A gene on the HER2 amplicon, C35, is an oncogene in breast cancer whose actions are prevented by inhibition of Syk.

BACKGROUND: C35 is a 12 kDa membrane-anchored protein endogenously over-expressed in many invasive breast cancers. C35 (C17orf37) is located on the HER2 amplicon, between HER2 and GRB7. The function of over-expressed C35 in invasive breast cancer is unknown. METHODS: Tissue microarrays containing 122 primary human breast cancer specimens were used to examine the association of C35 with HER2 expression. Cell lines over-expressing C35 were generated and tested for evidence of cell transformation in vitro. RESULTS: In primary breast cancers high levels of C35 mRNA expression were associated with HER2 gene amplification. High levels of C35 protein expression were associated with hallmarks of transformation, such as, colony growth in soft agar, invasion into collagen matrix and formation of large acinar structures in three-dimensional (3D) cell cultures. The transformed phenotype was also associated with characteristics of epithelial to mesenchymal transition, such as adoption of spindle cell morphology and down-regulation of epithelial markers, such as E-cadherin and keratin-8. Furthermore, C35-induced transformation in 3D cell cultures was dependent on Syk kinase, a downstream mediator of signalling from the immunoreceptor tyrosine-based activation motif, which is present in C35. CONCLUSION: C35 functions as an oncogene in breast cancer cell lines. Drug targeting of C35 or Syk kinase might be helpful in treating a subset of patients with HER2-amplified breast cancers.

Lethality-based selection of recombinant genes in mammalian cells: application to identifying tumor antigens.

Many biological processes result in either cell death or cessation of cell growth. However, plasmid- and retrovirus-based mammalian expression vectors in which it has been possible to construct representative cDNA libraries cannot be readily recovered from cells that are not actively dividing. This has limited the efficiency of selection of recombinant genes that mediate either lytic events or growth arrest. Examples include genes that encode the target antigens of cytotoxic T cells, genes that promote stem-cell differentiation and pro-apoptotic genes. We have successfully constructed representative cDNA libraries in a poxvirus-based vector that can be recovered from cells that have undergone lethality-based selection. This strategy has been applied to selection of a gene that encodes a cytotoxic T-cell target antigen common to several independently derived tumors.

Effect of the HELLP syndrome on maternal immune function.

The HELLP syndrome occurs in less than 1% of gravidas and is characterized by hemolysis, elevated liver enzymes and low platelet count. The status of immune function in these high-risk patients is not known but may be of great importance in better understanding the basis, if any, of immune dysfunction in pregnancy-associated hypertensive disorders and from the potential compounding effect of infection upon an already debilitated patient. We assessed maternal immune status in patients with the HELLP syndrome using conventional in vitro techniques. The results of these studies clearly show a depression of both T and B cell potential and impaired monocyte handling of intracellular pathogens (up to 33%, 11% and 17% of control values, respectively). The onset of this immunosuppression occurred before the clinical diagnosis of HELLP syndrome was made and persisted for at least 14 days after clinical resolution. Results of cell admixture studies suggest that these effects are mediated by accessory cells or their products and do not represent true lymphocyte dysfunction. The risk of opportunistic infections may therefore be increased in the patient with the HELLP syndrome because of this generalized immunosuppression and profound decrease in monocyte phagocytic and bactericidal activity.

The effects of betamethasone on maternal cellular resistance to infection.

Antenatal administration of glucocorticoids is often used to facilitate fetal lung maturation in cases of prematurity; however, the effects of betamethasone on maternal immune function have not been established. Therefore maternal immune function was assessed with the use of in vitro techniques. Transient and incomplete suppression of the proliferative response to the T-cell mitogen phytohemagglutinin was demonstrated as early as 24 hours after administration of betamethasone. The magnitude and duration of suppression showed a corresponding increase with advancing gestational age, but these effects were not cumulative and were always short-lived (less than 72 hours). No such suppression of the B-cell mitogen lipopolysaccharide was detected. The nonspecific cellular resistance to infection of maternal monocytes was determined through coincubation with the facultative intracellular pathogen Listeria monocytogenes. Increased phagocytic activity with a normal bactericidal effect was measured in the cell preparations obtained from recipients versus nonrecipients of betamethasone. Taken together, these findings clearly show that both specific and nonspecific immune function are intact in the preterm gravid woman after administration of betamethasone and should allay concerns over its use for reasons of infection control alone.

Hereditary pituitary prolactinomas: a new rat model.

Pituitary adenomas are known to occur spontaneously in several rat strains, especially during aging. Here we report the occurrence of adenomas (prolactinomas) in a new rat model at an early age. The adenomas were characterized by light and electron microscopy and histochemically by immunoperoxidase methods using specific antisera. Of the 53 pituitaries examined, one was normal, two were hyperplastic and the remainder were adenomas. Among the latter, there were 37 prolactinomas (70%), 3 GH-producing tumours and 2 ACTH-containing neoplasms. One microadenoma produced both PRL and GH. The remaining 8 adenomas were histochemically negative for all pituitary hormones. Serum PRL levels were elevated in both sexes while serum testosterone was high in females and low in the males. The serum estrogen levels were elevated in the males with prolactinomas. The prolactinomas appear to be transmitted as an autosomal dominant characteristic with incomplete penetrance and a greater incidence in the male.

Comparative immunology: sipunculid bactericidal responses.

This report continues comparative studies of inducible bactericidins of invertebrates. The bactericidin of the sipunculid worm Dendrostomum zostericolum Chamberlain was induced by intracoelomic injections of killed bacteria. With the assay system used, optimal results were obtained with 90 min of incubation at 20 to 35 C. Bactericidal titers in coelomic fluids from noninjected worms were either low or nonexistent. After injections of gram-negative bacteria, bactericidal titers of up to 1:1280 were reached within 7 days. Considerable individual variations were noted. Optimal stimulation was achieved with 4 x 10(8) bacteria per injection. The response was relatively nonspecific as measured by cross-immunization and absorption experiments. Injections of sterile sea water produced no response. Coelomic fluids could be inactivated by heating at 50 or 56 C for 20 min. Activity was not restored by a pool of unheated coelomic fluid from nonimmunized worms. The significance of these observations was discussed with reference to immunity of sipunculids and to comparative immunology.

Bactericidal antibody response in the Pacific hagfish, Eptatretus stoutii.

The hagfish, Eptatretus stoutii, has been shown to synthesize bactericidal antibodies after injections of gram-negative organisms. These bactericidins could be detected as early as 2 days after the primary injection. Secondary responses were accelerated, although titers were not significantly higher than those reached in the primary response. The bactericidins of hagfish were found to react with other gramnegative enteric bacteria, but not with gram-positive strains. The bactericidins show a lesser degree of specificity and a shorter induction period than other hagfish antibodies or most antibodies induced in higher vertebrates. The bactericidal activity was temperature dependent and could be irreversibly destroyed by heating at 50 C for 20 min. Immune sera were subjected to gel filtration on Sephadex G-200, and bactericidal activity was found to be associated with the excluded peak indicating a high-molecular-weight component. Evidence indicates that inducible immune responses of this bactericidal type represent a primitive capacity which arose before the emergence of vertebrate species.

Induced bactericidal response in the hagfish.

Hagfish were shown to be capable of synthesizing bactericidins after injection of gram-negative bacteria. The bactericidins could be detected as early as 2 days after injection. The degree of specificity was not as impressive as in mammalian systems.