IL-13 receptor-targeted cytotoxin cancer therapy leads to complete eradication of tumors with the aid of phagocytic cells in nude mice model of human cancer

Tumor-directed therapeutic approaches require unique or overexpressed specific Ag or receptor as a target to achieve selective tumor killing. However, heterogeneous expression of these targets on tumor cells limits the efficacy of this form of therapy. In this study, we forced abundant expression of IL-13Ralpha2 chain by plasmid-mediated gene transfer in head and neck, as well as prostate tumors to provide a potential target. This was followed by successfully treating xenograft tumor-bearing nude mice with IL-13R-directed cytotoxin (IL13-PE38QQR). Although we did not observe an indirect cytotoxic bystander effect conveyed to nontransduced tumor cells in vitro, our approach in vivo led to a complete regression of established tumors transfected with IL-13Ralpha2 chain in most animals. We found that the tumor eradication was achieved in part by infiltration of macrophages and NK cells, assessed by immunohistochemistry. Moreover, head and neck tumors xenografted in macrophage-depleted nude mice were less sensitive to the antitumor effect of IL-13 cytotoxin. Because we did not observe vector-related toxicity in any vital organs, our novel combination strategy of gene transfer of IL-13Ralpha2 chain and receptor-directed cytotoxin therapy may be a useful approach for the treatment of localized cancer.     

Microfilariae in association with other diseases. A report of six cases

BACKGROUND: Lymphatic filariasis is a major public health problem in tropical countries. Earlier reports have reported microfilariae as an incidental finding in body fluids and fine needle aspiration smears from various sites. CASES: The findings of body fluid cytology and fine needle aspiration smears from six patients with microfilariae in association with other conditions--tubercular pleural effusion/lymphadenitis, pregnancy and non-Hodgkin's lymphoma--are presented. Three patients demonstrated an associated eosinophilic cellular exudate. Adherence of inflammatory cells to microfilariae was seen in two patients. CONCLUSION: Although microfilariae in cytologic smears are considered incidental findings, the association of microfilariae with debilitating conditions suggests that it is an opportunistic infection and needs further study.     

Immunogenicity and protective efficacy of three DNA vaccines encoding pre-erythrocytic- and erythrocytic-stage antigens of Plasmodium cynomolgi in rhesus monkeys

Although several malaria vaccine candidate antigens have been identified, the most suitable methods for their delivery are still being investigated. In this regard, direct immunization with DNA encoding these vaccine target antigens is an attractive alternative. Here, we have investigated the immune responses to DNA immunization with three major vaccine target antigens: the apical membrane antigen-1 and the 19-kDa C-terminal fragment of merozoite surface protein-1 from the erythrocytic stage, and the thrombospondin-related adhesive protein from the pre-erythrocytic stage of Plasmodium cynomolgi in rhesus monkeys. Antigen-specific antibodies were developed in all the immunized monkeys and peripheral blood mononuclear cells from all immunized monkeys proliferated to different extents upon in vitro stimulation with the corresponding recombinant proteins. The immunized monkeys were challenged with P. cynomolgi sporozoites. All of the immunized animals developed infection but although there was no significant difference between the control and vaccinated animals in terms of pre-patent period, total duration of patency and primary peak parasitemia, the vaccinated animals had significantly lower secondary peak parasitemia than the control animals.     

In vivo active antimalarial isonitriles

Building on the lead from antimalarial isonitriles 1-4 of marine origin, several easily accessible synthetic isonitriles were assessed for their antimalarial activity against Plasmodium falciparum (in vitro) and multidrug resistant Plasmodium yoelii in Swiss mice model (in vivo). Isonitrile 11 has shown promising activity in both these assays.     

Solid-phase synthesis and bioevaluation of lupeol-based libraries as antimalarial agents

The use of the triterpenoid lupeol as a scaffold for the synthesis of lupeol-based libraries is described. Lupeol was anchored to a solid support (Rink amide/Sieber Amide) through aliphatic dicarboxylic acid moieties, which also served as a site for introducing diversity. The resulting polymer linked 3beta-O (resin-alkanoyl)-lup-20(29)-ene 3 was used to generate key intermediates 3beta-O (resin-alkanoyl)-30-bromo-lup-20(29)-ene 4 and 3beta-O (resin-alkanoyl)-30-amino-lup-20(29)-ene 6 for the generation of libraries based on disubstituted lupeol derivatives. A 96-member library was screened for its in-vitro antimalarial activity against Plasmodium falciparum.     

Progesterone receptor as an indicator of sperm function

Expression of progesterone receptor (PR) localization on spermatozoa was determined in men with normal and abnormal spermiograms. Studies were also carried out to evaluate the potential of PR as a marker of sperm function. Progesterone receptor expression on spermatozoa from men with normozoospermia (n = 8), oligozoospermia (n = 7), asthenozoospermia (n = 8), oligoasthenozoospermia (n = 7), and teratozoospermia (n = 11) was analyzed using an immunocytochemical method with monoclonal antibodies against PR, and flow cytometry using a cell-impermeable fluorescein-tagged progesterone coupled to BSA complex (P-FITC-BSA). Both methods revealed significantly fewer (P < 0.05) PR-positive spermatozoa in men with oligozoospermia, asthenozoospermia, oligoasthenozoospermia, and teratozoospermia compared with men with normozoospermia, thereby suggesting that down-regulation of PR expression in spermatozoa may be one of the causes of male infertility. Spermatozoa from men with normozoospermia (n = 12), oligozoospermia (n = 12), asthenozoospermia (n = 12), oligoasthenozoospermia (n = 9), and teratozoospermia (n = 10) were exposed to low osmotic conditions in the hypoosmotic swelling (HOS) test and then analyzed for PR expression using P-FITC-BSA complex. A significantly higher percentage (P < 0.05) of spermatozoa with physiologically active plasma membrane (HOS+) lacked PR expression (HOS+PR-) in all categories of men with infertility, thereby suggesting that compared to the HOS test, PR expression is a better indicator of sperm function. Furthermore, PR expression in spermatozoa showed a strong (P < 0.05) positive correlation with their ability to undergo an in vitro acrosome reaction. This was observed in all study groups (i.e., normozoospermia, r = 0.8545; oligozoospermia, r = 0.8711; asthenozoospermia, r = 0.7645; oligoasthenozoospermia, r = 0.9003; and teratozoospermia, r = 0.8676). This suggests a potential role for PR in the events leading to the acrosome reaction in sperm.     

Involvement of rho and rho-associated kinase in sphincteric smooth muscle contraction by angiotensin II

The tonic smooth muscles of lower esophageal sphincter (LES) and internal anal sphincter (IAS) are subject to modulation by the neurohumoral agents. We report that angiotensin (Ang) II-induced contraction of rat IAS and LES smooth muscle cells (SMC) was inhibited by Clostridium botulinum C3 exozyme, HA 1077 and Y 27632, suggesting a role for Rho kinase and a Rho-associated kinase (ROK). Ang II-induced contraction of the SMC was also attenuated by genistein, antibodies to the pp60(c-src), p(190) RhoGTPase-activating protein (p190 RhoGAP), carboxyl terminus of Galpha13, carboxyl terminus peptide, and ADP ribosylation factor (ARF) antibody. Ang II-induced increase in p(190) RhoGAP tyrosine phosphorylation was attenuated by genistein. Furthermore, Ang II-induced increase in smooth muscle tone and phosphorylation of myosin light chain (MLC; 20 kDa; MLC20-P) were attenuated by Y 27632 and genistein. The results suggest an important role for Galpha13 and pp60(c-src) in the intracellular events responsible for the activation of RhoA/ROK in Ang II-induced contraction of LES and IAS SMC.     

Expression of heat shock and cold shock proteins in the gorgonian Leptogorgia virgulata

In this study, we analyzed the response of the temperate, shallow-water gorgonian, Leptogorgia virgulata, to temperature stress. Proteins were pulse labeled with (35)S-methionine/cysteine for 1 h to 2 h at 22 degrees C (control), or 38 degrees C, or for 4 h at 12.5 degrees C. Heat shock induced synthesis of unique proteins of 112, 89, and 74 kDa, with 102, 98 and 56 kDa proteins present in the control as well. Cold shock from 22 degrees C-12.5 degrees C induced the synthesis of a 25 kDa protein, with a 44 kDa protein present in the control as well. Control samples expressed unique proteins of 38, and 33 kDa. Non-radioactive proteins expressed under the same conditions as above, as well as natural field conditions, were tested for reactivity with antibodies to heat shock proteins (HSPs). HSP60 was the major protein found in L. virgulata. Although HSP47, HSP60, and HSP104 were present in all samples, the expression of HSP60 was enhanced in heat stressed colonies, while HSP47 and HSP104 expression were greatest in cold shocked samples. Inducible HSP70 was expressed in cold-shocked, heat-shocked, and field samples. Constitutively expressed HSP70 was absent from all samples. The expression of HSP90 was limited to heat shocked colonies. The expression of both HSP70 and HSP104 suggests that the organism may also develop a stress tolerance response.     

DNA prime-protein boost immunization in monkeys: efficacy of a novel construct containing functional domains of Plasmodium cynomolgi CS and TRAP

We report the efficacy of a bimodal immunization regimen that involved priming with naked DNA (multiple doses) followed by a booster with recombinant protein in rhesus monkeys with a chimeric construct containing the N-terminus of thrombospondin-related adhesive protein and the C-terminus of circumsporozoite protein of Plasmodium cynomolgi. The vaccinated animals developed high titer antibodies against the chimeric antigen, the two components of the hybrid and the native proteins of sporozoites. The peripheral blood mononuclear cells isolated from the vaccinated animals had significant in vitro T cell proliferation activity when stimulated with the recombinant chimeric protein. Furthermore, following challenge with 1000 P. cynomolgi sporozoites, the peak and total parasitemia were significantly lower in vaccinated animals than in the control animals.