A long, naturally presented immunodominant epitope from NY-ESO-1 tumor antigen: implications for cancer vaccine design.

The tumor antigen NY-ESO-1 is a promising cancer vaccine target. We describe here a novel HLA-B7-restricted NY-ESO-1 epitope, encompassing amino acids 60-72 (APRGPHGGAASGL), which is naturally presented by melanoma cells. The tumor epitope bound to HLA-B7 by bulging outward from the peptide-binding cleft. This bulged epitope was not an impediment to T-cell recognition, however, because four of six HLA-B7(+) melanoma patients vaccinated with NY-ESO-1 ISCOMATRIX vaccine generated a potent T-cell response to this determinant. Moreover, the response to this epitope was immunodominant in three of these patients and, unlike the T-cell responses to bulged HLA class I viral epitopes, the responding T cells possessed a remarkably broad TCR repertoire. Interestingly, HLA-B7(+) melanoma patients who did not receive the NY-ESO-1 ISCOMATRIX vaccine rarely generated a spontaneous T-cell response to this cryptic epitope, suggesting a lack of priming of such T cells in the natural anti-NY-ESO-1 response, which may be corrected by vaccination. Together, our results reveal several surprising aspects of antitumor immunity and have implications for cancer vaccine design.

Increased expression of monocarboxylate transporters 1, 2, and 4 in colorectal carcinomas.

Tumour cells are known to be highly glycolytic, thus producing high amounts of lactic acid. Monocarboxylate transporters (MCTs), by promoting the efflux of the accumulating acids, constitute one of the most important mechanisms in the maintenance of tumour intracellular pH. Since data concerning MCT expression in colorectal carcinomas (CRC) are scarce and controversial, the present study aimed to assess the expressions of MCT1, 2, and 4 in a well characterized series of CRC and assess their role in CRC carcinogenesis. CRC samples (126 cases) were analyzed for MCT1, MCT2, and MCT4 immunoexpression and findings correlated with clinico-pathological parameters. Expression of all MCT isoforms in tumour cells was significantly increased when compared to adjacent normal epithelium. Remarkably, there was a significant gain of membrane expression for MCT1 and MCT4 and loss of plasma membrane expression for MCT2 in tumour cells. Plasma membrane expression of MCT1 was directly related to the presence of vascular invasion. This is the larger study on MCT expression in CRC and evaluates for the first time its clinico-pathological significance. The increased expression of these transporters suggests an important role in CRC, which might justify their use, especially MCT1 and MCT4, as targets in CRC drug therapy.

Differential requirement for CD4 help in the development of an antigen-specific CD8+ T cell response depending on the route of immunization.

Previous studies in our laboratory have shown that DBA/2 mice injected i.p. with syngeneic P815 tumor cells transfected with the HLA-CW3 gene (P815-CW3) showed a dramatic expansion of activated CD8+CD62L- T cells expressing exclusively the Vbeta10 segment. We have used this model to study the regulatory mechanisms involved in the development of the CW3-specific CD8+ response, with respect to different routes of immunization. Whereas both intradermal (i.d.) and i.p. immunization of DBA/2 mice with P815-CW3 cells led to a strong expansion of CD8+CD62L-Vbeta10+ cells, only the i.d. route allowed this expansion after immunization with P815 cells transfected with a minigene coding for the antigenic epitope CW3 170-179 (P815 miniCW3). Furthermore, depletion of CD4+ T cells in vivo completely abolished the specific response of CD8+CD62L-Vbeta10+ cells and prevented the rejection of P815-CW3 tumor cells injected i.p., whereas it did not affect CD8S+CD62L-Vbeta10+ cell expansion after i.d. immunization with either P815-CW3 or P815 miniCW3. Finally, the CW3-specific CD8+ memory response was identical whether or not CD4+ T cells were depleted during the primary response. Collectively, these results suggest that the CD8+ T cell response to P815-CW3 tumor cells injected i.p. is strictly dependent upon recognition of a helper epitope by CD4+ T cells, whereas no such requirement is observed for i.d. injection.

Reduction in the expression of glucose transporter protein GLUT 2 in preneoplastic and neoplastic hepatic lesions and reexpression of GLUT 1 in late stages of hepatocarcinogenesis.

Expression of two important glucose transporter proteins, GLUT 2 (which is the typical glucose transporter in hepatocytes of adult liver) and the erythroid/brain type glucose transporter GLUT 1 (representing the typical glucose transporter in fetal liver parenchyma), was studied immunocytochemically during hepatocarcinogenesis in rats at different time points between 7 and 65 wk after cessation of 7-wk administration of 12 mg/kg of body weight of N-nitrosomorpholine p.o. (stop model). Foci of altered hepatocytes excessively storing glycogen (GSF) and mixed cell foci (MCF) composed of both glycogenotic and glycogen-poor cells were present at all time points studied. Seven wk after withdrawal of the carcinogen, GSF were the predominant type of focus of altered hepatocytes. Morphometrical evaluation of the focal lesions revealed that the number and volume fraction of GSF increased steadily until Wk 65. MCF were rare at 7 wk, increased slightly in number and size until Wk 37, but showed a pronounced elevation in their number and volume fraction from Wk 37 to Wk 65. In both GSF and MCF, GLUT 2 was generally decreased or partially absent at all time points. Consequently, foci of decreased GLUT 2 expression showed a steady increase in number and volume fraction from Wk 7 to Wk 65. GLUT 1 was lacking in GSF but occurred in some MCF from Wk 50 onward. The liver type glucose transporter GLUT 2 was decreased in all adenomas and hepatocellular carcinomas (HCC). In three of seven adenomas and 10 of 12 carcinomas, expression of GLUT 1 was increased compared with normal liver parenchyma. In two cases of adenoid HCC, cells of ductular formations coexpressed GLUT 2 and GLUT 1. In contrast, normal bile ducts, bile duct proliferations, and cystic cholangiomas expressed only GLUT 1. Seven of 12 HCC contained many microvessels intensely stained for GLUT 1, a phenomenon never observed in normal liver. Whenever adenoid tumor formations occurred, GLUT 1-positive microvessels were located in the immediate vicinity of these formations. Only in one HCC were such microvessels found in the absence of adenoid formations. Our studies indicate that a reduction of GLUT 2 expression occurs already in early preneoplastic hepatic foci and is maintained throughout hepatocarcinogenesis, including benign and malignant neoplasms. Reexpression of GLUT 1, however, appears in a few MCF and in the majority of adenomas and carcinomas.

Carcinoembryonic antigen (CEA): demonstration of a partial identity between CEA and a normal glycoprotein.

The carcinoembryonic antigen of the human digestive tract (CEA), described by Gold as a glycoprotein specific for digestive carcinomatous and foetal tissues, was found to have common antigenic determinants with a glycoprotein of smaller size extracted from normal adult tissues. This observation suggests that only a part of the CEA molecule carries the onco-foetal specificity. It also has practical implications regarding the radioimmunoassay for CEA used for the diagnosis of certain carcinomas.

Decreased local control following radiation therapy alone in early-stage glottic carcinoma with anterior commissure extension.

PURPOSE: To assess the patterns of failure in the treatment of early-stage squamous cell carcinoma of the glottic larynx. PATIENTS AND METHODS: Between 1983-2000, 122 consecutive patients treated for early laryngeal cancer (UICC T1N0 and T2N0) by radical radiation therapy (RT) were retrospectively studied. Male-to-female ratio was 106 : 16, and median age 62 years (35-92 years). There were 68 patients with T1a, 18 with T1b, and 36 with T2 tumors. Diagnosis was made by biopsy in 104 patients, and by laser vaporization or stripping in 18. Treatment planning consisted of three-dimensional (3-D) conformal RT in 49 (40%) patients including nine patients irradiated using arytenoid protection. A median dose of 70 Gy (60-74 Gy) was given (2 Gy/fraction) over a median period of 46 days (21-79 days). Median follow-up period was 85 months. RESULTS: The 5-year overall, cancer-specific, and disease-free survival amounted to 80%, 94%, and 70%, respectively. 5-year local control was 83%. Median time to local recurrence in 19 patients was 13 months (5-58 months). Salvage treatment consisted of surgery in 17 patients (one patient refused salvage and one was inoperable; total laryngectomy in eleven, and partial laryngectomy or cordectomy in six patients). Six patients died because of laryngeal cancer. Univariate analyses revealed that prognostic factors negatively influencing local control were anterior commissure extension, arytenoid protection, and total RT dose < 66 Gy. Among the factors analyzed, multivariate analysis (Cox model) demonstrated that anterior commissure extension, arytenoid protection, and male gender were the worst independent prognostic factors in terms of local control. CONCLUSION: For early-stage laryngeal cancer, outcome after RT is excellent. In case of anterior commissure extension, surgery or higher RT doses are warranted. Because of a high relapse risk, arytenoid protection should not be attempted.

Perinatal arrhythmias.

Cardiac arrhythmias are very frequent in fetuses and newborns. The prognosis depends on the nature of the arrhythmias but is most often either spontaneously benign or following short-term medication administration. A correct diagnosis is essential for both management and prognosis. It is based on echocardiography during the fetal period and mainly on history, physical exam, and electrocardiogram after birth, but other modalities are available to record transient arrhythmic events. Irregular rhythms are mostly benign and rarely require therapy. In most fetuses and infants, tachyarrhythmias resolve spontaneously or require short-term administration of antiarrhythmics. Approximately one third of these may recur later on, especially during adolescence. Persistent bradyarrhythmias might require pacemaker implantation when associated with failure to thrive or with risk of sudden death. CONCLUSION: Arrhythmias in fetuses and infants are very common and mostly benign. History, physical exam, and recording of the arrhythmia are essential to make a correct diagnosis and establish an appropriate management for the rare potentially harmful arrhythmias.

Respiratory motion artifact suppression in diffusion-weighted MR imaging of the spine.

Diffusion-weighted spin-echo imaging of the spine has been successfully implemented for differentiation of benign fracture edema and tumor infiltration of the vertebral body. Nevertheless, this technique still suffers from insufficient image quality in numerous patients due to motion artifacts. The aim of this study was to investigate the impact of variable respiratory motion artifact suppression techniques on image quality in diffusion-weighted spin-echo imaging of the spine. In addition to phase-encoding reordering, a newly implemented right hemi-diaphragmaitc navigator for respiratory gating was used. Subjective and objective image quality parameters were compared. Respiratory motion artifact suppression has a major impact on image quality in diffusion-weighted imaging of the spine. Phase-encoding reordering does not enhance image quality while right hemi-diaphragmatic respiratory navigator gating significantly improves image quality at the cost of data acquisition time. Navigator gating should be used if standard spin-echo diffusion-weighted imaging demonstrates insufficient image quality.

On ubiquitin ligases and cancer.

Protein kinase genes account for almost 10% of all currently known cancer genes, highlighting the role of signal transduction in oncogenesis. A reexamination of the literature and available databases shows that E3 ubiquitin ligases are also key mediators of tumorigenesis. Altogether kinase and E3 genes represent more than 15% of the known cancer genes, underlining the importance of phosphorylation and ubiquitylation signaling pathways in cancer formation. Considering the recent literature reporting correlations between alterations in ubiquitylation processes and oncogenesis, this percentage is likely to increase even further in the future. Finally, E3 genes could serve as baits for the identification of additional cancer genes (e.g. their interacting partners). In contrast, deubiquitinases, like phosphatases, are not overrepresented among cancer genes. The same holds for E1 and E2 genes. Thus, kinase and E3 genes represent primary targets as cancer susceptibility genes for mutation screening and for the design of novel therapies.

Asymptomatic intrapericardial mature teratoma.

Benign mature teratomas are the most common type of germ cell tumors that arise in the anterior mediastinum. Intrapericardial teratomas are a rare manifestation of this tumor type, which are generally diagnosed during infancy because of the heart compression symptoms they produce. Here we report a rare case of intrapericardial mature teratoma that was incidentally discovered in an asymptomatic 51-year-old woman.

Toxicity at three years with and without irradiation of the internal mammary and medial supraclavicular lymph node chain in stage I to III breast cancer (EORTC trial 22922/10925).

INTRODUCTION: The EORTC 22922/10925 trial investigated the potential survival benefit and toxicity of elective irradiation of the internal mammary and medial supraclavicular (IM-MS) nodes Accrual completed in January 2004 and first results are expected in 2012. We present the toxicity reported until year 3 after treatment. PATIENTS AND METHODS: At each visit, toxicity was reported but severity was not graded routinely. Toxicity rates and performance status (PS) changes at three years were compared by chi(2) tests and logistic regression models in all the 3,866 of 4,004 patients eligible to the trial who received the allocated treatment. RESULTS: Only lung (fibrosis; dyspnoea; pneumonitis; any lung toxicities) (4.3% vs. 1.3%; p < 0.0001) but not cardiac toxicity (0.3% vs. 0.4%; p = 0.55) significantly increased with IM-MS treatment. No significant worsening of the PS was observed (p = 0.79), suggesting that treatment-related toxicity does not impair patient's daily activities. CONCLUSIONS: IM-MS irradiation seems well tolerated and does not significantly impair WHO PS at three years. A follow-up period of at least 10 years is needed to determine whether cardiac toxicity is increased after radiotherapy.

Comparison of copper-67- and iodine-125-labeled anti-CEA monoclonal antibody biodistribution in patients with colorectal tumors.

Copper-67 has comparable beta-particle emissions to that of 131I, but it displays more favorable gamma emission characteristics for application in radioimmunotherapy (RIT). This study investigates the potential of 67Cu-labeled monoclonal antibody (MAb) 35 for RIT of colorectal carcinoma. METHODS: Biokinetics of simultaneously injected 67Cu- and 125I-labeled MAb35 were studied in six patients scheduled for surgery of primary colorectal cancer. RESULTS: Whole-body clearance (T 1/2) of 67Cu, estimated from sequential anterior and posterior whole-body scans and corrected for decay of 67Cu, was 41 hr. Serum clearance of 67Cu was faster (27.41 hr) than that of 125I (38.33 hr). Mean tumor uptake of the 67Cu-labeled compound (0.0133% ID/g) exceeded that of 125I (0.0095% ID/g), and tumor-to-blood ratios were higher for 67Cu than for 125I, with averages of 6.07 and 2.41, respectively. The average 67Cu/125I ratio was 1.9 for tumor uptake, 0.7 for blood and 2.6 for tumor-to-blood ratios. Nonspecific liver uptake of 67Cu as calculated from whole-body scans was high in four patients, up to 25% of residual whole-body activity at 48 hr, but did not increase with time. We also observed some nonspecific bowel activity, as well as moderate to high uptake in benign polyps. CONCLUSION: Copper-67-labeled MAb35 is more favorable than its radioiodine-labeled counterpart for RIT of colorectal carcinoma due to higher tumor-to-blood ratios, but the problem of nonspecific liver and bowel uptake must first be overcome. The absolute accumulation of activity in tumor remains low, however, so the probability of cure with this compound alone is questionable. The use of 67Cu as one component of a multimodality adjuvant treatment seems to remain the most appropriate application for RIT.

Gemcitabine, oxaliplatin and 5-FU in advanced bile duct and gallbladder carcinoma: two parallel, multicentre phase-II trials.

BACKGROUND: Gemcitabine, oxaliplatin and 5-fluorouracil (5-FU) are active in biliary tract cancer and have a potentially synergistic mode of action and non-overlapping toxicity. The objective of these trials was to determine response, survival and toxicity separately in patients with bile duct cancer (BDC) and gallbladder cancer (GBC) treated with gemcitabine/oxaliplatin/5-FU chemotherapy. METHODS: Eligible patients with histologically proven, advanced or metastatic BDC (n=37) or GBC (n=35) were treated with gemcitabine (900 mg m(-2) over 30 min), oxaliplatin (65 mg m(-2)) and 5-FU (1500 mg m(-2) over 24 h) on days 1 and 8 of a 21-day cycle. Tumour response was the primary outcome measure. RESULTS: Response rates were 19% (95% CI: 6-32%) and 23% (95% CI: 9-37%) for BDC and GBC, respectively. Median survivals were 10.0 months (95% CI: 8.6-12.4) and 9.9 months (95% CI: 7.5-12.2) for BDC and GBC, respectively, and 1- and 2-year survival rates were 40 and 23% in BDC and 34 and 6% in GBC (intention-to-treat analysis). Major grade III and IV adverse events were neutropenia, thrombocytopenia, elevated bilirubin and anorexia. CONCLUSION: Triple-drug chemotherapy achieves comparable results for response and survival to previously reported regimens, but with more toxicity.

Germline mutations in retinoma patients: relevance to low-penetrance and low-expressivity molecular basis.

PURPOSE: To study phenotype-genotype correlation in patients who have retinoma, which is a benign tumor resembling the post irradiation regression pattern of retinoblastoma (RB). METHODS: We selected patients who had retinoma and positive family history for RB and patients who had retinoma in one eye and either retinoma or RB in the other eye. The study included 22 patients with available DNA: 18 from 11 families and four sporadic cases. DNA was extracted from peripheral blood leukocytes. The RB1 gene was screened by DHPLC and direct sequencing of the promoter and all the exons. RESULTS: We identified 17 occurrences of 11 distinct germline mutations in two sporadic and in 15 familial cases (nine families). The 11 identified mutations were located in exons 1, 10,11,13,14, and 19 to 23. Four of the identified mutations were not previously reported, including g.64407delT, g.153236A>T, g.156743delTCTG, and g.162078delA. Eight out the 11 mutations were truncating and three were nontruncating (missense). There was no correlation between the type of mutation and the number of tumor foci per eye (RB or retinomas). Highly heterogeneous intrafamilial expressivity was observed. CONCLUSIONS: To our knowledge, this study is the largest series of mutations of consecutive retinoma patients. The present data suggest that the type of inherited mutations underlying retinoma is undistinguishable from RB related ones, i.e., largely dominated by truncating mutants. This finding is in contrast with the RB1 genotypic spectrum of mutations associated with low-penetrance RB, i.e., nontruncating mutants. The molecular mechanism underlying low-penetrance and attenuated expressivity (retinomas) appeared to be distinct.

Melan-A-specific cytotoxic T cells are associated with tumor regression and autoimmunity following treatment with anti-CTLA-4.

PURPOSE: Ipilimumab is a monoclonal antibody that blocks the immune-inhibitory interaction between CTL antigen 4 (CTLA-4) and its ligands on T cells. Clinical trials in cancer patients with ipilimumab have shown promising antitumor activity, particularly in patients with advanced melanoma. Often, tumor regressions in these patients are correlated with immune-related side effects such as dermatitis, enterocolitis, and hypophysitis. Although these reactions are believed to be immune-mediated, the antigenic targets for the cellular or humoral immune response are not known. EXPERIMENTAL DESIGN: We enrolled patients with advanced melanoma in a phase II study with ipilimumab. One of these patients experienced a complete remission of his tumor. The specificity and functional properties of CD8-positive T cells in his peripheral blood, in regressing tumor tissue, and at the site of an immune-mediated skin rash were investigated. RESULTS: Regressing tumor tissue was infiltrated with CD8-positive T cells, a high proportion of which were specific for Melan-A. The skin rash was similarly infiltrated with Melan-A-specific CD8-positive T cells, and a dramatic (>30-fold) increase in Melan-A-specific CD8-positive T cells was apparent in peripheral blood. These cells had an effector phenotype and lysed Melan-A-expressing tumor cells. CONCLUSIONS: Our results show that Melan-A may be a major target for both the autoimmune and antitumor reactions in patients treated with anti-CTLA-4, and describe for the first time the antigen specificity of CD8-positive T cells that mediate tumor rejection in a patient undergoing treatment with an anti-CTLA-4 antibody. These findings may allow a better integration of ipilimumab into other forms of immunotherapy.