Gul Ahmad

Chemical reduction of three-dimensional silica micro-assemblies into microporous silicon replicas

The carbothermal reduction of silica into silicon requires the use of temperatures well above the silicon melting point (≥2,000 °C). Solid silicon has recently been generated directly from silica at much lower temperatures (≤850 °C) via electrochemical reduction in molten salts. However, the silicon products of such electrochemical reduction did not retain the microscale morphology of the starting silica reactants. Here we demonstrate a low-temperature (650 °C) magnesiothermic reduction process for converting three-dimensional nanostructured silica micro-assemblies into microporous nanocrystalline silicon replicas. The intricate nanostructured silica microshells (frustules) of diatoms (unicellular algae) were converted into co-continuous, nanocrystalline mixtures of silicon and magnesia by reaction with magnesium gas. Selective magnesia dissolution then yielded an interconnected network of silicon nanocrystals that retained the starting three-dimensional frustule morphology. The silicon replicas possessed a high specific surface area (>500 m2 g-1), and contained a significant population of micropores (≤20 Å). The silicon replicas were photoluminescent, and exhibited rapid changes in impedance upon exposure to gaseous nitric oxide (suggesting a possible application in microscale gas sensing). This process enables the syntheses of microporous nanocrystalline silicon micro-assemblies with multifarious three-dimensional shapes inherited from biological or synthetic silica templates for sensor, electronic, optical or biomedical applications.

Experimental vaccines in animal models for schistosomiasis

Considerable morbidity and mortality results from the affliction of an estimated 200 million people worldwide by several species of schistosomes; 779 million are exposed to the disease in 74 different countries. Even though anti-parasitic drugs and other control measures, including public hygiene and snail control are available, the advent of an effective vaccine still remains the most potentially powerful means for the control of this disease. The putative vaccine could be administered to small children prior to the time when their contact with infected water is maximal, so as to prevent severe infection in the subsequent years. This review attempts to summarize the status of schistosome vaccine development with special emphasis on functionally important vaccine candidates. The importance of utilizing both murine and nonhuman primate models as a prerequisite for clinical trials is discussed.

Protective and antifecundity effects of Sm-p80 based DNA vaccine formulation against Schistosoma mansoni in a nonhuman primate model

Schistosomiasis is an important parasitic disease for which there is no available vaccine. We have focused on a functionally important antigen of Schistosoma mansoni, Sm-p80, as a vaccine candidate because of its consistent immunogenicity, protective potential and antifecundity effect observed in murine models; and for its pivotal role in the immune evasion process. In the present study we report that an Sm-p80-based DNA vaccine formulation confers 38% reduction in worm burden in a nonhuman primate model, the baboon (Papio anubis). Animals immunized with Sm-p80-pcDNA3 exhibited a decrease in egg production by 32%. Sm-p80 DNA elicited specific immune responses that include IgG; its subtypes IgG1 and IgG2; and IgM in vaccinated animals. Peripheral blood mononuclear cells (PBMCs) from immunized animals when stimulated in vitro with Sm-p80 produced appreciably more Th1 response enhancing cytokines (IL-2, IFN-gamma) than Th2 response enhancing cytokines (IL-4, IL-10). PBMCs produced appreciably more spot-forming units for INF-gamma than for IL-4 in enzyme-linked immunosorbent spot (ELISPOT) assays. Overall it appears that even though a mixed (Th1/Th2) type of humoral antibody response was generated following immunization with Sm-p80; the dominant protective immune response is Th1 type. These data reinforce the potential of Sm-p80 as an excellent vaccine candidate for schistosomiasis.

Prime-boost and recombinant protein vaccination strategies using Sm-p80 protects against Schistosoma mansoni infection in the mouse model to levels previously attainable only by the irradiated cercarial vaccine

Advent of an effective schistosome vaccine would contribute significantly toward reducing the disease spectrum and transmission of schistosomiasis. We have targeted a functionally important antigen, Sm-p80, as a vaccine candidate because of its consistent immunogenicity, protective and antifecundity potentials, and important role in the immune evasion process. In this study, we report that using two vaccination approaches (prime boost and recombinant protein), Sm-p80-based vaccine formulation(s) confer up to 70% reduction in worm burden in mice. Animals immunized with the vaccine exhibited a decrease in egg production by up to 75%. The vaccine elicited strong immune responses that included IgM, IgA, and IgG (IgG1, IgG2a, IgG2b, and IgG3) in vaccinated animals. Splenocytes proliferated in response to Sm-p80 produced Th1 and Th17 response enhancing cytokines. These results again emphasize the potential of Sm-p80 as a viable vaccine candidate for schistosomiasis.

Preclinical Prophylactic Efficacy Testing of Sm-p80–Based Vaccine in a Nonhuman Primate Model of Schistosoma mansoni Infection and Immunoglobulin G and E Responses to Sm-p80 in Human Serum Samples From an Area Where Schistosomiasis Is Endemic

The prophylactic efficacy of a schistosome antigen (Sm-p80) was tested in a nonhuman primate model, the baboon. Using a total of 28 baboons, different vaccination strategies were used including recombinant Sm-p80 protein formulated in Toll-like receptor 7 and Toll-like receptor 9 agonists, and DNA priming followed by boosting with protein plus adjuvants. Recombinant protein approaches provided levels of prophylactic efficacy of 52%-58%, whereas prime-boost approaches conferred 38%-47% protection in baboons. An appropriately balanced pro-inflammatory (T-helper 17 [Th17] and Th1) and anti-inflammatory (Th2) type of response was generated; the Th1 and Th17 types of immune responses appear to be indicative of increased prophylactic efficacy. Production and expression of several cytokines (interleukin 2 [IL-2], interferon γ, IL-12α, IL-1β, IL-6, and IL-22) were up-regulated in vaccinated animals. Human correlate studies revealed Sm-p80 reactivity with immunoglobulin G in human serum samples from schistosome-infected individuals. In addition, a complete lack of prevailing Sm-p80-specific immunoglobulin E in a high-risk or infected population was observed, thus minimizing the risk of hypersensitivity reaction following vaccination with Sm-p80 in humans. This study provided the proof of concept to move Sm-p80 forward into further preclinical development leading to human clinical trials.

Sm-p80-based DNA vaccine formulation induces potent protective immunity against Schistosoma mansoni.

No effective vaccine exists for the human parasitic disease, schistosomiasis. We have targeted a functionally important antigen, Sm‐p80 as a vaccine candidate because of its consistent immunogenicity, protective potential and important role in the immune evasion process. In this study we report that a Sm‐p80‐based DNA vaccine formulation confers 59% reduction in worm burden in mice. Animals immunized with Sm‐p80‐pcDNA3 exhibited a decrease in egg production by 84%. Sm‐p80 DNA elicited strong immune responses that include IgG2A and IgG2B antibody isotypes in vaccinated animals. Splenocytes proliferated in response to Sm‐p80 produced appreciably more Th1 response enhancing cytokines (IL‐2, IFN‐γ) than Th2 response enhancing cytokines (IL‐4, IL‐10). These data reinforce the potential of Sm‐p80 as an excellent vaccine candidate for schistosomiasis.

Cross-species protection: Schistosoma mansoni Sm-p80 vaccine confers protection against Schistosoma haematobium in hamsters and baboons

The ability of the Schistosoma mansoni antigen, Sm-p80, to provide cross-species protection against Schistosoma haematobium challenge was evaluated in hamster and baboon models. Pronounced reduction in worm burden (48%) and in tissue egg load (64%) was observed in hamsters vaccinated with recombinant Sm-p80 admixed with glucopyranosyl lipid adjuvant-stable emulsion (GLA-SE). Similarly, in baboons, the Sm-p80/GLA-SE vaccine produced a 25% reduction in S. haematobium adult worms and decreased the egg load in the urinary bladder by 64%. A 40% and 53% reduction in fecal and urine egg output, respectively, was observed in vaccinated baboons. A balanced pro-inflammatory (Th17 and Th1) and Th2 type of response was generated after vaccination and appears indicative of augmented prophylactic efficacy. These data on cross-species protection coupled with the prophylactic, therapeutic and antifecundity efficacy against the homologous parasite, S. mansoni, reinforces Sm-p80 as a promising vaccine candidate. It is currently being prepared for GMP-compliant manufacture and for further pre-clinical development leading to human clinical trials. These results solidify the expectation that the Sm-p80 vaccine will provide relief for both the intestinal and the urinary schistosomiasis and thus will be greatly beneficial in reducing the overall burden of schistosomiasis.

Sm-p80—Based DNA Vaccine Provides Baboons with Levels of Protection against Schistosoma mansoni Infection Comparable to Those Achieved by the Irradiated Cercarial Vaccine

To date, no vaccine is available to prevent human schistosomiasis. We have targeted a protein of Schistosoma mansoni that plays an important role in the surface membrane renewal process, a mechanism widely believed to be utilized by the parasite as an immune evasion strategy. Sm-p80 antigen is a promising vaccine target because of its documented immunogenicity, protective efficacy, and antifecundity effects observed in both experimental murine and nonhuman primate models of this infectious disease. In the present study, we report that, in a vector approved for human use (VR1020), an Sm-p80-based DNA vaccine formulation confers a 46% reduction in the worm burden in a baboon (Papio anubis) model. Baboons vaccinated with Sm-p80-VR1020 had a 28% decrease in egg production after challenge with the infectious parasite. Sm-p80-VR1020 vaccine elicited robust immune responses to specific antigen Sm-p80, including immunoglobulin (Ig) G, its subtypes IgG1 and IgG2, and IgA and IgM in vaccinated animals. When stimulated in vitro with recombinant Sm-p80, peripheral blood mononuclear cells and splenocytes from baboons vaccinated with Sm-p80-VR1020 produced considerably higher levels of T helper 1 response-enhancing cytokines (interleukin [IL]-2 and interferon-gamma) than T helper 2 (Th2) response-enhancing cytokines (IL-4 and IL-10). Peripheral blood mononuclear cells produced a significantly higher number of spot-forming units for interferon-gamma than for IL-4 in enzyme-linked immunosorbent spot assays. A mixed T helper 1/T helper 2 type of humoral and T cell responses was generated after immunization with Sm-p80-VR1020. These findings again highlight the potential of Sm-p80 as a promising vaccine candidate for schistosomiasis.

Role of antibodies in Sm-p80-mediated protection against Schistosoma mansoni challenge infection in murine and nonhuman primate models

Schistosomiasis is an important public health concern in more than 76 developing countries. Advent of an anti-schistosome vaccine would undoubtedly add to the existing control measures and may eventually help in the elimination of this disease. In the present study we have attempted to dissect the role(s) of antibodies in Sm-p80 mediated protection by intravenously transferring pooled sera from mice immunized with Sm-p80-pcDNA3 or purified IgG from baboons immunized with Sm-p80-pcDNA3, into naïve C57BL/6 mice, respectively, prior to challenge with cercariae. The passive transfer of antibodies from protected mice (homologous transfers) as well as transfer of total IgG from baboons (heterologous transfers), into naïve mice showed statistically significant reductions in worm burden and in the number of eggs in the tissues. Immunizations of antibody knockout mice (μMt-/-; B10.129S2 (B6)-Igh-6(tm1Cgn)/J) with recombinant Sm-p80 in the presence of CpG-motif oligodeoxynucleotides as an adjuvant, resulted in substantial reduction of Sm-p80-mediated protection, compared to C57BL/6 (normal) control group of mice. Down regulation of cytokines that have important effects on B cell proliferation as well as the recovery of higher number of parasites in antibody knockout indicated a significant role(s) of antibodies in Sm-p80-mediated protection against Schistosoma mansoni in mice. In toto, these studies appear to suggest that antibodies play a significant role in Sm-p80 mediated protection.

Use of an Sm-p80–Based Therapeutic Vaccine to Kill Established Adult Schistosome Parasites in Chronically Infected Baboons

No vaccines are available for human use for any parasitic infections, including the helminthic disease schistosomiasis. Sm-p80, the large subunit of Schistosoma mansoni calpain, is a leading antigen candidate for a schistosomiasis vaccine. Prophylactic and antifecundity efficacies of Sm-p80 have been tested using a variety of vaccine approaches in both rodent and nonhuman primate models. However, the therapeutic efficacy of a Sm-p80-based vaccine had not been determined. In this study, we evaluated the therapeutic efficacy of Sm-p80 by using 2 different strategies and 3 Sm-p80-based vaccine formulations in baboons. Vaccine formulations were able to decrease established adult worms by 10%-36%, reduce retention of eggs in tissues by 10%-57%, and decrease egg excretion in feces by 13%-33%, compared with control formulations. Marked differences were observed in B and T cell immune correlates between vaccinated and control animals. This is the first report of killing of established adult schistosome worms by a vaccine. In addition to distinct prophylactic efficacy of Sm-p80, this study adds to the evidence that Sm-p80 is a potentially important antigen with both substantial prophylactic and therapeutic efficacies. These data reinforce that Sm-p80 should be moved forward along the path toward human clinical trials.