Immunoreactivity of the 14F7 Mab raised against N-Glycolyl GM3 Ganglioside in retinoblastoma tumours.

INTRODUCTION: The identification of molecules expressed selectively on the surface of retinoblastoma cells would allow applying targeted therapies. The Ganglioside, N-Glycolyl-GM3 (NeuGc-GM3), is an attractive candidate, as it has been detected in other paediatric neuroectodermic tumours, and it is not expressed in human normal tissues. The 14F7 antibody recognizes specifically the ganglioside NeuGc-GM3. PURPOSE: To characterize the expression of NeuGc-GM3 in retinoblastoma cell lines and in retinoblastoma tumours using the 14F7 monoclonal antibody. METHODS: We studied WERI-Rb1 and Y79 cell lines, 24 retinoblastoma primary tumours from unilateral and bilateral cases and two bone marrow biopsies from metastatic retinoblastoma. Tumours were classified into three groups: non-invasive (n = 13), invasive (n = 9) and metastatic (n = 2). Three eyes enucleated because of non-tumoural conditions were used as controls. Cell lines and tumour sections were studied by immunohistochemistry using the 14F7 antibody. NeuGc-GM3 expression was evaluated by analysing the percentage of positive tumoural cells and the staining intensity. These parameters were analysed comparatively among the three groups. RESULTS: Both retinoblastoma cell lines showed immunoreactivity to NeuGc-GM3 but WERI-Rb1 presented higher intensity than Y79. All the tumours studied showed strong immunoreactivity to NeuGc-GM3 with no significant differences among groups. In both bone marrow specimens, NeuGc-GM3 immunoreactivity was observed in retinoblastoma cells. In bilaterally enucleated cases, NeuGc-GM3 immunoreactivity was not altered before and after chemotherapy. Non-tumoural retinas were negative. CONCLUSIONS: NeuGc-GM3 is highly expressed in retinoblastoma cell lines, tumours and metastatic cells to the bone marrow, and it is not detectable in control eyes. There were no significant differences in the immunoreactivity to 14F7 among tumours from different disease stages. Its immunoreactivity did not change after chemotherapy.

Insight into the effect of the vasopressin analog desmopressin on lung colonization by mammary carcinoma cells in BALB/c mice.

BACKGROUND/AIM: Desmopressin (dDAVP) is a synthetic peptide analog of vasopressin with antidiuretic and hemostatic properties. Recent experimental evidence have suggested that dDAVP can inhibit metastasis formation by agonist action on V2 vasopressin receptors present in both tumor and endothelial cells. We have examined the kinetics of dDAVP effect during metastatic colonization and its potential association with hemostasis. MATERIALS AND METHODS: The experimental metastasis assay was performed by injecting F3II mammary carcinoma cells into the lateral tail vein of syngeneic female BALB/c mice. RESULTS: Clinically relevant doses of dDAVP (0.3 to 2 µg/kg intravenously (i.v.)) produced a dose-dependent inhibition in the formation of lung nodules when administered during the first 24 hours after F3II tumor cell injection. The hemostatic agent tranexamic acid (10 mg/kg, i.v.) had no effect on metastasis formation in the same experimental conditions, while the anticoagulant enoxaparin (1 mg/kg, subcutaneously (s.c.)) did not modify the antimetastatic action of dDAVP. In vitro, dDAVP had a strong inhibitory effect on F3II cell colony formation. CONCLUSION: dDAVP interferes with early metastatic disease, and direct association of this effect with hemostatic mechanisms is unlikely.

Detection of N-glycolyl GM3 ganglioside in neuroectodermal tumors by immunohistochemistry: an attractive vaccine target for aggressive pediatric cancer.

The N-glycolylated ganglioside NeuGc-GM3 has been described in solid tumors such as breast carcinoma, nonsmall cell lung cancer, and melanoma, but is usually not detected in normal human cells. Our aim was to evaluate the presence of NeuGc-GM3 in pediatric neuroectodermal tumors by immunohistochemistry. Twenty-seven archival cases of neuroblastoma and Ewing sarcoma family of tumors (ESFT) were analyzed. Formalin-fixed, paraffin-embedded tumor samples were cut into 5 µm sections. The monoclonal antibody 14F7, a mouse IgG1 that specifically recognizes NeuGc-GM3, and a peroxidase-labeled polymer conjugated to secondary antibodies were used. Presence of NeuGc-GM3 was evident in 23 of 27 cases (85%), with an average of about 70% of positive tumors cells. Immunoreactivity was moderate to intense in most tumors, showing a diffuse cytoplasmic and membranous staining, although cases of ESFT demonstrated a fine granular cytoplasmic pattern. No significant differences were observed between neuroblastoma with and without NMYC oncogene amplification, suggesting that expression of NeuGc-GM3 is preserved in more aggressive cancers. Until now, the expression of N-glycolylated gangliosides in pediatric neuroectodermal tumors has not been investigated. The present study evidenced the expression of NeuGc-GM3 in a high proportion of neuroectodermal tumors, suggesting its potential utility as a specific target of immunotherapy.

Effect of adjuvant perioperative desmopressin in locally advanced canine mammary carcinoma and its relation to histologic grade.

Desmopressin (DDAVP) is a vasopressin peptide analog with hemostatic properties that has been successfully used during surgery in patients with bleeding disorders. Recently published experimental and clinical data indicate that perioperative administration of DDAVP can minimize spread and survival of residual mammary cancer cells. The central aim of this study was to explore the effect of perioperative DDAVP and its relation to histologic grade in bitches with locally advanced mammary carcinoma. Of the 32 dogs initially recruited, 28 intact bitches with mammary carcinoma tumors stage III or IV were ultimately included. These dogs were randomized to receive DDAVP at intravenous doses of 1 µg/kg (n=18) or saline solution as placebo (n=10). En bloc mastectomy of the affected gland(s) was performed. Tumor malignancy was graded by the method of Elston and Ellis into well-differentiated (grade 1), moderately differentiated (grade 2), or poorly differentiated (grade 3). DDAVP therapy significantly prolonged the disease-free survival (P<0.001) and overall survival (P<0.01) in bitches with grade 2 or 3 carcinomas compared with bitches in the control group. No significant difference in disease-free period or overall survival was found between treatment groups in bitches with grade 1 tumors. The present data suggest that DDAVP may be an excellent candidate as a surgical adjuvant in the management of aggressive cancers in small animals. More research in this field is warranted.

Inmunohistoquímica y expresión de receptores de estrógenos y progesterona en neoplasias mamarias malignas caninas en distintos estadios clínicos / Immunohistochemistry and expression of estrogen and progesterone receptors in canine malignant mammary tumors in different clinical stages

La técnica de inmunohistoquímica descrita por Walker y col. (1998) y Mote y col. (2001) es de utilidad para la determinación de receptores de estrógenos y de progesterona en tumores mamarios de la hembra canina. Existe bastante controversia respecto al porcentaje de receptores de estrógenos y de progesterona, con valor pronóstico o predictivo en tumores malignos caninos debido, muy probablemente, a la falta de uniformidad en criterios de inclusión de los casos en los estudios realizados. El presente trabajo se realizó en una población de 51 perras. Los tumores fueron escindidos quirúrgicamente mediante mastectomía de las glándulas afectadas. Para la determinación de receptores hormonales, se analizó la muestra con mayores características histológicas de malignidad de cada animal. Se contabilizó el porcentaje de células tumorales positivas para cada uno de los receptores (receptor de estrógeno alfa, receptor de estrógeno beta y receptores de progesterona) en los distintos estadios clínicos (I, II, II, IV). Se consideró positivo todo corte ³ 20% de células tumorales positivas para cada uno de los receptores. Se realizó una estadística descriptiva (media y error estándar) con los animales positivos para cada uno de los receptores en los distintos estadios clínicos. En línea con reportes de otros autores, más de la mitad de caninos portadores de tumores mamarios malignos expresaron receptores de estrógenos y de progesterona. Conociendo que el comportamiento biológico de una neoplasia varía con el estadio clínico del paciente, resulta interesante estudiar la expresión de los receptores en cada uno de éstos, en animales con neoplasias mamarias malignas. Adicionalmente, a este objetivo incluye la puesta a punto la de técnica de inmunohistoquímica para la determinación de receptores hormonales en la especie.

receptors in canine mammary tumors. There is considerable controversy regarding the percentage of estrogen and progesterone receptors that having prognostic or predictive value in malignant tumors in dogs because, most likely, the lack of uniform criteria for inclusion of cases. This study was conducted in a population of 51 female dogs. The tumors were surgically treated by mastectomy of the affected glands. For the determination of hormone receptors, the sample analyzed was showed higher histological malignancy features of each animal. It was counted the percentage of tumor cells positive for each of the receptors (estrogen receptor alpha, estrogen receptor beta and progesterone receptor) in animals in various clinical stages (I, II, III, IV). It was considered positive cut-off ³ 20% tumor cells positive for each of the recipients. It was conducted a descriptive statistics (mean and standard error) which positive animals for each of the receptors in different clinical stages. In line with reports of other authors, more than half of canine carriers of malignant mammary tumors expressed estrogen and progesterone receptors. Knowing that the biological behavior of cancer varies with the clinical stage of the animal, objective was to study the expression of receptors in each of the clinical stages in animals with malignant mammary tumors. Additionally, this includes the preparation of the immunohistochemical technique for the determination of hormone receptors in this species.

Detection and characterization of N-glycolyated gangliosides in Wilms tumor by immunohistochemistry.

Gangliosides are glycolipids present on the cell surface. The N-glycolylated ganglioside NeuGc-GM3 has been described in some neoplasms, such as breast carcinoma and melanoma, but is usually not detected in normal human cells. Our aim was to evaluate the presence of NeuGc-GM3 in Wilms tumor by immunohistochemistry. Postchemotherapy tumors were grouped into different histologic subtypes considering the main preserved component. Formalin-fixed, paraffin-embedded tumor samples were cut into 5-microm sections. The monoclonal antibody 14F7, a mouse IgG1 that specifically recognizes NeuGc-GM3, and a peroxidase-labeled polymer conjugated to secondary antibodies were used. Sections from breast carcinoma were employed as positive controls. Presence of NeuGc-GM3 was evident in 22 of 25 (88%) cases. The staining was stronger in the epithelial component, with a membrane pattern and cytoplasmic diffusion. The stromal component expressed cytoplasmic NeuGc-GM3 in cells with rhabdomyoblastic differentiation. Tubules of adjacent renal tissue were also positive, but no expression of NeuGc-GM3 was detected in nontumoral fetal kidney. Until now, the expression of N-glycolylated gangliosides in pediatric solid tumors has not been investigated. The present study evidenced the expression of NeuGc-GM3 in a high proportion of Wilms tumors, suggesting its potential utility as a specific target of immunotherapy.

Perioperative desmopressin prolongs survival in surgically treated bitches with mammary gland tumours: a pilot study.

Desmopressin (1-deamino-8-d-arginine vasopressin, also known as DDAVP) is a safe haemostatic compound capable of inhibiting lymph node and lung metastasis in a mouse model of mammary tumour manipulation and surgical excision. The aim of this study was to test the efficacy and safety of perioperative DDAVP (1microg/kg) in surgically treated bitches with mammary gland tumours (MGT). Twenty-one, otherwise healthy, intact bitches, with malignant MGT stage III or IV were randomly allocated to DDAVP (n=11) or placebo (n=10) groups. En bloc mastectomy of the affected gland/s was performed. DDAVP had a significant beneficial effect on disease-free period (P<0.01) and overall survival time (P<0.05). No side effects were seen in any of the cases. Whatever the mechanism of action, it seems that DDAVP may have a novel use in cancer surgery to minimise spread or survival of residual malignant cells. Additional, large scale controlled trials are required to fully evaluate this adjuvant pharmacological protocol.