Cancer-malaria: hidden connections.

Cancer and malaria exemplify two maladies historically assigned to separated research spaces. Cancer, on the one hand, ranks among the top priorities in the research agenda of developed countries. Its rise is mostly explained by the ageing of these populations and linked to environment and lifestyle. Malaria, on the other hand, represents a major health burden for developing countries in the Southern Hemisphere. These two diseases also belong to separate fields of medicine: non-communicable diseases for cancer and communicable diseases for malaria.

Prognostic value of serum CYFRA 21-1 1 in patients with anal canal squamous cell carcinoma treated with radio(chemo)therapy.

BACKGROUND: We aimed to assess the prognostic value of CYFRA 21-1 in a series of patients with anal canal squamous cell carcinoma treated by radiation-based therapy. METHODS: All patients with anal cancer referred between September 2005 and July 2013 were considered. Patients with diagnosis of anal squamous cell carcinoma and in whom pre- and post-treatment serum CYFRA 21-1 levels were available were included. Serum CYFRA 21-1 levels at initial workup and after therapy were collected. Survival rates were estimated using the Kaplan-Meier method. Cox regression analysis was used to evaluate prognostic variables for prediction of outcomes. RESULTS: Eighty-two patients were included. Median follow-up was 60 months (range: 8-128). Pre-treatment serum CYFRA 21-1 levels were significantly correlated with tumour stage (p < 0.001). Normal post-treatment serum CYFRA 21-1 level was significantly correlated with tumour complete response (p = 0.004). Elevated post-treatment serum CYFRA 21-1 level was significantly associated with poorer progression-free survival (p = 0.02) and overall survival (p = 0.003). T stage and post-treatment serum CYFRA 21-1 were independent prognostic factors for overall survival (p = 0.04 and 0.03, respectively). CONCLUSIONS: Serum CYFRA 21-1 appears to be a useful marker for the monitoring of anal squamous cell carcinoma patients. Elevated post-treatment value appears to be correlated with treatment failure.

The early pregnancy placenta foreshadows DNA methylation alterations of solid tumors.

The placenta relies on phenotypes that are characteristic of cancer to successfully implant the embryo in the uterus during early pregnancy. Notably, it has to invade its host tissues, promote angiogenesis-while surviving hypoxia-, and escape the immune system. Similarities in DNA methylation patterns between the placenta and cancers suggest that common epigenetic mechanisms may be involved in regulating these behaviors. We show here that megabase-scale patterns of hypomethylation distinguish first from third trimester chorionic villi in the placenta, and that these patterns mirror those that distinguish many tumors from corresponding normal tissues. We confirmed these findings in villous cytotrophoblasts isolated from the placenta and identified a time window at the end of the first trimester, when these cells come into contact with maternal blood, as the likely time period for the methylome alterations. Furthermore, the large genomic regions affected by these patterns of hypomethylation encompass genes involved in pathways related to epithelial-mesenchymal transition, immune response, and inflammation. Analyses of expression profiles corresponding to genes in these hypomethylated regions in colon adenocarcinoma tumors point to networks of differentially expressed genes previously implicated in carcinogenesis and placentogenesis, where nuclear factor kappa B is a key hub. Taken together, our results suggest the existence of epigenetic switches involving large-scale changes of methylation in the placenta during pregnancy and in tumors during neoplastic transformation. The characterization of such epigenetic switches might lead to the identification of biomarkers and drug targets in oncology as well as in obstetrics and gynecology.

Circulating human papillomavirus DNA detected using droplet digital PCR in the serum of patients diagnosed with early stage human papillomavirus-associated invasive carcinoma.

Specific human papillomavirus genotypes are associated with most ano-genital carcinomas and a large subset of oro-pharyngeal carcinomas. Human papillomavirus DNA is thus a tumour marker that can be detected in the blood of patients for clinical monitoring. However, data concerning circulating human papillomavirus DNA in cervical cancer patients has provided little clinical value, due to insufficient sensitivity of the assays used for the detection of small sized tumours. Here we took advantage of the sensitive droplet digital PCR method to identify circulating human papillomavirus DNA in patients with human papillomavirus-associated carcinomas. A series of 70 serum specimens, taken at the time of diagnosis, between 2002 and 2013, were retrospectively analyzed in patients with human papillomavirus-16 or human papillomavirus-18-associated carcinomas, composed of 47 cases from the uterine cervix, 15 from the anal canal and 8 from the oro-pharynx. As negative controls, 18 serum samples from women with human papillomavirus-16-associated high-grade cervical intraepithelial neoplasia were also analyzed. Serum samples were stored at -80°C (27 cases) or at -20°C (43 cases). DNA was isolated from 200 µl of serum or plasma and droplet digital PCR was performed using human papillomavirus-16 E7 and human papillomavirus-18 E7 specific primers. Circulating human papillomavirus DNA was detected in 61/70 (87%) serum samples from patients with carcinoma and in no serum from patients with cervical intraepithelial neoplasia. The positivity rate increased to 93% when using only serum stored at -80°C. Importantly, the two patients with microinvasive carcinomas in this series were positive. Quantitative evaluation showed that circulating viral DNA levels in cervical cancer patients were related to the clinical stage and tumour size, ranging from 55 ± 85 copies/ml (stage I) to 1774 ± 3676 copies/ml (stage IV). Circulating human papillomavirus DNA is present in patients with human papillomavirus-associated invasive cancers even at sub-clinical stages and its level is related to tumour dynamics. Droplet digital PCR is a promising method for circulating human papillomavirus DNA detection and quantification. No positivity was found in patients with human papillomavirus-associated high grade cervical intraepithelial neoplasia.

Serum CA125 and HE4 levels as predictors for optimal interval surgery and platinum sensitivity after neoadjuvant platinum-based chemotherapy in patients with advanced epithelial ovarian cancer.

BACKGROUND: The aim of this study is to evaluate a new tumour marker, HE4, and to compare it with CA125 in predicting optimal cytoreduction and response to chemotherapy. Thirty patients with advanced epithelial ovarian cancer and multiple sera harvested during neoadjuvant chemotherapy (NAC) were included. RESULTS: Based on ROC curves analysis, CA125 ≤ 75 UI/ml and HE4 ≤ 252 pmol/L after the 3rd cycles of NAC, with a sensitivity of 93.7 % and a specificity of 92.3 % (PPV = 93.7 % and NPV = 92.3 %), offered the best combination for predicting optimal cytoreduction. In addition, the HE4 value of 115 pmol/L is the best cut-off level for identifying platinum-sensitive patients. CONCLUSIONS: The introduction of HE4 as a new tool for predicting platinum-sensitivity and interval optimal cytoreduction is promising.

Spherical cancer models in tumor biology.

Three-dimensional (3D) in vitro models have been used in cancer research as an intermediate model between in vitro cancer cell line cultures and in vivo tumor. Spherical cancer models represent major 3D in vitro models that have been described over the past 4 decades. These models have gained popularity in cancer stem cell research using tumorospheres. Thus, it is crucial to define and clarify the different spherical cancer models thus far described. Here, we focus on in vitro multicellular spheres used in cancer research. All these spherelike structures are characterized by their well-rounded shape, the presence of cancer cells, and their capacity to be maintained as free-floating cultures. We propose a rational classification of the four most commonly used spherical cancer models in cancer research based on culture methods for obtaining them and on subsequent differences in sphere biology: the multicellular tumor spheroid model, first described in the early 70s and obtained by culture of cancer cell lines under nonadherent conditions; tumorospheres, a model of cancer stem cell expansion established in a serum-free medium supplemented with growth factors; tissue-derived tumor spheres and organotypic multicellular spheroids, obtained by tumor tissue mechanical dissociation and cutting. In addition, we describe their applications to and interest in cancer research; in particular, we describe their contribution to chemoresistance, radioresistance, tumorigenicity, and invasion and migration studies. Although these models share a common 3D conformation, each displays its own intrinsic properties. Therefore, the most relevant spherical cancer model must be carefully selected, as a function of the study aim and cancer type.

¹8F-FDG PET/CT imaging versus dynamic contrast-enhanced CT for staging and prognosis of inflammatory breast cancer.

PURPOSE: Inflammatory breast cancer (IBC) is the most aggressive type of breast cancer with a poor prognosis. Locoregional staging is based on dynamic contrast-enhanced (DCE) CT or MRI. The aim of this study was to compare the performances of FDG PET/CT and DCE CT in locoregional staging of IBC and to assess their respective prognostic values. METHODS: The study group comprised 50 women (median age: 51 ± 11 years) followed in our institution for IBC who underwent FDG PET/CT and DCE CT scans (median interval 5 ± 9 days). CT enhancement parameters were net maximal enhancement, net early enhancement and perfusion. RESULTS: The PET/CT scans showed intense FDG uptake in all primary tumours. Concordance rate between PET/CT and DCE CT for breast tumour localization was 92%. No significant correlation was found between SUVmax and CT enhancement parameters in primary tumours (p > 0.6). PET/CT and DCE CT results were poorly correlated for skin infiltration (kappa = 0.19). Ipsilateral foci of increased axillary FDG uptake were found in 47 patients (median SUV: 7.9 ± 5.4), whereas enlarged axillary lymph nodes were observed on DCE CT in 43 patients. Results for axillary node involvement were fairly well correlated (kappa = 0.55). Nineteen patients (38%) were found to be metastatic on PET/CT scan with a significant shorter progression-free survival than patients without distant lesions (p = 0.01). In the primary tumour, no statistically significant difference was observed between high and moderate tumour FDG uptake on survival, using an SUVmax cut-off of 5 (p = 0.7 and 0.9), or between high and low tumour enhancement on DCE CT (p > 0.8). CONCLUSION: FDG PET/CT imaging provided additional information concerning locoregional involvement to that provided by DCE CT on and allowed detection of distant metastases in the same whole-body procedure. Tumour FDG uptake or CT enhancement parameters were not correlated and were not found to have any prognostic value.

Immuno-SPET/CT and immuno-PET/CT: a step ahead to translational imaging.

Malignant tumours have the remarkable property to express cell surface antigens. Pressman was first reporting that radiolabeled antibodies were capable of organ localization. It was a promising challenge but the expected success and the development of this imaging method was limited by a poor imaging resolution despite a rather good specificity of the antibodies used. Identification of key cell surface markers is opening a new era as potential molecular imaging biomarkers in oncologic applications. Antibodies production has been promoted by the development of engineered fragments with preserved immunological properties and pharmacokinetics optimized for molecular imaging. A good compromise has to be obtained between the biological properties of the antibody and the physical half-life of the radionuclide. Several positron emission tomography (PET) radionuclides such as iodine-124, copper-64, yttrium-86 or zirconium-89 have been the focus of recent immuno-PET studies with interesting informative images in preclinical and clinical studies. Thanks to the development of more sensitive new detectors and specific software, molecular imaging methods, particularly PET imaging, allow nowadays the detection of lesions smaller than 5 mm in human. Immuno-PET can potentially be used for tumour detection and identification at diagnosis, staging and restaging, for treatment selection and monitoring, and during follow-up. Moreover the availability of matched imaging or therapeutic radionuclide pairs, such as (124)I/(131)I, (64)Cu/(67)Cu and (86)Y/(90)Y, make easier the quantification of tissue uptake and dosimetry calculation for radioimmunotherapy.

Assessment of response to endocrine therapy using FDG PET/CT in metastatic breast cancer: a pilot study.

PURPOSE: The purpose of this pilot study was to assess whether outcome in metastatic or recurrent breast cancer patients is related to metabolic response to endocrine therapy determined by (18)F-FDG PET/CT. METHODS: The study group comprised 22 patients with breast cancer (age 58 ± 11 years, mean ± SD) who were scheduled to receive endocrine therapy. They were systematically assessed by PET/CT at baseline and after a mean of 10 ± 4 weeks for evaluation of response after induction. All patients demonstrated FDG-avid lesions on the baseline PET/CT scan. The metabolic response was assessed according to EORTC criteria and based on the mean difference in SUV(max) between the two PET/CT scans, and the patients were classified into four groups: complete or partial metabolic response, or stable or progressive metabolic disease (CMR, PMR, SMD and PMD, respectively). All patients were followed in our institution. RESULTS: Metastatic sites were localized in bone (n = 15), lymph nodes (n = 11), chest wall (n = 3), breast (n = 5), lung (n = 3), soft tissue (n = 1) and liver (n = 1). PMR was observed in 11 patients (50%), SMD in 5 (23%) and PMD in 6 (27%). The median progression-free survival (PFS) times were 20, 27 and 6 months in the PMR, SMD and PMD groups, respectively. PFS in the SMD group differed from that in the PMR and SMD groups (p < 0.0001). CONCLUSION: Metabolic response assessed by FDG PET/CT imaging in patients with metastatic breast cancer treated with endocrine therapy is predictive of the patients' PFS.

Breast cancer recurrence diagnosis suspected on tumor marker rising: value of whole-body 18FDG-PET/CT imaging and impact on patient management.

BACKGROUND: Breast cancer recurrence is often suspected on tumor marker rising in asymptomatic patients. The value of fluorine-18 fluorodeoxyglucose (18FDG)-positron emission tomography/computed tomography (PET/CT) imaging to detect recurrence and its subsequent impact on patient management were retrospectively assessed. METHODS: PET/CT scans were performed on 228 asymptomatic patients (mean, 60.8 years; range, 30-91 years) presenting with rising CA 15-3 and/or CEA serum levels. RESULTS: PET/CT scans were positive in 181 patients (79.5%) and normal in 47 patients, whereas 187 true recurrences were diagnosed. Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of PET/CT imaging for detection of breast cancer recurrence were 93.6%, 85.4%, 96.7%, 74.5%, and 92.1%, respectively. When compared with the standard workup available in 67 patients, PET/CT imaging had a higher sensitivity and accuracy (94.5% vs 33% and 94% vs 48%, respectively). Recurrences were confirmed by pathology, conventional imaging techniques, or radiological and clinical follow-up beyond 1 year (mean, 34 months; range, 12-67 years) in 32, 130, and 25 patients, respectively. The diagnosis of recurrence led to a treatment modification in 123 patients (54%). CONCLUSIONS: 18FDG-PET/CT imaging is an efficient technique to detect breast cancer recurrence suspected on tumor marker rising in asymptomatic patients. It may thus contribute to improve patient management, providing an earlier diagnosis with complete whole-body staging as a "one-stop shop" procedure.

Single photon emission tomography/computed tomography (SPET/CT) and positron emission tomography/computed tomography (PET/CT) to image cancer.

Hybrid systems associating the sharpness of anatomic images coming from computed tomography (CT) and radionuclide functional imaging (SPET or PET) are opening a new era in oncology. This multimodal imaging method is now routinely used for the diagnosis, extent, follow up, treatment response and detection of occult disease in different types of malignancies with a significant impact on the treatment strategy leading for a change for more than 68% of all investigated patients.

In situ protein expression in tumour spheres: development of an immunostaining protocol for confocal microscopy.

BACKGROUND: Multicellular tumour sphere models have been shown to closely mimic phenotype characteristics of in vivo solid tumours, or to allow in vitro propagation of cancer stem cells (CSCs). CSCs are usually characterized by the expression of specific membrane markers using flow cytometry (FC) after enzymatic dissociation. Consequently, the spatial location of positive cells within spheres is not documented. Confocal microscopy is the best technique for the imaging of thick biological specimens after multi-labelling but suffers from poor antibody penetration. Thus, we describe here a new protocol for in situ confocal imaging of protein expression in intact spheroids. METHODS: Protein expression in whole spheroids (150 mum in diameter) from two human colon cancer cell lines, HT29 and CT320X6, has been investigated with confocal immunostaining, then compared with profiles obtained through paraffin immunohistochemistry (pIHC) and FC. Target antigens, relevant for colon cancer and with different expression patterns, have been studied. RESULTS: We first demonstrate that our procedure overcomes the well-known problem of antibody penetration in compact structures by performing immunostaining of EpCAM, a membrane protein expressed by all cells within our spheroids. EpCAM expression is detected in all cells, even the deepest ones. Likewise, antibody access is confirmed with CK20 and CD44 immunostaining. Confocal imaging shows that 100% of cells express beta-catenin, mainly present in the plasma membrane with also cytoplasmic and nuclear staining, in agreement with FC and pIHC data. pIHC and confocal imaging show similar CA 19-9 cytoplasmic and membranar expression profile in a cell subpopulation. CA 19-9+ cell count confirms confocal imaging as a highly sensitive method (75%, 62% and 51%, for FC, confocal imaging and pIHC, respectively). Finally, confocal imaging reveals that the weak expression of CD133, a putative colon CSC marker, is restricted to the luminal cell surface of colorectal cancer acini, with CD133+ cellular debris into glandular lumina. CONCLUSION: The present protocol enables in situ visualization of protein expression in compact three-dimensional models by whole mount confocal imaging, allowing the accurate localization and quantification of cells expressing specific markers. It should prove useful to study rare events like CSCs within tumour spheres.

Inflammation in reproductive disorders.

Inflammatory disorders account for a significant percentage of gynecologic disease, particularly in reproductive age women. Inflammation is a basic method by which we respond to infection, irritation, or injury. Inflammation is now recognized as a type of nonspecific immune response, either acute or chronic. In gynecology, inflammation leads to anatomic disorders primarily as a result of infectious disease; however inflammation can affect ovulation and hormone production as well as be associated with endometriosis. Similarly, immune cell trafficking is an important component of cyclic endometrial development in each menstrual cycle. These immune cells are required for endometrial function, producing a vast array of inflammatory cytokines. Inflammation alters endometrial receptivity, however it may also play a role in tissue repair and remodeling. Finally, inflammation affects the trophoblast and trophoblast-endometrial interaction. Some components of the immune response are required for optimal fertility and normal tissue remodeling. A better understanding of the necessary role of inflammation in reproduction will allow more rational and targeted treatment of inflammatory disorders in reproductive medicine.

Establishment of human colon cancer cell lines from fresh tumors versus xenografts: comparison of success rate and cell line features.

Obtaining representative human colon cancer cell lines from fresh tumors is technically difficult. Using 32 tumor fragments from patients with colon cancer, the present study shows that prior xenograft leads to more efficient cell line establishment compared with direct establishment from fresh tumors (P < 0.05). From 26 tumor specimens, we successfully established 20 tumor xenografts in nude mice (77%); among 19 of these xenografts, 9 (47%) led to cell lines, including four from liver metastases. Only 3 of 31 tumor specimens (9.7%) grew immediately in vitro, and all were derived from primary tumors. To compare major phenotypic and genotypic characteristics of human colon cancer cell lines derived from the same tumor fragment using two protocols, the two pairs of cell lines obtained from 2 of 32 tumor fragments were extensively studied. They displayed similar morphology and were able to form compact spheroids. Chemosensitivity to 5-fluorouracil, CPT11, and L-OHP differed between cell lines obtained from patient tumors and those derived from xenografts. Matched cell lines shared a common core of karyotype alterations and distinctive additional chromosomal aberrations. Expression levels of genes selected for their role in oncogenesis evaluated by real-time quantitative PCR were found to be statistically correlated whatever the in vitro culture model used. In conclusion, xenotransplantation in mice of tumor fragments before establishment of cell lines enables generation of more novel human cancer cell lines for investigation of colon cancer cell biology, opening up the opportunity of reproducing the diversity of this disease.

[Humanized antibodies as therapeutics]. / Anticorps humanisés en thérapeutique.

Since 1997, nine humanized antibodies received the approval of the FDA to be used as drugs for the treatment of various diseases including transplant rejections, metastatic breast and colon cancers, leukaemia, non-Hodgkin lymphomas, allergic conditions or multiple sclerosis. This review describes techniques used to engineer these antibodies and presents the recent evolutions of these techniques : SDRs grafting or << abbreviated >> CDRs grafting. Based on the illustrative examples of several antibodies, Mylotarg, Herceptin or Xolair, the therapeutic effectiveness of humanized antibodies are underlined and, with the example of Tysabri, the sometimes dramatic adverse effects associated with their clinical use is stressed. In a second part, this review presents some future and realistic avenues to improve the effectiveness of the humanized antibodies, to decrease their immunogenicity and to reduce their cost.

Development and validation of a real-time PCR assay for the detection and quantitation of p53 recombinant adenovirus in clinical samples from patients treated with Ad5CMV-p53 (INGN 201).

The purpose of this study was to assess the usefulness of real-time PCR as a quantitative, highly reproducible, and sensitive method, for detecting and quantifying p53 recombinant adenovirus in biological samples from cancer patients receiving injections of Ad5CMV-p53. The dynamic range of this real-time PCR-based assay was wide (at least five orders of magnitude). Our assay used an internal positive control in the same PCR tube that is capable of detecting residual PCR inhibitors. Serial spiked samples in plasma with known quantities of Ad5CMV-p53 were evaluated. The minimum detection limit was 2 pfu per PCR (approximately 50 pfu per ml of plasma) and the quantification values were reproducible. A total of 2069 controls tested with 1780 plasma samples from 286 patients enrolled in gene therapy trials using Ad5CMV-p53 were investigated. Using calibrators to adjust the quantitation value, the results confirmed the good performance of the assay. In conclusion, the high sensitivity, simplicity and reproducibility of the real-time Ad5CMV-p53 assay, allowing screening of large numbers of samples, combined with its wide dynamic range, make this method particularly suitable for monitoring gene therapy trials.

A three-dimensional tumor cell defect in activating autologous CTLs is associated with inefficient antigen presentation correlated with heat shock protein-70 down-regulation.

We described previously a CTL clone able to lyse the autologous carcinoma cell line IGR-Heu after specific recognition of an HLA-A2/mutated alpha-actinin-4 peptide complex. Here, we used IGR-Heu, cultured either as standard two-dimensional monolayers or as three-dimensional spheroids, to further analyze the influence of target architecture on CTL reactivity. Interestingly, we found that changes in the tumor structure from two- to three-dimensional induced a dramatic decrease in its capacity to activate autologous CTL, as measured by IFN-gamma and tumor necrosis factor-alpha secretion. These functional alterations were attributable neither to MHC class I expression nor to tumor antigen (Ag) down-regulation, because IGR-Heu, cultured as two- or three-dimensional, expressed similar levels of HLA-A2 and alpha-actinin-4. More importantly, incubation of three-dimensional cells with synthetic epitope completely restored cytokine release by CTL. This defective Ag presentation correlated with a decrease in heat shock protein (hsp)70 expression by three-dimensional tumors compared with two-dimensional cells. Furthermore, transfection of the tumor cells with hsp70 cDNA completely restored the Ag-presenting potential of spheroids and, therefore, cytokine production by T cells. These data strongly suggest that hsp70 down-regulation in three-dimensional cells may result in tumor resistance to the immune response.

Transcriptional expression of genes involved in cell invasion and migration by normal and tumoral trophoblast cells.

Once initiated, invasion of trophoblast cells must be tightly regulated, particularly in early pregnancy. The mechanisms necessary for the invasion and migration of trophoblast cells are thought to be related to those involved in the invasive and metastatic properties of cancer cells. Quantitative PCR was used to measure, in trophoblast cells, the transcriptional expression profiles of four genes, INSL4, BRMS1, KiSS-1 and KiSS-1R, reported to be implicated in tumor invasion and metastasis. Laser capture microdissection and purification of trophoblast cells demonstrate that, as already known for INSL4, BRMS1, KiSS-1 and KiSS-1R are expressed by the trophoblast subset of placental tissues. Expression profiles of these genes studied in early placentas (7-9 weeks, n=55) and term placentas (n=11) showed that expression levels of BRMS1 are higher in term than in early placentas, while expression levels of KiSS-1R are higher in early than in term placentas. Low levels of expression of BRMS1 were observed in normal pregnancies, in molar pregnancies and in choriocarcinoma cell lines BeWo, JAR and JEG3 while, in striking contrast, the expression levels of INSL4, KiSS-1 and Kiss-1R were increased in both early placentas and molar pregnancies and were reduced in choriocarcinoma cells. These transcriptional expression profiles are in favor of a predominant role of INSL4, KiSS-1 and KiSS-1R in the control of the invasive and migratory properties of trophoblast cells.

Tumor-associated antigen human chorionic gonadotropin beta contains numerous antigenic determinants recognized by in vitro-induced CD8+ and CD4+ T lymphocytes.

The beta subunit of human chorionic gonadotropin (hCG beta) is markedly overexpressed by neoplastic cells of differing histological origin including those present in colon, breast, prostate and bladder tumors. We have previously shown that some patients with hCG beta-producing urothelial tumors have circulating T cells that proliferate in response to hCG beta. To make a comprehensive study of hCG beta as a potential target for cancer immunotherapy, we investigated whether hCG beta peptides could induce CD4+ or CD8+ T-cell responses in vitro. By stimulating peripheral blood mononuclear cells (PBMCs) from three donors with mixtures of overlapping 16-mer synthetic peptides analogous to portions of either the hCG beta 20-71 or the hCG beta 102-129 region, we established six CD4+ T-cell lines that proliferated specifically in response to five distinct determinants located within these two hCG beta regions. Three antigenic determinants (hCG beta 52-67, 106-121 and 114-125) were presented by HLA-DR molecules, while the two other antigenic determinants (hCG beta 48-63 and 56-67) were presented by HLA-DQ molecules. Interestingly, one T-cell line specific for peptide hCG beta 106-121 recognized hCG beta peptides comprising, at position 117, either an alanine or an aspartic acid residue, with the latter residue being present within the protein expressed by some tumor cells. In addition, three other hCG beta-derived peptides that exhibited HLA-A*0201 binding ability were able to stimulate CD8+ cytotoxic T cells from two HLA-A*0201 donors. These three immunogenic peptides corresponded to regions hCG beta 40-48, hCG beta 44-52 and hCG beta 75-84. Our results indicate that the tumor-associated antigen hCG beta possesses numerous antigenic determinants liable to stimulate CD4+ and CD8+ T lymphocytes, and might thus be an effective target antigen for the immunotherapy of hCG beta-producing tumors.