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Triple negative breast cancers (TNBC) are characterized by aggressive tumor biology, lack of targeted treatments and poor prognosis. Anthracyclins were shown to induce immunogenic death in target cells, potentially leading to "endogenous" vaccination. We comparatively assessed expression of cancer testis antigens (CTA) and topoisomerase 2-alpha (TOPO2A), a well defined molecular target of anthracyclins, in TNBC fully characterized for basal-like (BL) immunophenotype, BL morphology and conventional clinicopathological factors. The study included 83 patients undergoing surgery between January 2003 and December 2009. Tissue sections were stained with CK5/6, CK14, EGFR, Ki-67, TOPO2A, MAGE-A1, MAGE-A10, NY-ESO and multi-MAGE-A specific reagents. Of the 83 TNBC, >66.3% had BL immunophenotype and 48.2% had BL morphology. MAGE-A1 specific staining was most frequently detectable (69.2%), followed by multi-MAGE-A (58%), NY-ESO (27.1%) and MAGE-A10 (16%) specific staining. MAGE-A10 expression significantly correlated with tumor size (p=0.026). Furthermore, MAGE-A1, MAGE-A10 and multi-MAGE-A specific stainings significantly correlated with advanced clinical stage (p=0.024, p=0.041, p=0.031, respectively). We found no significant association between CTA expression and disease free (DFS) or overall survival (OS). Most interestingly, a significant correlation was observed between expression of MAGE-A10 and NY-ESO and expression of TOPO2A (p=0.005, p=0.013). Expression of defined CTA and TOPO2A are significantly correlated in TNBC. Considering the limited therapeutic options for TNBC, these findings might suggest novel forms of combination therapies that should be further explored. [ABSTRACT FROM AUTHOR]

We have explored MICA/B expression and its relationship with innate inflammatory infiltrate in renal cell carcinoma (RCC). The expression of MICA/B, CD16, CD56, and CD68 in 140 RCC lesions contained in a tissue microarray (TMA) was investigated by immunohistochemistry. MICA/B gene and protein expressions in Caki-1 cells were analyzed by reverse transcription-polymerase chain reaction and flow cytometry, respectively. Natural killer (NK) cells were studied by flow cytometry. All the RCC lesions (n = 140)wereMICA/B-positive. MICA/B was mainly expressed in the cytoplasm of tumor cells, whereas stromal cells were negative. Renal cell carcinoma lesions showed low NK cell infiltration, although they were rich in CD16+CD56- cells, strongly resembling macrophages. CD16+ macrophage infiltration was more frequently detectable in metastatic lesions compared with primary tumors (P = .0223) and was associated with poor RCC differentiation (P = .007). To investigate mechanisms potentially underlying the lack of NK cells infiltration into MICA/B-positive RCC lesions, we used Caki-1 RCC cells. Caki-1 expressed MICA and MICB genes. However, MICA protein was not detectable in Caki-1 cells, whereas MICB protein was detectable in their cytoplasm and on the cell membrane. Coculture of peripheral blood mononuclear cells with Caki-1, K562, HCT116, respectively, resulted in CD56+CD16+ NK cells deletion without affecting CD56+/CD16- NK subset and immature NK cells generated in vitro from CD34+ cells. Natural killer cell apoptosis seemed to be preferentially triggered by cancer cells because HLA-A0201+ NK cells were only marginally affected by allogeneic HLA-A0201- peripheral blood mononuclear cells. Caki-1 cell-mediated NK cell apoptosis was reduced by an anti-β2-integrin (CD18) monoclonal antibody but was NKG2D-, granule exocytosis-, and caspase-independent. [ABSTRACT FROM AUTHOR]

Cancer/testis tumour-associated antigens (C/T TAA) were the first human tumour-associated antigens to be characterised at the molecular level. Specific genes are expressed in the testis and in tumours of varying histological origin. The tissue expression pattern supports the notion that these antigens could be targets for active specific immunotherapy. Specific serological reagents have been developed and have helped to clarify biochemical characteristics of C/T TAA and to assess their distribution within clinical tumour samples. We review immunohistochemical evidence of the expression of C/T TAA known to be recognised by specific cytotoxic T lymphocytes. The emerging picture is consistent with a mostly heterogeneous expression in human cancers. These findings support the concept of multiantigenic tumour vaccine preparations. Moreover, the wide range of tumours in which C/T TAA have been detected urges further efforts to develop effective specific immunotherapeutic procedures. [Copyright &y& Elsevier]

Objective: The aim of this study was to evaluate the prevalence of cancer testis tumor-associated antigens MAGE-A and NY-ESO-1 in cervical cancer and correlate expression patterns with clinicopathologic parameters and prognosis. Study Design: One hundred sixty-two cervical cancer samples from 109 patients who were treated with radical hysterectomy, neoadjuvant chemotherapy, or pelvic disease recurrence were analyzed by immunohistochemistry. Results: MAGE-A was expressed by 32/94 (34%) and 7/15 (47%) previously untreated and recurrent tumors, respectively. NY-ESO-1 was expressed by 46/94 (49%) and 6/15 (40%) previously untreated and recurrent tumors, respectively. MAGE-A in early stage tumors was correlated to tumor size and lymph node metastases (P = .024 and P = .046, respectively) whereas NY-ESO-1 to tumor grading (P = .039). Conclusion: Cervical cancer frequently expresses cancer testis tumor-associated antigens. MAGE-A and NY-ESO-1 expression rates are not influenced by systemic therapies. Cancer testis tumor-associated antigens are correlated to common prognostic factors. [Copyright &y& Elsevier]

Spontaneous Ca2+ oscillations have been observed in a number of excitable and non-excitable cells, but in most cases their biological role remains elusive. In the present study we demonstrate that spontaneous Ca2+ oscillations occur in immature human monocyte-derived dendritic cells but not in dendritic cells stimulated to undergo maturation with lipopolysaccharide or other toll like-receptor agonists. We investigated the mechanism and role of spontaneous Ca2+ oscillations in immature dendritic cells and found that they are mediated by the inositol 1,4,5-trisphosphate receptor as they were blocked by pretreatment of cells with the inositol 1,4,5-trisphosphate receptor antagonist Xestospongin C and 2-aminoethoxydiphenylborate. A component of the Ca2+ signal is also due to influx from the extracellular environment and may be involved in maintaining the level of the intracellular Ca2+ stores. As to their biological role, our results indicate that they are intimately linked to the "immature" phenotype and are associated with the translocation of the transcription factor NFAT into the nucleus. In fact, once the Ca2+ oscillations are blocked with 2-aminoethoxydiphenylborate or by treating the cells with lipopolysaccharide, NFAT remains cytoplasmic. The results presented in this report provide novel insights into the physiology of monocyte-derived dendritic cells and into the mechanisms involved in maintaining the cells in the immature stage. [ABSTRACT FROM AUTHOR]

Abstract: The capacity of pro-inflammatory cytokines to modulate proteolysis was analyzed by liquid chromatography using human fibroblasts as cell model and enzyme source, and the immunodominant epitope gp100280–288 (YLEPGPVTA) as substrate. The measurements made after fibroblast pre-incubation with either IL-1, TNF, or IL-6 plus its soluble receptors have been compared with those made with un-stimulated fibroblasts. The results obtained suggest an uneven association of cytokine treatment with substrate degradation, and with a prevailingly positive – but also negative – association with release of smaller peptides and free amino acids. Data obtained by separately measuring these two groups of by-products indicate that, after IL-1 cell pre-treatment, the velocity of formation of both groups of by-products increased, resulting in a net increase of substrate degradation. After TNF and IL-6 pre-treatment, the increase of one group was compensated by a decrease of the other group; specifically, the compensation was only partial for TNF, and overall substrate hydrolysis increased. In the case of IL-6, the increase of free amino acids was almost exactly compensated by a reduction of peptidic by-products, resulting in a negligible increase of substrate hydrolysis. In addition, the existence of reaction time-related modifications in the apparent velocity of substrate degradation and formation of by-products, allows hypothesizing different effects of cytokines on the enzymes degrading the substrate with different time constants. Taken together, these data can be interpreted as indicating different, positive and negative, effects of the three cytokines on the individual enzymes expressed by fibroblasts and capable of degrading peptidic substrates. [Copyright &y& Elsevier]

Anticancer compound screening on 2D cell cultures poorly predicts “in vivo” performance, while conventional 3D culture systems are usually characterized by limited cell proliferation, failing to produce tissue-like-structures (TLS) suitable for drug testing. We addressed engineering of TLS by culturing cancer cells in porous scaffolds under perfusion flow. Colorectal cancer (CRC) HT-29 cells were cultured in 2D, on collagen sponges in static conditions or in perfused bioreactors, or injected subcutaneously in immunodeficient mice. Perfused 3D (p3D) cultures resulted in significantly higher (p < 0.0001) cell proliferation than static 3D (s3D) cultures and yielded more homogeneous TLS, with morphology and phenotypes similar to xenografts. Transcriptome analysis revealed a high correlation between xenografts and p3D cultures, particularly for gene clusters regulating apoptotic processes and response to hypoxia. Treatment with 5-Fluorouracil (5-FU), a frequently used but often clinically ineffective chemotherapy drug, induced apoptosis, down-regulation of anti-apoptotic genes ( BCL-2 , TRAF1 , and c-FLIP ) and decreased cell numbers in 2D, but only “nucleolar stress” in p3D and xenografts. Conversely, BCL-2 inhibitor ABT-199 induced cytotoxic effects in p3D but not in 2D cultures. Our findings advocate the importance of perfusion flow in 3D cultures of tumor cells to efficiently mimic functional features observed “in vivo” and to test anticancer compounds. [ABSTRACT FROM AUTHOR]

Abstract: We have investigated monocyte and T cell responsiveness to silk based biomaterials of different physico-chemical characteristics. Here we report that untransformed CD14+ human monocytes respond to overnight exposure to silk fibroin-based biomaterials in tridimensional form by IL-1β and IL-6, but not IL-10 gene expression and protein production. In contrast, fibroin based materials in bidimensional form are unable to stimulate monocyte responsiveness. The elicitation of these effects critically requires contact between biomaterials and responding cells, is not sustained and becomes undetectable in longer term cultures. We also observed that NF-κβ and p38 MAP kinase play key roles in monocyte activation by silk-based biomaterials. On the other hand, fibroin based materials, irrespective of their physico-chemical characteristics appeared to be unable to induce the activation of peripheral blood T cells from healthy donors, as evaluated by the expression of activation markers and IFN-γ gene. [Copyright &y& Elsevier]

Abstract: T-cadherin is gaining recognition as a determinant for the development of incipient invasive squamous cell carcinoma (SCC). However, effects of T-cadherin expression on the metastatic potential of SCC have not been studied. Here, using a murine model of experimental metastasis following tail vein injection of A431 SCC cells we report that loss of T-cadherin increased both the incidence and rate of appearance of lung metastases. T-cadherin-silenced SCC metastases were highly disordered with evidence of single cell dissemination away from main foci whereas SCC metastases overexpressing T-cadherin developed as compact, tightly organised sheets. SCC cell adhesion to vascular endothelial cells (EC) in culture was increased for T-cadherin-silenced SCC and decreased for T-cadherin-overexpressing SCC. Confocal microscopy showed that T-cadherin-silenced SCC adherent on EC display an elongated morphology with long thin extensions and a high degree of intercalation within the EC monolayer, whereas SCC overexpressing T-cadherin formed poorly-spread multicellular aggregates that remain on the outer surface of the EC monolayer. T-cadherin-deficient SCC or human keratinocyte cells exhibited increased transendothelial migration in vitro which could be attenuated in the presence of EGFR inhibitor gefitinib. Our data suggest that loss of T-cadherin can increase metastatic potential and aggressiveness of SCC, possibly due to facilitating arrest and extravasation through the vascular wall and/or more efficient establishment of metastases in the new microenvironment. [Copyright &y& Elsevier]

Abstract: Background: Cancer initiation and progression might be driven by small populations of cells endowed with stem cell-like properties. Here we comparatively addressed the expression of genes encoding putative stemness regulators including c-Myc, Klf4, Nanog, Oct4A and Sox2 genes in benign prostatic hyperplasia (BPH) and prostate cancer (PCA). Methods: Fifty-eight PCA and thirty-nine BPH tissues samples were used for gene expression analysis, as evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). The expression of specific Klf4 isoforms was tested by conventional PCR. Klf4 specific antibodies were used for protein detection in a tissue microarray including 404 prostate samples. Results: Nanog, Oct4A and Sox2 genes were comparably expressed in BPH and PCA samples, whereas c-Myc and Klf4 genes were expressed to significantly higher extents in PCA than in BPH specimens. Immunohistochemical studies revealed that Klf4 protein is detectable in a large majority of epithelial prostatic cells, irrespective of malignant transformation. However, in PCA, a predominantly cytoplasmic location was observed, consistent with the expression of a differentially spliced Klf4α isoform. Conclusion: Klf4 is highly expressed at gene and protein level in BPH and PCA tissues but a cytoplasmic location of the specific gene product is predominantly detectable in malignant cells. Klf4 location might be of critical relevance to steer its functions during oncogenesis. [Copyright &y& Elsevier]

Recombinant vaccinia virus (rVV) encoding tumor-associated antigens (TAAs) and adhesion or costimulatory molecules may represent important immunogenic reagents for cancer immunotherapy. Recently, intranodal (IN) antigen administration was suggested to be more immunogenic than intradermal (ID) vaccination. However, IN rVV administration has not been attempted so far. We used a rVV encoding gp100280–288, Melan-A/MART-127–35 and tyrosinase1–9 HLA-A0201 restricted epitopes and CD80 and CD86 costimulatory molecules in stage III and IV melanoma patients in a phase 1/2 trial. Of 15 patients initiating treatment, including two cycles of IN immunization, each comprising one rVV administration and three recall injections of the corresponding peptides, accompanied by subcutaneous granulocyte macrophage–colony stimulating factor supplementation, five withdrew due to progressing disease. Of 10 remaining patients seven showed evidence of induction of cytotoxic T lymphocytes (CTLs) directed against at least one epitope under investigation, as detectable by limiting dilution analysis (LDA) of specific precursors and multimer staining. Adverse reactions were mild (National Cancer Institute (NCI) grade 1–2) and mainly represented by fever, skin rashes, and pruritus. These data indicate that IN administration of rVV encoding melanoma-associated epitopes and costimulatory molecules is safe and immunogenic. [ABSTRACT FROM AUTHOR]

Abstract: Arginase 2, inducible- and endothelial-nitric-oxide synthase (iNOS and eNOS), indoleamine 2,3-dioxygenase (IDO) and TGF-β, might impair immune functions in prostate cancer (PCA) patients. However, their expression was not comparatively analysed in PCA and benign prostatic hyperplasia (BPH). We evaluated the expression of these genes in PCA and BPH tissues. Seventy-six patients (42 BPH, 34 PCA) were enrolled. Arginase 2, eNOS and iNOS gene expression was similar in BPH and PCA tissues. TGF-β1 gene expression was higher in BPH than in PCA tissues (p =0.035). IDO gene expression was more frequently detectable (p =0.00007) and quantitatively higher (p =0.00001) in PCA tissues than in BPH. IDO protein, expressed in endothelial cells from both BPH and PCA, was detectable in tumour cells in PCA showing evidence of high specific gene expression. In these patients, IDO gene expression correlated with kynurenine/tryptophan ratio in sera. Thus high expression of IDO gene is specifically detectable in PCA. [Copyright &y& Elsevier]

Experimental models indicate that tumor cells in suspension, unlike solid tumor fragments, might be unable to produce life-threatening cancer outgrowth when transferred to animal models, irrespective of the number of cells transferred, although they induce specific immune responses. Human tumor cells cultured in three dimensions display increased pro-angiogenic capacities and resistance to interferons, chemotherapeutic agents or irradiation, as compared with cells cultured in two-dimensional (2D) monolayers. Tumor cells cultured in three dimensions were also shown to be characterized by defective immune recognition by cytotoxic T lymphocytes (CTLs) specific for tumor-associated antigens (TAAs) and by a capacity to inhibit CTL proliferation and dendritic cell (DC) functions. Downregulation of human leukocyte antigen (HLA) or TAA expression and high production of lactic acid might play a role in the elicitation of these effects. Here, we propose that growth in 3D architectures might provide new insights into tumor immunology and could represent an integral missing component in pathophysiological tumor immune escape mechanisms. [Copyright &y& Elsevier]

Abstract: To the exception of early stages of disease, the morbidity and mortality of melanoma is considerable, with no acknowledged therapeutic options beyond surgery. Immunotherapy of melanoma has achieved some success, but further refinements are urgently needed in order to realize its potential. This paper describes a multi-centre phase I/II open labeled, controlled clinical trial investigating 2 innovative immunotherapeutic reagents. Two successive groups of 20 resected AJCC stages IIb–IV melanoma patients will be treated, first with melanoma epitopes included into Influenza virosomes (group 1), and second with a heterologous prime-boost protocol priming with a recombinant Vaccinia virus, and boosting with Influenza virosomes (group 2). Five melanoma epitopes from three different melanoma differentiation antigens were included into Influenza virosomes, that cross-stimulate CD4+ T cells and are endowed with high adjuvant capacity in the generation of CTL. The same five melanoma epitopes, two co-stimulatory molecules CD80 and CD86, and the CD40 ligand, a marker known to play a crucial role in CTL generation and memory maintenance were encoded in a recombinant Vaccinia virus. GM-CSF will be administered as a supporting cytokine. Both Influenza virosomes and octo-recombinant Vaccinia virus are innovative and original constructs assessed for the first time in human. Immunotherapy foresees 12 weekly immunizations for each group. Toxicity and adverse events will be monitored clinically. Immunological efficacy will be assessed dynamically by ex-vivo multimer analysis, Elispot, and quantitative real-time PCR for up to 3 months following completion of immunotherapy schedule. Disease free survival will be assessed by 4-monthly serial clinic visits, including physical and FDG-PET examinations, for a follow-up time of 2 years. Quality of life will be assessed with a dedicated FACT–BRM 4 questionnaire. [Copyright &y& Elsevier]

Abstract: Cancer testis tumour associated antigens (C/T-TAAs) were investigated in several gynaecologic and non-gynaecologic neoplasms as possible prognostic markers and targets for immunotherapy. The objective of the present study was to evaluate C/T-TAA expression patterns and prognostic significance in patients affected by vulvar cancer. Melanoma antigen E (MAGE)-A1, MAGE-A4 and NY-ESO-1 expression was determined by immunohistochemistry in paraffin-embedded tissue specimens from 45 primary and 14 recurrent vulvar carcinomas treated with surgery. MAGE-A1, MAGE-A4 and NY-ESO-1 were expressed in 25 (42%), 38 (64%) and 40 (68%) of the 59 samples, respectively. MAGE-A4 was significantly more frequently expressed in tumours with lymph node metastases (p <0.002) and in recurrent tumours (p <0.02). NY-ESO-1 was more highly expressed by moderately or poorly differentiated tumours (p <0.01). This study demonstrates that vulvar cancer frequently expresses C/T-TAAs. Antigen expression correlates with the presence of lymph node metastases and poor tumour differentiation. [Copyright &y& Elsevier]

Buruli ulcer (BU) caused by Mycobacterium ulcerans is a chronic necrotizing disease of the skin and the underlying soft tissue. Fat tissue necrosis accompanied by minimal inflammation is considered the most reliable histopathologic feature of BU. There may be a constant influx of inflammatory cells to the sites of active infection but these are thought to be killed by mycolactone, a polyketide toxin produced by M. ulcerans, through apoptosis and necrosis. Here we describe the spatial correlations between mycobacterial load and the expression of dendritic cell (DC) surface markers (cluster of differentiation (CD)83, CD11c, and CD123), the Toll-like receptor (TLR) 9 and pro- and anti-inflammatory cytokines (IL-8, IL-6, tumor necrosis factor-alpha (TNF-α), IFN-α, IL-12p40, IL-10, and IFN-γ) within BU lesions. Although IL-8, IL-6, and TNF-α messenger RNA (mRNA) was detectable by real-time PCR in all lesions, the expression of the other cytokines was only found as small foci in some lesions. Correlations of the distribution of mRNA encoding the activation marker CD83 and the DC subset markers CD123 and CD11c indicate that both activated plasmacytoid and myeloid dendritic cells were present in the lesions. Results suggest that M. ulcerans specific immune responses may develop once therapeutic interventions have limited the production of mycolactone.Journal of Investigative Dermatology (2007) 127, 638–645. doi:10.1038/sj.jid.5700593; published online 19 October 2006 [ABSTRACT FROM AUTHOR]

The evaluation and characterization of epitope-specific human leukocyte antigen (HLA)-restricted memory T-cell reactivity is an important step for the development of preventive vaccines and peptide-based immunotherapies for viral and tumor diseases. The past decade has witnessed the use of HLA-restricted peptides as tools to activate strong immune responses of naïve or memory T cells specifically. This has fuelled an active search for methodological approaches focusing on HLA and peptide associations. Here, we outline new perspective on the emerging opportunity of evaluating HLA and peptide restriction by using novel approaches, such as quantitative real-time PCR, that can identify epitope specificities that are potentially useful in clinical settings. [Copyright &y& Elsevier]

Abstract: Cancer immunotherapy requires the induction of HLA class I restricted cytotoxic T lymphocytes (CTL) specific for tumor associated antigens (TAA). While a number of TAA have been identified, there is an urgent need for the development of adjuvants capable of stimulating CTL responsiveness. Previously, we reported the capacity of immunopotentiating reconstituted influenza virosomes (IRIV) to enhance CTL responses specific for synthetic peptides simultaneously added to cultures in soluble form. This effect was based on IRIV mediated activation of CD4+ T cells. Here we investigated the “in vitro” immunogenicity of a novel virosome formulation coupling in a single reagent the adjuvant power of IRIV to the capacity of liposomes to efficiently encapsulate synthetic peptides. As a model epitope we chose L27Melan-A/Mart-126–35 HLA-A0201restricted peptide from a melanoma-associated antigen widely used in tumor immunotherapy. The reagent thus developed induced the proliferation of CD4+ T cells characterized by a T helper 1 cytokine profile and CXCR3 expression. Most importantly, it significantly enhanced the generation of L27Melan-A/Mart-126–35 specific CTL, as compared to soluble peptides, in particular at low nominal epitope concentrations (<1μg/ml). These effector cells were able to efficiently kill HBL melanoma cells expressing Melan-A/MART-1 and HLA-A0201. The adjuvant effects observed were also detectable in the absence of CD4+ T cells. Taken together our results suggest that this highly immunogenic antigenic formulation might qualify for clinical use in active, antigen-specific, melanoma immunotherapy. [Copyright &y& Elsevier]

Degradation of the tumor antigen epitope gp100280–288 (YLEPGPVTA) was investigated in the presence of cultured human fibroblasts, and acellular supernatants obtained from these cells; the possible effect of substrate degradation on in vitro immunorecognition was also addressed. In the presence of fibroblasts, gp100280–288 was degraded to free amino acids with a half-life of less than 4 min; hydrolysis data support the hypothesis that substrate degradation was mainly caused by the activity of cell-expressed amino- and carboxypeptidases. Gp100280–288 was also degraded in the presence of acellular supernatants: under these conditions, the hydrolysis pattern was similar to that observed in the presence of whole cells, but degradation kinetics was slower. As a result of these phenomena, immunorecognition of gp100280–288-specific cytotoxic T lymphocyte (CTL) clones was severely hampered, and was totally suppressed after 90 min. In conclusion, the high activity of fibroblast-expressed proteases, and the presence of wide-scope soluble enzymes, may explain, at least in part, the low activity of peptide-based antineoplastic vaccines, as well as the transient effectiveness of subcutaneously administered peptides in general. [Copyright &y& Elsevier]

Immunopotentiating reconstituted influenza virosomes (IRIV) are one of the few adjuvants currently licensed for human use. While their adjuvant capacity in the induction of humoral responses is clearly documented, few data exist on their effects on T cell immune response. Here we addressed IRIV adjuvance in the induction of HLA class I restricted cytotoxic T lymphocytes (CTL) in vitro.Lymphocyte stimulation with IM58–66 and IRIV resulted in marked expansion of specific CTL as compared to cultures performed in the presence of either antigen alone or antigen and control liposomes (L). Studies addressing underlying adjuvant mechanisms demonstrated that IRIV activated CD4/CD45RO+ T cells, induced a cytokine profile consistent with T helper 1 (Th1) stimulation and increased the percentage of CD4+ T cells expressing CXCR3. Furthermore, supernatants from IRIV stimulated PBMC cultures promoted dendritic cell maturation. Most importantly, IRIV mediated CTL adjuvance required the presence of live CD4+ T cells. Powerful adjuvant effects of IRIV were also observed in the induction of CTL specific for the melanoma associated Melan-A/MART-127–35, HLA-A0201 restricted epitope.Taken together these findings indicate that IRIV are endowed with a high adjuvant capacity for HLA class I restricted CTL induction, largely attributable to their ability to antigenically stimulate CD4+ T cells. [Copyright &y& Elsevier]

The possible degradation of the tumor antigen epitope gp100280–288 (YLEPGPVTA) in the presence of the monocyte-like line U937, and the effect of degradation on the in vitro-measured immune recognition, were investigated by chromatographic techniques and immunological assays. Results indicate a rapid hydrolysis of the substrate in the presence of the model cells, which is consistent with the hypothesis that degradation of gp100280–288 is caused by the activity of U937-expressed enzymes, specifically amino- and carboxypeptidases. On the other hand, these results do not support the involvement of other enzymes known to be expressed by U937 cells. From a functional point of view, these data indicate that the degradation of gp100280–288 severely hampered recognition by specific CTL clones. The results obtained may provide a model for epitope degradation by the antigen-presenting cells found in defined anatomical compartments and may, at least in part, account for the low activity of peptide-based antineoplastic vaccines, as well as for the transience of the effects of subcutaneously administered peptides in general. [Copyright &y& Elsevier]

Abstract: Objectives: To investigate by immunohistochemistry the expression of MAGE-A and NY-ESO-1/LAGE-1, cancer testis antigens (CTAs), in squamous cell carcinoma of the penis. Methods: A total of 30 penile carcinoma samples from patients undergoing penile amputation at the Urology Clinics at the Zagreb Clinical Hospital Center and University Hospital “Sestre milosrdnice” from 1997 to 2004 were investigated in this study. Three monoclonal antibodies were used for immunohistochemical staining: 77B specific for MAGE-A1, 57B recognizing multiple MAGE-A CTAs, and D8.38, specific for NY-ESO-1 antigen. Results: The expression of MAGE-A1 was not observed in the carcinoma samples, but both multi-MAGE-A and NY-ESO-1-specific reagents stained 29 (97%) of 30 samples. Immunohistochemical staining was prevailingly detected in the cytoplasm. A significant heterogeneity was observed within the same specimen, in which areas with strong positivity coexisted with CTA-negative areas. The extent of CTA expression did not correlate significantly with tumor grade. Conclusions: The results of this study have documented for the first time the expression of CTAs in squamous cell carcinoma of the penis. Additional research is warranted to explore the potential implications regarding both diagnosis and therapy. [Copyright &y& Elsevier]