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Dive into the research topics where Robert W. Kaminski is active.

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Featured researches published by Robert W. Kaminski.


Infection and Immunity | 2006

IpaD Localizes to the Tip of the Type III Secretion System Needle of Shigella flexneri

Marianela Espina; Andrew J. Olive; Roma Kenjale; David S. Moore; S. Fernando Ausar; Robert W. Kaminski; Edwin V. Oaks; C. Russell Middaugh; William D. Picking; Wendy L. Picking

ABSTRACT Shigella flexneri, the causative agent of shigellosis, is a gram-negative bacterial pathogen that initiates infection by invading cells within the colonic epithelium. Contact with host cell surfaces induces a rapid burst of protein secretion via the Shigella type III secretion system (TTSS). The first proteins secreted are IpaD, IpaB, and IpaC, with IpaB and IpaC being inserted into the host cell membrane to form a pore for translocating late effectors into the target cell cytoplasm. The resulting pathogen-host cross talk results in localized actin polymerization, membrane ruffling, and, ultimately, pathogen entry. IpaD is essential for host cell invasion, but its role in this process is just now coming to light. IpaD is a multifunctional protein that controls the secretion and presentation of IpaB and IpaC at the pathogen-host interface. We show here that antibodies recognizing the surface-exposed N terminus of IpaD neutralize Shigellas ability to promote pore formation in erythrocyte membranes. We further show that MxiH and IpaD colocalize on the bacterial surface. When TTSS needles were sheared from the Shigella surface, IpaD was found at only the needle tips. Consistent with this, IpaD localized to the exposed tips of needles that were still attached to the bacterium. Molecular analyses then showed that the IpaD C terminus is required for this surface localization and function. Furthermore, mutations that prevent IpaD surface localization also eliminate all IpaD-related functions. Thus, this study demonstrates that IpaD localizes to the TTSA needle tip, where it functions to control the secretion and proper insertion of translocators into host cell membranes.


Expert Review of Vaccines | 2009

Inactivated and subunit vaccines to prevent shigellosis.

Robert W. Kaminski; Edwin V. Oaks

Shigellosis remains a formidable disease globally, with children of the developing world bearing the greatest number of infections. The need for an affordable, safe and efficacious vaccine has persisted for decades. Vaccines to prevent shigellosis can be divided into living and nonliving approaches. Several nonliving Shigella vaccines are currently at different stages of development and show substantial promise. Outlined here is an overview of multiple nonliving vaccine technologies, highlighting their current status and recent advances in testing. In addition, gaps in the knowledge base regarding immune mechanisms of protection are explored.


Vaccine | 2011

Safety and immunogenicity of an intranasal Shigella flexneri 2a Invaplex 50 vaccine

Mark S. Riddle; Robert W. Kaminski; Carlos Williams; Chad K. Porter; Shahida Baqar; Alexis A. Kordis; Theron Gilliland; Joyce Lapa; Melissa Coughlin; Chris Soltis; Erica Jones; Jackie Saunders; Paul B. Keiser; Ryan T. Ranallo; Robert Gormley; M.R. Nelson; K. Ross Turbyfill; David R. Tribble; Edwin V. Oaks

BACKGROUND Shigella flexneri 2a lipopolysaccharide 50 is a nasally delivered subunit vaccine consisting of a macromolecular complex composed of LPS, IpaB, IpaC and IpaD. The current study examined vaccine safety and immunogenicity across a dose range and the clinical performance of a new intranasal delivery device. METHODS Volunteers (N=36) were randomized to receive vaccine via the Dolphin™ (Valois of America, Congers, New York) intranasal spray device at one of three doses (240, 480, and 690 μg) on days 0, 14, and 28. Another group (N=8) received the 240 μg dose via pipette. Vaccine safety was actively monitored and antigen-specific humoral and mucosal immune responses were determined. RESULTS There were no serious adverse events and the majority of adverse events (98%) were mild. Antibody secreting cells (ASC), plasma, and mucosal immune responses to Shigella antigens were detected at all three dose levels with the 690 μg dose inducing the highest magnitude and frequency of responses. Vaccination with comparable doses of Invaplex 50 via the Dolphin™ resulted in higher plasma and ASC immune responses as compared to pipette delivery. CONCLUSION In this trial the S. flexneri 2a Invaplex 50 vaccine was safe, well-tolerated and induced robust levels of antigen-specific intestinal IgA and ASC responses. The spray device performed well and offered an advantage over pipette intranasal delivery.


Vaccine | 2008

Immunogenicity and efficacy of highly purified invasin complex vaccine from Shigella flexneri 2a

K. Ross Turbyfill; Robert W. Kaminski; Edwin V. Oaks

Development of a subunit vaccine for shigellosis requires identification of protective antigens and delivering these antigens in a manner that stimulates immunity comparable to that induced by natural infection. The Shigella invasin complex (Invaplex) vaccine is an ion-exchange-purified extract from virulent Shigella that consists of LPS and several other proteins, including the invasins IpaB and IpaC. Intranasal delivery of Invaplex stimulates protective immunity in small animal models for shigellosis. To identify the active component(s) of Invaplex responsible for its immunogenicity and efficacy, size-exclusion chromatography (SEC) was used to separate Invaplex into several different fractions. A high-molecular mass complex with a molecular mass between 669 MDa and 2 MDa consisted primarily of LPS, IpaB and IpaC and was considered to be a highly purified (HP) form of Invaplex. Using the mouse lung model to evaluate the immunogenicity and efficacy of the SEC fractions it was clearly demonstrated that the high-molecular mass complex of the invasins and LPS was responsible for the protective capacity of parent native Invaplex. Other smaller mass SEC fractions were mostly non-immunogenic and did not stimulate solid protection. In guinea pigs, the HP Invaplex stimulated an enhanced immune response as compared to the parent Invaplex and was fully protective. Isolation and characterization of the immunogenic and protective moiety within Invaplex will allow better standardization of the Invaplex product and may allow future development of an Invaplex assembled from purified components.


Infection and Immunity | 2006

Mucosal Adjuvant Properties of the Shigella Invasin Complex

Robert W. Kaminski; K. Ross Turbyfill; Edwin V. Oaks

ABSTRACT The Shigella invasin complex (Invaplex) is an effective mucosal vaccine capable of protecting against Shigella challenge in animal models. The major antigenic constituents of Invaplex are the Ipa proteins and lipopolysaccharide. The cell-binding capacity of the Ipa proteins prompted the investigation into the adjuvanticity of Invaplex. Using ovalbumin (OVA) as a model antigen, intranasal immunization with OVA combined with Invaplex was found to enhance anti-OVA serum immunoglobulin G (IgG) and IgA responses and induce OVA-specific mucosal antibody responses at sites located both proximal and distal to the immunization site. The immune responses induced with OVA and Invaplex were comparable in both magnitude and duration to the immune responses induced after immunization with OVA and cholera toxin. The OVA-specific immune response was characterized by high levels of serum IgG1 and increased production of interleukin-4 (IL-4), IL-5, or IL-10 from lymphoid cells of immunized animals, suggesting a Th2 response. In addition to enhancing the immunogenicity of OVA, Invaplex-specific immune responses were also induced, indicating the potential for the development of a combination vaccine consisting of Invaplex and other immunogens. Preexisting Invaplex-specific immunity did not interfere with the capacity to enhance the immunogenicity of a second, unrelated vaccine antigen, suggesting that Invaplex could be used as a mucosal adjuvant in multiple vaccine regimens.


Vaccine | 2010

Safety and immunogenicity of a Shigella flexneri 2a Invaplex 50 intranasal vaccine in adult volunteers.

David R. Tribble; Robert W. Kaminski; J. Cantrell; M.R. Nelson; Chad K. Porter; Shahida Baqar; C. Williams; R. Arora; J. Saunders; M. Ananthakrishnan; J. Sanders; G. Zaucha; R. Turbyfill; Edwin V. Oaks

Shigellosis is a leading cause of diarrhea worldwide prompting vaccine development. The Shigella flexneri Invaplex 50 is a macromolecular complex containing IpaB, IpaC, and LPS, formulated from an aqueous extract of virulent Shigella delivered via nasal administration. Preclinical vaccine testing demonstrated safety, immunogenicity and efficacy. An open-label dose-escalating phase 1 study evaluated a 3-dose (2-week intervals) regimen via nasal pipette delivery. Thirty-two subjects were enrolled into one of four vaccine dose groups (10, 50, 240, or 480 microg). The vaccine was well tolerated with minor short-lived nasal symptoms without evidence of dose effect. Antibody-secreting cell (ASC) responses were elicited at doses > or =50 microg with the highest IgG ASC, Invaplex 50 (100%) and S. flexneri 2a LPS (71%), as well as, serologic responses (43%) occurring with the 240 microg dose. Fecal IgA responses, Invaplex 50 (38.5%) and LPS (30.8%), were observed at doses > or =240 microg. The Invaplex 50 nasal vaccine was safe with encouraging mucosal immune responses. Follow-on studies will optimize dose, delivery mechanism and assess efficacy in a S. flexneri 2a challenge study.


Infection and Immunity | 2010

Virulence, inflammatory potential, and adaptive immunity induced by Shigella flexneri msbB mutants.

Ryan T. Ranallo; Robert W. Kaminski; Tonia George; Alexis A. Kordis; Qing Chen; Kathleen Szabo; Malabi M. Venkatesan

ABSTRACT The ability of genetically detoxified lipopolysaccharide (LPS) to stimulate adaptive immune responses is an ongoing area of investigation with significant consequences for the development of safe and effective bacterial vaccines and adjuvants. One approach to genetic detoxification is the deletion of genes whose products modify LPS. The msbB1 and msbB2 genes, which encode late acyltransferases, were deleted in the Shigella flexneri 2a human challenge strain 2457T to evaluate the virulence, inflammatory potential, and acquired immunity induced by strains producing underacylated lipid A. Consistent with a reduced endotoxic potential, S. flexneri 2a msbB mutants were attenuated in an acute mouse pulmonary challenge model. Attenuation correlated with decreases in the production of proinflammatory cytokines and in chemokine release without significant changes in lung histopathology. The levels of specific proinflammatory cytokines (interleukin-1β [IL-1β], macrophage inflammatory protein 1α [MIP-1α], and tumor necrosis factor alpha [TNF-α]) were also significantly reduced after infection of mouse macrophages with either single or double msbB mutants. Surprisingly, the msbB double mutant displayed defects in the ability to invade, replicate, and spread within epithelial cells. Complementation restored these phenotypes, but the exact nature of the defects was not determined. Acquired immunity and protective efficacy were also assayed in the mouse lung model, using a vaccination-challenge study. Both humoral and cellular responses were generally robust in msbB-immunized mice and afforded significant protection from lethal challenge. These data suggest that the loss of either msbB gene reduces the endotoxicity of Shigella LPS but does not coincide with a reduction in protective immune responses.


Vaccine | 2011

Shigella sonnei vaccine candidates WRSs2 and WRSs3 are as immunogenic as WRSS1, a clinically tested vaccine candidate, in a primate model of infection

Shoshana Barnoy; Shahida Baqar; Robert W. Kaminski; T. Collins; K. Nemelka; Thomas L. Hale; Ryan T. Ranallo; Malabi M. Venkatesan

Shigella causes diarrhea and dysentery through contaminated food and water. Shigella sonnei live vaccine candidates WRSs2 and WRSs3 are attenuated principally by the loss of VirG(IcsA) that prevents bacterial spread within the colonic epithelium. In this respect they are similar to the clinically tested vaccine candidate WRSS1. However, WRSs2 and WRSs3 are further attenuated by loss of senA, senB and WRSs3 also lacks msbB2. As previously shown in cell culture assays and in small animal models, these additional gene deletions reduced the levels of enterotoxicity and endotoxicity of WRSs2 and WRSs3, potentially making them safer than WRSS1. However the behavior of these second-generation VirG(IcsA)-based vaccine candidates in eliciting an immune response in a gastrointestinal model of infection has not been evaluated. In this study, WRSs2 and WRSs3 were nasogastrically administered to rhesus monkeys that were evaluated for colonization, as well as for systemic and mucosal immune responses. Both vaccine candidates were safe in rhesus monkeys and behaved comparably to WRSS1 in bacterial excretion rates that demonstrated robust intestinal colonization. Furthermore, humoral and mucosal immune responses elicited against bacterial antigens appeared similar in all categories across all three strains indicating that the additional gene deletions did not compromise the immunogenicity of these vaccine candidates. Based on data from previous clinical trials with WRSS1, it is likely that, WRSs2 and WRSs3 will not only be safer in human volunteers but will generate comparable levels of systemic and mucosal immune responses that were achieved with WRSS1.


Clinical and Vaccine Immunology | 2014

Development and preclinical evaluation of a trivalent, formalin-inactivated Shigella whole-cell vaccine.

Robert W. Kaminski; M. Wu; K. R. Turbyfill; K. Clarkson; B. Tai; August L. Bourgeois; L. L. Van De Verg; Richard I. Walker; Edwin V. Oaks

ABSTRACT Studies were undertaken to manufacture a multivalent Shigella inactivated whole-cell vaccine that is safe, effective, and inexpensive. By using several formalin concentrations, temperatures, and incubation periods, an optimized set of inactivation conditions was established for Shigella flexneri 2a, S. sonnei, and S. flexneri 3a to produce inactivated whole cells expressing a full repertoire of Ipa proteins and lipopolysaccharide (LPS). The inactivation conditions selected were treatment with 0.2% formalin (S. flexneri 2a and 3a) or 0.6% formalin (S. sonnei) for 48 h at 25°C. Vaccine formulations prepared under different inactivation conditions, in different doses (10E5, 10E7, and 10E9 cells), and with or without the inclusion of double-mutant heat-labile toxin (dmLT) were evaluated in mice. Two intranasal immunizations with ≥10E7 inactivated whole cells resulted in high levels of anti-Invaplex and moderate levels of LPS-specific IgG and IgA in serum and in lung and intestinal wash samples. Addition of dmLT to the vaccine formulations did not significantly enhance humoral immunogenicity. Minimal humoral responses for IpaB, IpaC, or IpaD were detected after immunization with inactivated whole Shigella cells regardless of the vaccine inactivation conditions. In guinea pigs, monovalent formulations of S. flexneri 2a of 3a or S. sonnei consisting of 10E8, 10E9, or 10E10 cells were protective in a keratoconjunctivitis assay. A trivalent formulation provided protection against all three serotypes (S. flexneri 2a, P = 0.018; S. flexneri 3a, P = 0.04; S. sonnei, P < 0.0001). The inactivated Shigella whole-cell vaccine approach incorporates an uncomplicated manufacturing process that is compatible with multivalency and the future development of a broadly protective Shigella vaccine.


Vaccine | 2012

Two live attenuated Shigella flexneri 2a strains WRSf2G12 and WRSf2G15: a new combination of gene deletions for 2nd generation live attenuated vaccine candidates.

Ryan T. Ranallo; Suramya Fonseka; Tara L. Boren; Lisa Bedford; Robert W. Kaminski; Sejal Thakkar; Malabi M. Venkatesan

Shigella infections are a major cause of inflammatory diarrhea and dysentery worldwide. First-generation virG-based live attenuated Shigella strains have been successfully tested in phase I and II clinical trials and are a leading approach for Shigella vaccine development. Additional gene deletions in senA, senB and msbB2 have been engineered into second-generation virG-based Shigella flexneri 2a strains producing WRSf2G12 and WRSf2G15. Both strains harbor a unique combination of gene deletions designed to increase the safety of live Shigella vaccines. WRSf2G12 and WRSf2G15 are genetically stable and highly attenuated in both cell culture and animal models of infection. Ocular immunization of guinea pigs with either strain induces robust systemic and mucosal immune responses that protect against homologous challenge with wild-type Shigella. The data support further evaluation of the second-generation strains in a phase I clinical trial.

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Edwin V. Oaks

Walter Reed Army Institute of Research

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Malabi M. Venkatesan

Walter Reed Army Institute of Research

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Ryan T. Ranallo

Walter Reed Army Institute of Research

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K. Ross Turbyfill

Walter Reed Army Institute of Research

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Shahida Baqar

Naval Medical Research Center

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George H. Lowell

Walter Reed Army Institute of Research

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Thomas C. VanCott

Walter Reed Army Institute of Research

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Alexis A. Kordis

Walter Reed Army Institute of Research

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Chad K. Porter

Naval Medical Research Center

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Kristen A. Clarkson

Walter Reed Army Institute of Research

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