, respectively. The specificity of the chemotactic response of MCF-7 to MIP-1alpha and MIP-1beta was confirmed using chemokine-specific neutralising antibodies and heat denaturation, and was demonstrated to involve G protein and cyclic AMP signalling pathways. MIP-1beta and MIP-1alpha were shown to induce changes in the organisation of the actin cytoskeleton and the level of F-actin in MCF-7 cells, as determined using flow cytometric analysis and confocal microscopy. Our results show that breast carcinoma cells can respond to chemokines, and suggests a potential role for these molecules in the process of tumour cell migration, invasion and metastasis.

Actins↗
PubMed1997

Cytokine and chemokine regulation of proMMP-9 and TIMP-1 production by human peripheral blood lymphocytes.

The production and activation of matrix-degrading proteinases such as the matrix metalloproteinases (MMP) by lymphocytes is likely to be an important factor in facilitating lymphocyte trafficking through the endothelial barrier and the extracellular matrix. Leukocyte infiltration into inflammatory sites occurs as a response to members of the chemokine superfamily and other inflammatory mediators. In the present study, highly purified leukocyte subpopulations were cultured with or without chemokines or cytokines, and their ability to express gelatinolytic MMPs and their inhibitors, the tissue inhibitors of metalloproteinase (TIMPs), was analyzed. In the absence of exogenous stimuli, purified CD4+ T lymphocytes produced similar quantities of proMMP-9 and elevated levels of TIMP-1 compared with PBMC, while purified CD8+ and CD3+ populations exhibited less MMP-9 and TIMP-1 activity. In comparison, CD56+ (NK) cells secreted barely detectable levels of proMMP-9 and TIMP-1. The secretion of proMMP-9 by PBMC and purified CD3+, CD4+, and CD8+ lymphocytes was selectively modulated by beta chemokines and proinflammatory cytokines. ProMMP-9 secretion by CD3+ and CD4+, but not by CD8+ T cells was augmented in response to TNF-alpha and IL-1, and down-regulated by IFN-gamma, while macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, and RANTES (regulated upon activation and normally T cell expressed and secreted) (beta chemokines) up-regulated the secretion of proMMP-9 by all of the lymphocyte subsets tested. These results demonstrate that a number of proinflammatory cytokines and chemokines differentially regulate proMMP-9 secretion from purified CD4+ and CD8+ lymphocytes, while for purified CD3+ T cells (consisting of CD4+ and CD8+ cells in approximately a 2:1 ratio), a predominantly CD4+ lymphocyte response profile was demonstrated.

Chemokines↗
PubMed1997

Abnormalities of chromosomes 3 and 8 in posterior uveal melanoma correlate with prognosis.

Posterior uveal melanomas have nonrandom alterations affecting chromosomes 3, 6, and 8. Loss of chromosome 3 in uveal melanoma has been shown to act as a predictor of disease-free and overall survival. To confirm the significance of chromosome 3 loss and to extend the observations to include those of the associated alterations of chromosome 8, we have conducted a cytogenetic analysis on a series of 42 tumours from patients with primary uveal melanoma who were followed up for a median of 31 months (range = 8-96 months). Abnormalities of chromosomes 3 and 8 were the commonest changes and were confirmed in 10 tumours using fluorescence in situ hybridization. Monosomy of chromosome 3 was found in 21 (50%) of the tumours, and 23 (54%) tumours had additional copies of 8q. Alterations of chromosomes 3 and 8 were found occurring together in 19 (45%) of the tumours and were significantly associated with a ciliary body component (P < 0.0001). Prognostic indicators and changes of chromosomes 3 and 8 were analysed for correlation with patient survival. Of the chosen parameters, only ciliary body involvement (P = 0.003), monosomy of chromosome 3 (P = 0.0007), and additional copies of 8q (P = 0.003) correlated with reduced survival. Evaluation of the dosage effect of additional copies of chromosome arm 8q showed a significant association with reduced survival (P = 0.0001), which was also predictive of a decreased disease-free interval (P = 0.01). Thus, the cytogenetic analysis of uveal melanoma may provide a valuable predictor of prognosis.

Adult↗
PubMed1997

Migration responses of human monocytic cell lines to alpha- and beta-chemokines.

The beta-chemokines monocyte chemotactic protein 1 (MCP-1), macrophage inflammatory protein 1 alpha (MIP-1alpha), MIP-1beta and regulated on activation, normal T cells, expressed and secreted (RANTES) induced the in vitro migration of the monocytic cell line MonoMac-6. MCP-1 exhibits the most potent chemotactic effect on this cell line while MIP-1alpha, RANTES and to a lesser extent MIP-1beta were more moderate inducers of cell migration. MonoMac-6 migration in response to chemokines was shown to be a chemotactic and not a chemokinetic response, which was inhibited by pertussis and cholera toxins suggesting a role for G proteins in chemokine receptor-mediated signalling in these cells; chemotaxis of MonoMac-6 cells in response to MCP-1 was abrogated by the addition of anti-MCP-1 antibody. The response of MonoMac-6 cells to the alpha-chemokines IL-8, IP-10, growth-related peptide (Gro) alpha and MIP-2beta was substantially weaker than to the beta-chemokines. MCP-1 caused an alteration in cellular morphology by increasing ruffling at the cell membrane and the number of cells exhibiting extended pseudopodia. The chemotactic response of MonoMac-6 cells to beta-chemokines was compared with less well-differentiated myelomonocytic cell lines. THP-1 showed a similar, but weaker response to the beta-chemokines while both HL60 and U937 failed to respond to any member of this subfamily when tested under the same conditions. These results suggest that the differentiation status of cells of monocytic lineage may affect their response to beta-chemokines.

Cell Line↗
PubMed1997

The association between nm23 gene expression and survival in patients with sarcomas.

The relationship between the expression of nm23, a putative metastasis-suppressor gene and prognosis was determined for 88 patients with sarcomas. Immunohistochemistry using immunopurified anti-nm23 peptide antibodies was performed and the results of each case graded according to the degree of staining. Univariate and multivariate analyses were carried out to determine the prognostic significance of nm23 staining for sarcoma patients. Expression of nm23 was found to increase in line with metastatic potential in many cases but this did not reach significance for the study as a whole. However, the possibility of nm23 loss occurring in association with metastasis cannot be ruled out in some more aggressive sarcomas, as was demonstrated for six patients with low-scoring, unclassified and synovial sarcomas that had metastasized. The time to metastasis was longer for patients with grade 3 sarcomas (50-75% of tumour cells staining) than similar patients in other staining groups. These results suggest that expression of nm23 genes in sarcomas is variable and has no value as a prognostic indicator for these mesenchymal tumours.

Adolescent↗
PubMed1997

An immunohistochemical study of altered immunomodulatory molecule expression in head and neck squamous cell carcinoma.

For the presentation of peptide antigens to cytotoxic CD8+ T lymphocytes of the immune system, the expression of human leukocyte antigen (HLA) class I molecules on the cell surface is necessary. There is increasing evidence that surface HLA class I antigen expression is altered in a variety of human tumours by either loss or down-regulation of these molecules, which may be a strategy for evasion of immunosurveillance by malignant cells. This study has examined the expression of HLA class I molecules in head and neck squamous cell carcinoma (HNSCC) specimens by immunohistochemistry, using a wide panel of antibodies directed against allele-specific as well as monomorphic determinants of these molecules. The expression of TAP proteins, HLA-DR and the co-stimulatory molecule ICAM-1 were also studied. In addition, the expression of the tumour-associated antigens (TAA) p53 and MAGE genes was determined. Aberrant allelic expression of HLA class I antigens was detected in 17 out of 34 (50%) of the specimens stained, whereas HLA class I expression determined by W6/32 staining was found to be heterogeneous in only 2 out of 34 (6%) cases. Decreased expression of ICAM-1 was observed in 12 out of 34 (35%) tumour specimens and de novo expression of HLA-DR (HLA class II) by carcinoma cells in 13 out of 34 (38%) cases. Aberrant expression of HLA class I antigens was frequently observed in cases in which MAGE genes and p53 overexpression were detected. The altered expression of these immunomodulatory molecules in HNSCC may affect prognosis and has important implications for peptide-based immunotherapy strategies for these patients.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

Trafficking of activated lymphocytes into the RENCA tumour microcirculation in vivo in mice.

The aim of the study was to establish a model of tumour microcirculation in vivo using the murine renal cell carcinoma cell line (RENCA) implanted into the mouse cremaster muscle, and subsequently to investigate the trafficking of syngeneic lymphocyte subpopulations into both the RENCA tumour and the surrounding normal cremaster muscle microcirculation. We have demonstrated that RENCA tumour cells, at a dose of 1.5 x 10(5) per 30 microl injected into the cremaster muscle, reproducibly produced a vascularized tumour suitable for in vivo microscopy at 10-14 days. Injection of fluorescently labelled effector cells (1 x 10(6)) including naive splenocytes, T-cell enriched populations and ex vivo interleukin 2 (IL-2)-activated splenocytes all migrated to and flowed through both the tumour and the normal microcirculation, with negligible adhesion. However, we observed the selective recruitment, localization and arrest of IL-2-activated splenocytes (P < 0.05) into the tumour microcirculation, and the subsequent extravasation of cells into the tumour intestitium in some instances. This did not occur with the other effector cells. We also observed the absence of leucocyte rolling in the tumour microcirculation, suggesting an impairment in adhesion molecule expression on the tumour endothelium. We have therefore established the potential of this model for defining further effector cell-tumour-endothelium interactions.

Animals↗
PubMed1997

DNA index and % S phase fraction in posterior uveal melanoma: a 5 year prospective study of fresh tissue using flow cytometry.

In many different solid tumours the DNA content of cells has been reported to be associated with malignancy. With one exception, studies using flow cytometry to estimate cellular DNA in uveal melanoma have used archival material. We report the findings of a prospective study using fresh uveal melanoma where the DNA index (DI) and percentage of cells in S phase (%SPF) have been determined by flow cytometry in 51 patients with a minimum follow-up of 5 years. Forty-one (80%) of the tumours were large, 9 (18%) were medium and only 1 was small. Eight (16%) were epithelioid tumours, 41 (80%) were mixed cell and only 2 were spindle B cell tumours. Twenty-seven tumours were diploid, of which 10 (37%) developed metastases. Of the 24 aneuploid tumours, metastases developed in 6 (25%). The median %SPF was significantly reduced in patients with diploid tumours compared with aneuploid tumours (p < 0.0001), but there were no significant differences in %SPF between tumours associated with or without metastatic disease. The DI and %SPF were not significantly associated with the operation performed, cell morphology, tumour size or location. Neither %SPF nor aneuploidy was associated with a poor prognosis. Our results suggest that the estimation of DI or %SPF by flow cytometry is of little value in predicting survival from uveal melanomas after a minimum follow-up of 5 years.

Adult↗
PubMed1997

Cytokine modulation of antigen expression in human melanoma cell lines derived from primary and metastatic tumour tissues.

The constitutive and cytokine-mediated expression of MHC class I and II antigens and intercellular adhesion molecule-1 (ICAM-1) was evaluated on eight human melanoma cell lines derived from primary and metastatic malignancies from patients (WM human melanoma series) including three pairs of related cell lines derived from the same individual. The cytokines IL-1 beta, IL-4, IL-6, TNF alpha, TGF beta 2, IFN gamma and IFN alpha were assessed for their ability to modulate the expression of cell surface antigens. MHC class I and class II antigen expression was unregulated by IFN gamma, IFN alpha and/or TNF alpha in cell lines established from primary melanoma. In contrast the cell lines derived from metastatic deposits did not show an increase in expression of MHC antigens in response to these cytokines. Both primary and metastatic WM cell lines were shown to be resistant to spontaneous natural killer cell (NK) activity, but susceptible to effector lymphocytes mediating lymphotine activated killer (LAK) cytotoxicity as a result of activation by IL-2. Although the constitutive and cytokine-induced level of expression of ICAM-1 and MHC antigens varied between paired primary and metastatic cell lines, this did not correlate with susceptibility of the cell line target to NK or LAK cytotoxicity. Whereas the IFNs, TNF alpha, TGF beta 2 and IL-1 beta differentially modulated the expression of ICAM-1 and MHC class I, treatment with IFNs (but not IL-1 beta, TNF alpha or TGF beta 2) resulted in a significant reduction in the sensitivity of the melanoma cells to NK and LAK cytotoxicity. Constitutive ICAM-1 expression was positively correlated with the ability of WM cell lines to colonise the lungs of SCID mice upon i.v. injection. The acquisition of cytokine resistance and inability to demonstrate enhanced cell surface expression may represent an important feature associated with the development of the metastatic phenotype.

Animals↗
PubMed1997

Natural killer and lymphokine-activated cytotoxicity following anaesthesia in patients with uveal malignant melanoma.

The role of enucleation in the treatment of ocular melanoma has been questioned by those who believe that it may increase circulating melanoma cells, increasing the incidence of metastases. Some aspects of the immune system are depressed by general anaesthesia, therefore this study was initiated to assess the effect of general anaesthesia on natural killer (NK) cell and interleukin-2 (IL-2)-activated cytotoxicity in 19 patients using 51Cr-labelled target cells (K 562 and SW 742). NK cytotoxicity was increased at induction and during the operation which was due to significant increases in female, but not male patients. At 1 week post-surgery, female patients had decreased NK cytotoxicity compared with males at this interval. NK cytotoxicity compared with males at this interval. NK cytotoxicity was increased intra-operatively when alfentanyl was used, and if the mean tumour diameter was > 15.0 mm the NK cytotoxicity was increased intra-operatively compared with that in patients with smaller tumours. IL-2-stimulated cytotoxicity was reduced at 1 day post-surgery in females only. There was an increased IL-2-stimulated cytotoxicity at induction when propofol was used and IL-2-stimulated cytotoxicity was reduced at 1 week post-operatively if the patient's mean tumour diameter was > 15.0 mm. If NK cell and lymphokine-stimulated cell cytotoxicity are involved in limiting metastatic disease, the patient' sex and tumour size and the anaesthetic agent used may influence the survival rates following surgery.

Adult↗
PubMed1996

Activation of STAT4 by IL-12 and IFN-alpha: evidence for the involvement of ligand-induced tyrosine and serine phosphorylation.

The immunoregulatory cytokine IL-12 plays a central role in cell-mediated immune responses through its effects on NK cells and T lymphocytes. While IL-12 is known to share some functions with other cytokines, such as IFN-alpha, it also maintains distinct roles, such as its ability to induce Th1 differentiation. The molecular basis for these unique and overlapping functions is not well understood. IL-12 has previously been shown to induce tyrosine phosphorylation and DNA-binding of STAT3 and STAT4, members of the signal transducers and activators of transcription (STAT) family. Because STAT4 has only been shown to be activated in response to IL-12, this specificity has been suggested to be a basis for the unique actions of IL-12. In this study, we demonstrated that STAT4 activation by IL-12 is not unique; IL-12 and IFN-alpha, but not IFN-gamma, induced tyrosine phosphorylation and DNA binding of STAT4. Since tyrosine and serine phosphorylation of STAT1 have previously been shown to be important in IFN-alpha-mediated signaling, we also investigated IL-12- and IFN-alpha-induced serine phosphorylation of STAT4. We demonstrated that both cytokines induced serine phosphorylation. This modification was not required for DNA binding, but may be important in STAT-mediated transcription. Thus, STAT4 activation was not IL-12 specific, and IL-12 and IFN-alpha activated STAT4 through both tyrosine and serine phosphorylation. These findings have significant implications for understanding the role of STAT4 in mediating biologic functions; specifically, the data argue that the unique effects of IL-12 cannot be solely explained by STAT4 activation.

Animals↗
PubMed1996

Investigation of the effect of BB-94 (batimastat) on the colonization potential of human lymphoma cells in SCID mice.

We have assessed the effectiveness of the metalloproteinase inhibitor BB-94 (batimastat) in reducing the colonization potential of the human Burkitt lymphoma Namalwa cell line. In this study Namalwa cells were injected intraperitoneally into SCID mice and their spread to the spleen, liver and lung studied over a 3 week period. The human cells were detected in the mouse tissues by polymerase chain reaction (PCR) amplification of a human alu repeat sequence. Comparison of BB-94-treated animals with an untreated control group provided no evidence for a significant reduction in the colonization of mouse tissues by the human lymphoma cells in the presence of the drug. Tumour growth, after subcutaneous injection of the Namalwa cells into SCID mice, was similarly unaffected by BB-94. The significance of these results is discussed.

Animals↗
PubMed1996

Infrequent expression of the MAGE gene family in uveal melanomas.

It has previously been reported that MAGE-1, -2, -3 and -4 genes are expressed in human cancers including cutaneous melanoma. MAGE-1 and MAGE-3 represent targets for specific immunotherapy because they encode peptide antigens which are recognised by cytotoxic T lymphocytes (CTL) when presented by HLA class I molecules, and pilot clinical trials with these peptides are currently in progress. It is likely that other members of the MAGE gene family may also encode antigens recognised by CTL. Uveal melanomas, like cutaneous melanomas, arise from melanocytes that are derived from the neural crest. To determine if uveal melanoma patients would be suitable for MAGE-peptide immunotherapy, the expression of MAGE-1, -2, -3 and -4 genes was assessed by reverse transcription followed by polymerase chain reaction (RT-PCR) amplification and ethidium bromide staining. Expression of MAGE genes was not detected in any of 27 primary tumours. Either MAGE-1 or MAGE-4 was expressed in only 2 of 26 metastatic samples, but expression of MAGE-2 or -3 was not detected. Our data suggest that, unlike cutaneous melanomas, uveal melanomas may not be suitable candidates for MAGE-peptide immunotherapy.

Antigens, Neoplasm↗