Interleukin-2 increases the antibody response in patients receiving autologous intralymphatic tumor cell vaccine immunotherapy.

The production of tumor-binding antibodies was studied in a group of cancer patients undergoing active specific immunotherapy with irradiated, cholesterol-treated, cell culture-derived autologous tumor cells injected by the intralymphatic route. Fifteen patients were analyzed: nine patients (four melanoma, one breast, one sarcoma, one colon, and one undifferentiated cancer) received three injections of 10 to 15 x 10(6) tumor cells, spaced 2 weeks apart, and six patients (two melanoma, two renal, one breast, and one colon cancer) received tumor cells admixed with 3 x 10(6) U recombinant interleukin-2 (IL-2) (Proleukin, Cetus, Emeryville, CA, USA) plus a 10-day intravenous infusion of 15 x 10(6) U/kg/day IL-2 after each immunization. Serum antibody binding to autologous tumor cells was measured at 2 and 4 weeks after initiation of therapy using an enzyme-linked immunosorbent assay with patient serum being added to adherent tumor cells bound to 96-well microtiter plates. After 4 weeks, we found a significant difference (0.02 less than P less than 0.04) in serum titer in the group receiving IL-2 (33% mean increase) compared with the non-IL-2 group (8% mean increase). Although neither group showed clinical improvement in response to the therapy, the results clearly demonstrated the efficacy of IL-2 in augmenting patient antibody response to autologous intralymphatic tumor cell immunization.     

Antiserum directed against cell surface antigens is lethal toLeishmania donovani promastigotes

The purified flagellar fraction ofLeishmania donovani promastigotes consists of 30–35 polypeptides. Antiserum raised against this fraction reacts with both flagella and pellicular membrane antigens as evident from immunoblot and immunofluorescence studies. Only 3 of these immunoreactive polypeptides are flagellum-specific. The antiserum agglutinates the cells and inhibits their growth in liquid culture medium. Moreover, glucose uptake and glucose-stimulated oxygen uptake of the promastigotes are significantly inhibited by the antiserum. The results indicate that the antiserum has a profound lethal effect on the invitro propagation of the parasite.     

Identification and characterization of a Mg+2-dependent and an independent Ca+2-ATPase in microsomal membranes of rat testis

Rat testicular microsomal membrane fraction contains both Mg⁺²-dependent and Mg⁺²-independent Ca⁺²-ATPase activity. The latter activity is about two times higher than the former. Calcium ion required for maximum activation of Mg⁺²-independent Ca⁺²-ATPase in 3.0 mM, whereas for the dependent one it is 2.5 mM. Both the enzymes are resistant to cold shock upto seven days. Histidine and imidazole buffers are found to be the most suitable for dependent and independent enzyme activities, respectively. The pH optima for dependent one is 7.5, whereas for the independent one it is 8.5. Temperature optima for the former is 37°C and for latter one it is 40°C. Among all the nuclestides tested, ATP is found to be the best substrate for both the enzymes. The optimum concentration of ATP for dependent and independent enzyme activities are 3.0 mM and 1.5 mM respectively. Divalent metal ions like Zn⁺², Ba⁺² and Mn⁺² have been found to inhibit Mg⁺²-dependent Ca⁺²-ATPase activity whereas Mg⁺²-independent Ca⁺²-ATPase activity is inhibited by the divalent ions except zinc which is found to stimulate the enzyme activity. Both the enzymes are inhibited by vanadata, EDTA and EGTA. I₅₀, for vanadate is 0.05 and 0.125 mM for dependent and independent activities, respectively. Sulfhydral groups modifying agents e.g., NEM, DTNB and chlorpromazine are found to affect the enzyme activities in different ways. Thus NEM and chlorpromazine are found to inhibit and DTNB stimulate the enzyme activities in both the cases.     

Psilorhynchus nahlongthai,a new fish species (Teleostei: Psilorhynchidae) from the Brahmaputra drainage,northeast India

Psilorhynchus nahlongthai, a new psilorhynchid fish, is described from the Diyung River, a tributary to the Kopili River (itself a southern tributary of the Brahmaputra drainage) in Assam, northeast India. It is placed in the Psilorhynchus balitora species group and can be easily distinguished from all other members of this group by a combination of the following characters: dense and prominent tuberculation on the head region; thick and long pre- and post-epiphyseal fontanelles on the neurocranium; 9 + 8 caudal-fin rays; and 34 (24 + 10) vertebrae. Genetic divergence between P. nahlongthai and members of the P. balitora species group from the Brahmaputra and neighbouring drainages, with K2P distances ranging 3.7%–14.7% and 7.4%–20.7% in the mitochondrial COI and cyt b gene datasets respectively, support its report as a new species.     

Heregulin induces increase in sensitivity of an erbB-2-overexpressing breast cancer cell type to lysis by lymphokine-activated killer cells

 The erbB-2 oncoprotein is overexpressed in 30% of tumors from breast and ovarian cancer patients and it is related to poor overall and disease-free survival. In vitro studies on erbB-2-overexpressing cells have found a strong correlation between this oncogene overexpression and relative resistance to lymphokine-activated killer (LAK) cell lysis. gp30/heregulin/NDF (neu differentiation factor), indirect activators of erbB-2, are able to induce a more differentiated phenotype on erbB-2-overexpressing, erbB-3- and/or erbB-4-positive breast cancer cells. We tested the ability of these highly homologous growth factors to stimulate LAK cell lysis of breast cancer cells. Our experiments demonstrated a marked increase in LAK cell cytotoxicity towards an erbB-2-overexpressing, erbB-3-positive cell line by treatment of these cells with heregulin for 72 h. In contrast we did not observe any enhancement of lysis of MCF-7, a cell line that does not overexpress erbB-2 and is positive for the erbB-3 and erbB-4 receptors, after treatment with heregulin. The increased lysis was associated with up-regulation of intercellular adhesion molecule 1 (ICAM-1), down-regulation of erbB-2 and increased binding between breast cancer cells and LAK cells. Pre incubation of target (SKBR3) cells with blocking anti-ICAM-1 antibody completely abrogated the enhanced cytotoxicity. A similar effect was observed by pretreatment of the effector (LAK) cells with antibodies directed against LFA-1, the receptor for ICAM-1. These results suggest the possible utilization of gp30/heregulin in the treatment of breast cancer patients by its ability to stimulate patient immune responses. Received: 6 March 1995 / Accepted: 7 June 1996     

Human germline and pan-cancer variomes and their distinct functional profiles

Identification of non-synonymous single nucleotide variations (nsSNVs) has exponentially increased due to advances in Next-Generation Sequencing technologies. The functional impacts of these variations have been difficult to ascertain because the corresponding knowledge about sequence functional sites is quite fragmented. It is clear that mapping of variations to sequence functional features can help us better understand the pathophysiological role of variations. In this study, we investigated the effect of nsSNVs on more than 17 common types of post-translational modification (PTM) sites, active sites and binding sites. Out of 1 705 285 distinct nsSNVs on 259 216 functional sites we identified 38 549 variations that significantly affect 10 major functional sites. Furthermore, we found distinct patterns of site disruptions due to germline and somatic nsSNVs. Pan-cancer analysis across 12 different cancer types led to the identification of 51 genes with 106 nsSNV affected functional sites found in 3 or more cancer types. 13 of the 51 genes overlap with previously identified Significantly Mutated Genes (Nature. 2013 Oct 17;502(7471)). 62 mutations in these 13 genes affecting functional sites such as DNA, ATP binding and various PTM sites occur across several cancers and can be prioritized for additional validation and investigations.     

Characterization of Membrane Protein Interactions by Isothermal Titration Calorimetry

Understanding the structure, folding, and interaction of membrane proteins requires experimental tools to quantify the association of transmembrane (TM) helices. Here, we introduce isothermal titration calorimetry (ITC) to measure integrin αIIbβ3 TM complex affinity, to study the consequences of helix–helix preorientation in lipid bilayers, and to examine protein-induced lipid reorganization. Phospholipid bicelles served as membrane mimics. The association of αIIbβ3 proceeded with a free energy change of − 4.61 ± 0.04 kcal/mol at bicelle conditions where the sampling of random helix–helix orientations leads to complex formation. At bicelle conditions that approach a true bilayer structure in effect, an entropy saving of > 1 kcal/mol was obtained from helix–helix preorientation. The magnitudes of enthalpy and entropy changes increased distinctly with bicelle dimensions, indicating long-range changes in bicelle lipid properties upon αIIbβ3 TM association. NMR spectroscopy confirmed ITC affinity measurements and revealed αIIbβ3 association and dissociation rates of 4500 ± 100 s^− 1 and 2.1 ± 0.1 s^− 1, respectively. Thus, ITC is able to provide comprehensive insight into the interaction of membrane proteins.