Publications by authors named "Joseph Bower"

25 Publications

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Recommendations for the content and management of Certificates of Analysis for reference standards from the GCC for bioanalysis.

Bioanalysis 2021 Apr 13;13(8):609-619. Epub 2021 Apr 13.

Worldwide Clinical Trials, Austin, TX, USA.

The 13th Global CRO Council (GCC) closed forum for bioanalysis was held in New Orleans, LA, USA on 5 April 2019. This GCC meeting was organized to discuss the contents of the 2019 ICH M10 Bioanalytical Method Validation Draft Guideline published in February 2019 and consolidate the feedback of the GCC members. While ICH M10 will cover requirements for reference standards, one of the biggest challenges facing the CRO community is the lack of consistency and completeness of Certificates of Analysis for reference standards used in regulated bioanalysis. Similar challenges exist with critical reagents (e.g., capture and detection antibodies) used for assays supporting biologics. The recommendations provided in this publication are the minimum requirements for the content that GCC members believe should be included in Certificates of Analysis for reference standards obtained from commercial vendors, sponsors and compendial suppliers, for use in regulated bioanalytical studies. In addition, recommendations for internal standards, metabolites and critical reagents are discussed.
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http://dx.doi.org/10.4155/bio-2021-0046DOI Listing
April 2021

Lipoprotein Drug Delivery Vehicles for Cancer: Rationale and Reason.

Int J Mol Sci 2019 Dec 15;20(24). Epub 2019 Dec 15.

Advanced Imaging Research Center, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390, USA.

Lipoproteins are a family of naturally occurring macromolecular complexes consisting amphiphilic apoproteins, phospholipids, and neutral lipids. The physiological role of mammalian plasma lipoproteins is to transport their apolar cargo (primarily cholesterol and triglyceride) to their respective destinations through a highly organized ligand-receptor recognition system. Current day synthetic nanoparticle delivery systems attempt to accomplish this task; however, many only manage to achieve limited results. In recent years, many research labs have employed the use of lipoprotein or lipoprotein-like carriers to transport imaging agents or drugs to tumors. The purpose of this review is to highlight the pharmacologic, clinical, and molecular evidence for utilizing lipoprotein-based formulations and discuss their scientific rationale. To accomplish this task, evidence of dynamic drug interactions with circulating plasma lipoproteins are presented. This is followed by epidemiologic and molecular data describing the association between cholesterol and cancer.
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http://dx.doi.org/10.3390/ijms20246327DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6940806PMC
December 2019

GCC Consolidated Feedback to ICH on the 2019 ICH M10 Bioanalytical Method Validation Draft Guideline.

Bioanalysis 2019 Sep 30;11(18s):1-228. Epub 2019 Sep 30.

WuXi Apptec, Shanghai, China.

The 13 GCC Closed Forum for Bioanalysis was held in New Orleans, Louisiana, USA on April 5, 2019. This GCC meeting was organized to discuss the contents of the 2019 ICH M10 Bioanalytical Method Validation Draft Guideline published in February 2019 and consolidate the feedback of the GCC members. In attendance were 63 senior-level participants from eight countries representing 44 bioanalytical CRO companies/sites. This event represented a unique opportunity for CRO bioanalytical experts to share their opinions and concerns regarding the ICH M10 Bioanalytical Method Validation Draft Guideline and to build unified comments to be provided to the ICH.
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http://dx.doi.org/10.4155/bio-2019-0207DOI Listing
September 2019

HER2/HER3 regulates lactate secretion and expression of lactate receptor mRNA through the MAP3K4 associated protein GIT1.

Sci Rep 2019 07 25;9(1):10823. Epub 2019 Jul 25.

The Department of Pharmacology and Toxicology, College of Pharmacy University of Arizona, Tucson, Arizona, 85721, USA.

One of the major features of cancer is Otto Warburg's observation that many tumors have increased extracellular acidification compared to healthy tissues. Since Warburg's observation, the importance of extracellular acidification in cancer is now considered a hallmark of cancer. Human MAP3K4 functions upstream of the p38 and JNK mitogen activated protein kinases (MAPKs). Additionally, MAP3K4 is required for cell migration and extracellular acidification of breast cancer cells in response to HER2/HER3 signaling. Here, we demonstrate that GIT1 interacts with MAP3K4 by immunoprecipitation, while cellular lactate production and the capacity of MCF-7 cells for anchorage independent growth in soft agar were dependent on GIT1. Additionally, we show that activation of HER2/HER3 signaling leads to reduced expression of lactate receptor (GPR81) mRNA and that both, GIT1 and MAP3K4, are necessary for constitutive expression of GPR81 mRNA. Our study suggests that targeting downstream proteins in the HER2/HER3-induced extracellular lactate signaling pathway may be a way to inhibit the Warburg Effect to disrupt tumor growth.
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http://dx.doi.org/10.1038/s41598-019-46954-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6658559PMC
July 2019

11th GCC Closed Forum: cumulative stability; matrix stability; immunogenicity assays; laboratory manuals; biosimilars; chiral methods; hybrid LBA/LCMS assays; fit-for-purpose validation; China Food and Drug Administration bioanalytical method validation.

Bioanalysis 2018 Apr 27;10(7):433-444. Epub 2018 Apr 27.

Worldwide Clinical Trials, Austin, TX, USA.

The 11th Global CRO Council Closed Forum was held in Universal City, CA, USA on 3 April 2017. Representatives from international CRO members offering bioanalytical services were in attendance in order to discuss scientific and regulatory issues specific to bioanalysis. The second CRO-Pharma Scientific Interchange Meeting was held on 7 April 2017, which included Pharma representatives' sharing perspectives on the topics discussed earlier in the week with the CRO members. The issues discussed at the meetings included cumulative stability evaluations, matrix stability evaluations, the 2016 US FDA Immunogenicity Guidance and recent and unexpected FDA Form 483s on immunogenicity assays, the bioanalytical laboratory's role in writing PK sample collection instructions, biosimilars, CRO perspectives on the use of chiral versus achiral methods, hybrid LBA/LCMS assays, applications of fit-for-purpose validation and, at the Global CRO Council Closed Forum only, the status and trend of current regulated bioanalytical practice in China under CFDA's new BMV policy. Conclusions from discussions of these topics at both meetings are included in this report.
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http://dx.doi.org/10.4155/bio-2018-0014DOI Listing
April 2018

Downregulation of GLUT4 contributes to effective intervention of estrogen receptor-negative/HER2-overexpressing early stage breast disease progression by lapatinib.

Am J Cancer Res 2016 1;6(5):981-95. Epub 2016 May 1.

Department of Molecular and Cellular Oncology, The University of Texas M. D. Anderson Cancer CenterHouston, TX 77030, USA; Cancer Biology Program, The University of Texas Graduate School of Biomedical Sciences at HoustonHouston, TX 77030, USA.

Tamoxifen and aromatase inhibitors (AIs) have shown efficacy in prevention of estrogen receptor-positive (ER+) breast cancer; however, there exists no proven prevention strategy for estrogen receptor-negative (ER-) breast cancer. Up to 40% of ER- breast cancers have human epidermal growth factor receptor 2 overexpression (HER2+), suggesting HER2 signaling might be a good target for chemoprevention for certain ER- breast cancers. Here, we tested the feasibility of the HER2-targeting agent lapatinib in prevention and/or early intervention of an ER-/HER2+ early-stage breast disease model. We found that lapatinib treatment forestalled the progression of atypical ductal hyperplasia (ADH)-like acini to ductal carcinoma in situ (DCIS)-like acini in ER-/HER2+ human mammary epithelial cells (HMECs) in 3D culture. Mechanistically, we found that inhibition of HER2/Akt signaling by lapatinib led to downregulation of GLUT4 and a reduced glucose uptake in HER2-overexpressing cells, resulting in decreased proliferation and increased apoptosis of these cells in 3D culture. Additionally, our data suggest that HER2-driven glycolytic metabolic dysregulation in ER-/HER2+ HMECs might promote early-stage breast disease progression, which can be reversed by lapatinib treatment. Furthermore, low-dose lapatinib treatment, starting at the early stages of mammary grand transformation in the MMTV-neu* mouse model, significantly delayed mammary tumor initiation and progression, extended tumor-free survival, which corresponded to effective inhibition of HER2/Akt signaling and downregulation of GLUT4 in vivo. Taken together, our results indicate that lapatinib, through its inhibition of key signaling pathways and tumor-promoting metabolic events, is a promising agent for the prevention/early intervention of ER-/HER2+ breast cancer progression.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4889714PMC
June 2016

8th GCC: consolidated feedback to US FDA on the 2013 draft FDA guidance on bioanalytical method validation.

Bioanalysis 2014 ;6(22):2957-63

Covance Laboratories, Chantilly, VA, USA.

The 8th GCC Closed Forum for Bioanalysis was held in Baltimore, MD, USA on 5 December 2013, immediately following the 2013 AAPS Workshop (Crystal City V): Quantitative Bioanalytical Methods Validation and Implementation--The 2013 Revised FDA Guidance. This GCC meeting was organized to discuss the contents of the draft revised FDA Guidance on bioanalytical method validation that was published in September 2013 and consolidate the feedback of the GCC members. In attendance were 63 senior-level participants, from seven countries, representing 46 bioanalytical CRO companies/sites. This event represented a unique opportunity for CRO bioanalytical experts to share their opinions and concerns regarding the draft FDA Guidance, and to build unified comments to be provided to the FDA.
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http://dx.doi.org/10.4155/bio.14.287DOI Listing
July 2015

Recommendations on incurred sample stability (ISS) by GCC.

Bioanalysis 2014 Sep;6(18):2385-90

Quintiles Bioanalytical & ADME Labs, Ithaca, NY, USA.

The topic of incurred sample stability (ISS) has generated considerable discussion within the bioanalytical community in recent years. The subject was an integral part of the seventh annual Workshop on Recent Issues in Bioanalysis (WRIB) held in Long Beach, CA, USA, in April 2013, and at the Global CRO Council for Bioanalysis (GCC) meeting preceding it. Discussion at both events focused on the use of incurred samples for ISS purposes in light of results from a recent GCC survey completed by member companies. This paper reports the consensus resulting from these discussions and serves as a useful reference for depicting ISS issues and concerns, summarizing the GCC survey results and providing helpful recommendations on ISS in the context of bioanalytical method development and application.
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http://dx.doi.org/10.4155/bio.14.155DOI Listing
September 2014

Recommendations and best practices for reference standards and reagents used in bioanalytical method validation.

AAPS J 2014 Mar 6;16(2):352-6. Epub 2014 Feb 6.

Covance Laboratories Inc., 3635 Concorde Parkway, Suite 100, Chantilly, Virginia, 20151, USA,

The continued globalization of pharmaceutics has increased the demand for companies to know and understand the regulations that exist across the globe. One hurdle facing pharmaceutical and biotechnology companies developing new drug candidates is interpreting the current regulatory guidance documents and industry publications associated with bioanalytical method validation (BMV) from each of the different agencies throughout the world. The objective of this commentary is to provide our opinions on the best practices for reference standards and key reagents, such as metabolites and internal standards used in the support of regulated bioanalysis based on a review of current regulatory guidance documents and industry white papers for BMV.
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http://dx.doi.org/10.1208/s12248-014-9566-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3933579PMC
March 2014

Racial difference in Acylation Stimulating Protein (ASP) correlates to triglyceride in non-obese and obese African American and Caucasian women.

Nutr Metab (Lond) 2009 Apr 17;6:18. Epub 2009 Apr 17.

Department of Biological and Chemical Sciences, Faculty of Pure and Applied Sciences, University of The West Indies, Cave Hill Campus, Bridgetown, St Michael, Barbados.

Background: Acylation Stimulating Protein (ASP) has been shown to influence adipose tissue triglyceride (TG) storage. The aim was to examine ethnic differences in ASP and leptin levels in relation to lipid profiles and postprandial changes amongst African American (AA) and Caucasian American (CA) women matched for BMI.

Methods: 129 women were recruited in total (age 21 - 73 y): 24 non-obese (BMI < 30 kg/m²) CA, 27 obese (BMI ≥ 30 kg/m²) CA, 13 obese diabetic CA, 25 non-obese AA, 25 obese AA, and 15 obese diabetic AA. Cholesterol, HDL-C, LDL-C, apoB, glucose and insulin were measured at baseline. TG, non-esterified fatty acids, leptin, and ASP were measured at baseline and postprandially following a fat meal.

Results: ASP, leptin, insulin and TG were significantly increased in obese subjects within each race. However, AA women had significantly lower ASP and TG than CA women at all BMI. Obese and diabetic AA women had significantly lower apoB levels than CA women when compared to their respective counterparts. For AA women, fasting ASP was positively correlated with BMI, cholesterol, apoB, LDL-C and glucose. For CA women, fasting ASP was positively correlated with BMI, leptin, glucose and insulin. However, for any given BMI, ASP was significantly reduced in AA vs CA (p = 0.0004). Similarly, for any given leptin level or TG levels, ASP was significantly lower in AA women (p = 0.041 and p = 0.003, respectively).

Conclusion: CA women have higher baseline TG levels and an earlier TG peak that is accompanied with higher ASP levels suggesting increased ASP resistance, while AA women have lower baseline TG levels and a later TG peak at lower ASP levels suggesting increased ASP sensitivity. This may explain why AA women may have fewer metabolic complications, such as diabetes and CVD, when compared to their Caucasian counterparts at the same level of obesity.
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http://dx.doi.org/10.1186/1743-7075-6-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2679015PMC
April 2009

Solve the succession crisis by growing inside-outside leaders.

Authors:
Joseph L Bower

Harv Bus Rev 2007 Nov;85(11):90-6, 153

Busniess Administration, harvard Business School, Boston, USA.

In his analysis of 1800 successions, Harvard Business School professor Bower found that companies performed significantly better when they appointed insiders to the job of CEO. Other researchers, including Jim Collins in Good to Great, have come to similar conclusions working from different data sets. Yet Bower finds far too many companies have no succession plans; as a result, when the time comes to name a new chief executive, more firms turn to outsiders. Both insider and outsider CEOs have strengths and weaknesses at the start. Insiders know the company and its people but are often blind to the need for radical change. Outsiders see the need for a new approach but can't make the necessary changes because they don't know the organization or industry sector well enough. What companies must do, then, is find a way to nurture what Bower calls inside-outsiders--internal candidates who have outside perspective. Often such executives have spent much of their time away from the mainstream of the organization, and away from headquarters, living with new opportunities and threats. Before becoming CEO, Procter & Gamble's A.G. Lafley, for instance, worked for years building P&G's Chinese cosmetics operation rather than the core detergent business. IBM's Sam Palmisano was a champion of software and open systems at a time when Big Blue was essentially a closed-system, hardware-oriented company. Nascent inside-outsiders should enter the CEO-training process by the time they are 30 and be given the opportunity to manage a whole business, so that they become good insiders. But they also need to be mentored with an eye toward preserving their outsider perspective, so they learn how to turn their new ideas into great businesses and are protected from old-timers who might be inclined to teach them a lesson.
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November 2007

How Managers' everyday decisions create or destroy your company's strategy.

Harv Bus Rev 2007 Feb;85(2):72-9, 154

Harvard Business School, Boston, USA.

Senior executives have long been frustrated by the disconnection between the plans and strategies they devise and the actual behavior of the managers throughout the company. This article approaches the problem from the ground up, recognizing that every time a manager allocates resources, that decision moves the company either into or out of alignment with its announced strategy. A well-known story--Intel's exit from the memory business--illustrates this point. When discussing what businesses Intel should be in, Andy Grove asked Gordon Moore what they would do if Intel were a company that they had just acquired. When Moore answered, "Get out of memory," they decided to do just that. It turned out, though, that Intel's revenues from memory were by this time only 4% of total sales. Intel's lower-level managers had already exited the business. What Intel hadn't done was to shut down the flow of research funding into memory (which was still eating up one-third of all research expenditures); nor had the company announced its exit to the outside world. Because divisional and operating managers-as well as customers and capital markets-have such a powerful impact on the realized strategy of the firm, senior management might consider focusing less on the company's formal strategy and more on the processes by which the company allocates resources. Top managers must know the track record of the people who are making resource allocation proposals; recognize the strategic issues at stake; reach down to operational managers to work across division lines; frame resource questions to reflect the corporate perspective, especially when large sums of money are involved and conditions are highly uncertain; and create a new context that allows top executives to circumvent the regular resource allocation process when necessary.
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February 2007

A minimum CR2 binding domain of C3d enhances immunity following vaccination.

Adv Exp Med Biol 2006 ;586:249-64

Department of Medicine, Division of Infectious Diseases, University of Pittsburgh, School of Medicine, Pittsburgh, PA, 15261, USA.

The degradation product of the third (C3) complement component, C3d, links innate and adaptive immunity, and the covalent attachment of C3d to an antigen enhances antigen-specific immune responses. C3d has been hypothesized to enhance immunity by direct interaction with complement receptor 2 (CR2/CD21) on immune cells. However, the domains on C3d important for CR2 binding have been controversial, with various studies reaching contradictory conclusions. In addition, the concept of B-cell activation via CR2 by C3d has been questioned, since mice lacking CR2 still elicit C3d-enhanced immunity following vaccination. Therefore, the goal of this study was to determine if a peptide representing one of the proposed CR2 binding domains of C3d could substitute for the entire protein and enhance antigen-specific immunity. Mice (BALB/c) were vaccinated with the HIV-1 gp120 envelope glycoprotein (Env(gp120)) alone or fused to multiple copies of the murine C3d or a twenty-eight amino-acid peptide (P28) containing a minimum CR2 binding domain. Each immunogen was expressed from DNA plasmid in vivo or injected as purified recombinant protein. The fusion of the P28 peptide to Env(gp120) enhanced both humoral and cell-mediated immune responses with similar efficiency as Env(gp120) conjugated to C3d. The fusion of C3d or P28 to Env(gp120) elicited higher-titer anti-Env specific antibody, enhanced avidity maturation of the elicited antibody, and elicited higher numbers of IFN-gamma and IL-4 secreting cells compared to Env(gp120) immunizations. This CR2-binding domain specific 28 amino acid peptide can substitute for the entire C3d molecule and enhance immunity. These results indicate that the adjuvant properties of C3d are associated with CR2 interaction.
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http://dx.doi.org/10.1007/0-387-34134-X_17DOI Listing
February 2007

HIV-1 Envgp140 trimers elicit neutralizing antibodies without efficient induction of conformational antibodies.

Vaccine 2006 Jun 3;24(26):5442-51. Epub 2006 Apr 3.

University of Pittsburgh, School of Medicine, Department of Medicine, Division of Infectious Diseases, Scaife Hall, Room S871, 3550 Terrace Street, Pittsburgh, PA 15261, USA.

Currently, no vaccine for human immunodeficiency virus (HIV-1) provides protection from virus infection. One reason for these disappointing results has been the difficulty of current vaccine candidates to elicit high-titer, broadly reactive immunity to a large number of viral proteins. Recently, our laboratory demonstrated that the coupling of C3d to a soluble trimerized HIV-1 envelope (Env(gp140(FT))) elicited higher titers of neutralizing antibodies than monomers of Env(gp120) coupled to C3d [Bower JF, Yang X, Sodroski J, Ross TM. Elicitation of neutralizing antibodies with DNA vaccines expressing soluble stabilized human immunodeficiency virus type 1 envelope glycoprotein trimers conjugated to C3d. J Virol 2004;78(9):4710-9]. To determine if the induction of conformational antibodies correlated with neutralization, mice (BALB/c) were primed (2x) with DNA plasmids expressing monomeric Env(gp120) or trimeric Env(gp140) alone or fused to mC3d(3) at one of two doses (2.0microg or 0.2microg), followed by a boost of recombinant uncleaved, trimeric Env(gp140). Regardless of the priming dose of DNA, all mice had high-titer anti-Env IgG antibodies. Interestingly, Env(gp140) trimers did not elicit higher titers of antibodies that recognized conformational Env epitopes compared to monomers of Env(gp120). Therefore, additional parameters were examined for correlation with neutralization. For neutralization-resistant HIV-1 isolates, ADA and YU-2, neutralization correlated with high-titer, high avidity antibodies, with Env(gp140) eliciting slightly higher neutralization titers than Env(gp120). In contrast, none of the measured parameters correlated with neutralization for the more neutralization-sensitive isolates, MN or 89.6. Therefore, even though soluble, uncleaved Env(gp140) trimers may be marginally more effective at eliciting neutralizing antibodies than Env(gp120), neutralization does not appear to correlate with the elicitation of conformationally dependent antibodies.
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http://dx.doi.org/10.1016/j.vaccine.2006.03.063DOI Listing
June 2006

Differences in transport of fatty acids and expression of fatty acid transporting proteins in adipose tissue of obese black and white women.

Am J Physiol Endocrinol Metab 2006 Jan;290(1):E87-E91

Department of Internal Medicine, Brody School of Medicine, East Carolina University, Greenville, NC 27834, USA.

We have reported that the rate of de novo triglyceride (TG) synthesis by omental, but not subcutaneous, adipose tissue was higher in African-American women (AAW) than in Caucasian women (CAW). The purpose of this study was to explore the potential mechanisms underlying this increase. Toward that end, we determined the activities of key enzymes in the pathway of TG synthesis, the rates of uptake of fatty acids by adipocytes, mRNA and protein levels of the fatty acid-transporting proteins FAT/CD36 and FATP, and mRNA and protein levels of PPARgamma in omental fat of AAW and CAW. The results showed 1) no difference in the activity of phosphofructokinase, glycerol-3-phosphate dehydrogenase, or diacylglycerol acyltransferase; 2) a higher rate of fatty acid uptake by adipocytes of the AAW; 3) an increase in the mRNA and protein levels of CD36 and FATP4 in the fat of the AAW; and 4) an increase in the mRNA and protein levels of PPARgamma, which can stimulate the expression of CD36 and FATP. These results suggest that the increase in the transport of fatty acid, which is mediated by the overexpression of the transport proteins in the omental adipose tissue of the AAW, might contribute to the higher prevalence of obesity in AAW.
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http://dx.doi.org/10.1152/ajpendo.00194.2005DOI Listing
January 2006

C3d enhances immune responses using low doses of DNA expressing the HIV-1 envelope from codon-optimized gene sequences.

Curr HIV Res 2005 Apr;3(2):191-8

Department of Medicine, Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA.

DNA vaccinations effectively induce both humoral and cellular immune responses to immunogens from diverse infectious agents. However, DNA vaccines expressing the HIV-1 envelope glycoprotein (Env) are poorly immunogenic when expressed from wild-type (wt) DNA sequences. Two recent approaches used to enhance the immunogenicity of Env expressed from a DNA vaccine are the fusion of the molecular adjuvant, C3d, to a soluble form of Env and the use of codon-optimized (co) env gene inserts. Independently, each approach enhances antibody titer and cellular responses against Env expressed from gene inserts. The goal of this study was to examine if both codon-optimization of env gene inserts and C3d conjugation to Env could function in a synergistic manner to enhance immunogenicity. Mice (BALB/c) were inoculated with decreasing doses (2.0 microg, 0.2 microg or 0.02 microg) of co DNA expressing Env alone or fused to three copies of murine C3d (mC3d3) gene. Mice vaccinated with the highest dose (2.0 microg) of DNA had high anti-Env specific antibody titers regardless of the addition of mC3d3. At lower doses (0.2 microg and 0.02 microg) of DNA, mice vaccinated with Env-mC3d3 had enhanced immune responses compared to mice vaccinated with DNA expressing Env only. In addition, mice vaccinated with Env-mC3d3 at the highest doses of DNA had enhanced interleukin-4 secreting cells, while mice vaccinated with the lowest dose of DNA had enhanced interferon-gamma secreting cells. Therefore, both codon-optimization of env sequences and C3d conjugation to Env appear to enhance anti-Env antibodies in an independent and additive manner.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1266284PMC
http://dx.doi.org/10.2174/1570162053506937DOI Listing
April 2005

Unique V3 loop sequence derived from the R2 strain of HIV-type 1 elicits broad neutralizing antibodies.

AIDS Res Hum Retroviruses 2004 Nov;20(11):1259-68

Department of Medicine, Division of Infectious Diseases, University of Pittsburgh, School of Medicine, Pittsburgh, Pennsylvania 15261, USA.

DNA vaccines expressing the envelope (Env) of the human immunodeficiency virus type 1 (HIV-1) have been relatively ineffective at generating high-titer, long-lasting, neutralizing antibodies. In this study, DNA vaccines were constructed to express the gp120 subunit of Env from the isolate HIV-1(R2) using both wild-type and codon-optimized gene sequences. Three copies of the murine C3d were added to the carboxyl terminus to enhance the immunogenicity of the expressed fusion protein. Mice (BALB/c) vaccinated with DNA plasmid expressing the gp120(R2) using codon-optimized Env sequences elicited high-titer anti-Env antibodies regardless of conjugation to C3d. In contrast, only mice vaccinated with DNA using wild-type gp120(R2) sequences fused to mC3d(3), had detectable anti-Env antibodies. Interestingly, mice vaccinated with DNA expressing gp120(R2) from codon-optimized sequences elicited antibodies that neutralized both homologous and heterologous HIV-1 isolates. To determine if the unique sequence found in the crown of the V3 loop of the Env(R2) was responsible for the elicitation of the cross-clade neutralizing antibodies, the codons encoding for the Pro-Met (amino acids 313-314) were introduced into the sequences encoding the gp120(ADA) (R5) or gp120(89.6) (R5X4). Mice vaccinated with gp120(ADA)-mC3d(3)-DNA with the Pro-Met mutation had antibodies that neutralized HIV-1 infection, but not the gp120(89.6)-mC3d(3)-DNA. Therefore, the use of the unique sequences in the Env(R2) introduced into an R5 tropic envelope, in conjunction with C3d fusion, was effective at broadening the number of viruses that could be neutralized. However, the introduction of this same sequence into an R5X4-tropic envelope was ineffective in eliciting improved cross-clade neutralizing antibodies.
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http://dx.doi.org/10.1089/aid.2004.20.1259DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1550980PMC
November 2004

DNA vaccines expressing soluble CD4-envelope proteins fused to C3d elicit cross-reactive neutralizing antibodies to HIV-1.

Virology 2004 Oct;328(2):292-300

Department of Medicine, Division of Infectious Diseases, University of Pittsburgh, School of Medicine, Pittsburgh, PA 15261, USA.

DNA vaccines expressing the envelope (Env) of the human immunodeficiency virus type 1 (HIV-1) have been relatively ineffective at generating high-titer, long-lasting, neutralizing antibodies in a variety of animal models. In this study, DNA vaccines were constructed to express a fusion protein of the soluble human CD4 (sCD4) and the gp120 subunit of the HIV-1 envelope. To enhance the immunogenicity of the expressed fusion protein, three copies of the murine C3d (mC3d3) were added to the carboxyl terminus of the complex. Monoclonal antibodies that recognize CD4-induced epitopes on gp120 efficiently bound to sCD4-gp120 or sCD4-gp120-mC3d3. In addition, both sCD4-gp120 and sCD4-gp120-mC3d3 bound to cells expressing appropriate coreceptors in the absence of cell surface hCD4. Mice (BALB/c) vaccinated with DNA vaccines expressing either gp120-mC3d3 or sCD4-gp120-mC3d3 elicited antibodies that neutralized homologous virus infection. However, the use of sCD4-gp120-mC3d3-DNA elicited the highest titers of neutralizing antibodies that persisted after depletion of anti-hCD4 antibodies. Interestingly, only mice vaccinated with DNA expressing sCD4-gp120-mC3d3 had antibodies that elicited cross-protective neutralizing antibodies. The fusion of sCD4 to the HIV-1 envelope exposes neutralizing epitopes that elicit broad protective immunity when the fusion complex is coupled with the molecular adjuvant, C3d.
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http://dx.doi.org/10.1016/j.virol.2004.07.031DOI Listing
October 2004

Cutting edge: C3d functions as a molecular adjuvant in the absence of CD21/35 expression.

J Immunol 2004 May;172(10):5833-7

Department of Immunology, Duke University Medical Center, Durham, NC 27710, USA.

Complement component C3 covalently attaches to Ags following activation, where the C3d cleavage fragment can function as a molecular adjuvant to augment humoral immune responses. C3d is proposed to exert its adjuvant-like activities by targeting Ags to the C3d receptor (CD21/35) expressed by B cells and follicular dendritic cells. To directly assess the importance of CD21/35 in mediating the immunostimulatory effects of C3d, CD21/35-deficient (CD21/35(-/-)) mice were immunized with streptavidin (SA), SA-C3dg tetramers, recombinant HIV gp120 (gp120), or gp120 fused with linear multimers of C3d. Remarkably, SA- and gp120-specific Ab responses were significantly augmented in CD21/35(-/-) mice when these Ags were complexed with C3d in comparison to Ag alone. In fact, primary and secondary Ab responses and Ab-forming cell responses of CD21/35(-/-) mice approached those of wild-type mice immunized with SA-C3dg and gp120-C3d. Thus, C3d can function as a molecular adjuvant in the absence of CD21/35 expression.
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http://dx.doi.org/10.4049/jimmunol.172.10.5833DOI Listing
May 2004

Elicitation of neutralizing antibodies with DNA vaccines expressing soluble stabilized human immunodeficiency virus type 1 envelope glycoprotein trimers conjugated to C3d.

J Virol 2004 May;78(9):4710-9

Department of Medicine, Division of Infectious Diseases, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261, USA.

DNA vaccines expressing the envelope (Env) of human immunodeficiency virus type 1 (HIV-1) have been relatively ineffective at generating high-titer, long-lasting immune responses. Oligomeric or trimeric (gp140) forms of Env that more closely mimic the native proteins on the virion are often more effective immunogens than monomeric (gp120) envelopes. In this study, several forms of Env constructed from the HIV-1 isolate YU-2 (HIV-1(YU-2)) were tested for their immunogenic potential: a trimeric form of uncleaved (-) Env stabilized with a synthetic trimer motif isolated from the fibritin (FT) protein of the T4 bacteriophage, sgp140(YU-2)(-/FT), was compared to sgp140(YU-2)(-) without a synthetic trimerization domain, as well as to monomeric gp120(YU-2). DNA plasmids were constructed to express Env alone or fused to various copies of murine C3d (mC3d). BALB/c mice were vaccinated (day 1 and week 4) with DNA expressing a codon-optimized envelope gene insert, alone or fused to mC3d. Mice were subsequently boosted (week 8) with the DNA or recombinant Env protein. All mice had high anti-Env antibody titers regardless of the use of mC3d. Sera from mice vaccinated with DNA expressing non-C3d-fused trimers elicited neutralizing antibodies against homologous HIV-1(YU-2) virus infection in vitro. In contrast, sera from mice inoculated with DNA expressing Env-C3d protein trimers elicited antibody that neutralized both homologous HIV-1(YU-2) and heterologous HIV-1(ADA), albeit at low titers. Therefore, DNA vaccines expressing trimeric envelopes coupled to mC3d, expressed in vivo from codon-optimized sequences, elicit low titers of neutralizing antibodies against primary isolates of HIV-1.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC387675PMC
http://dx.doi.org/10.1128/jvi.78.9.4710-4719.2004DOI Listing
May 2004

Cholesteryl ester transfer protein expression prevents diet-induced atherosclerotic lesions in male db/db mice.

Arterioscler Thromb Vasc Biol 2003 Aug 5;23(8):1412-5. Epub 2003 Jun 5.

Department of Biochemistry, Brody School of Medicine, East Carolina University, Greenville, NC, USA.

Objective: Accompanying more atherogenic lipoprotein profiles and an increased incidence of atherosclerosis, plasma cholesteryl ester transfer protein (CETP) is depressed in diabetic obese patients compared with nondiabetic obese counterparts. The depressed levels of CETP in the plasma of diabetic obese individuals may contribute to the development of an atherogenic lipoprotein profile and atherogenesis. We have examined the effect of CETP expression on vascular health in the db/db model of diabetic obesity.

Methods And Results: Transgenic mice expressing the human CETP minigene were crossed with db/db strain, and 3 groups of offspring (CETP, db, and db/CETP) were placed on an atherogenic diet for 16 weeks. The proximal aorta was then excised and examined for the presence of atherosclerotic plaques. In db mice, 9 of 11 had intimal lesions with a mean area of 26 098+/-7486 microm2. No lesions greater than 1000 microm2 were observed in db/CETP or CETP mice. CETP-expressing mice had lower circulating cholesterol concentrations than db mice. Fractionating plasma lipids by FPLC indicated that the difference in total cholesterol was primarily attributable to differences in VLDL and LDL.

Conclusions: The expression of human CETP in db/db mice prevented the formation of diet-induced lesions, suggesting an antiatherogenic effect of CETP in the context of diabetic obesity.
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http://dx.doi.org/10.1161/01.ATV.0000080687.94313.67DOI Listing
August 2003

Differences in the lipolytic function of adipose tissue preparations from Black American and Caucasian women.

Metabolism 2002 Nov;51(11):1514-8

Department of Biochemistry, East Carolina University, Greenville, NC 27858, USA.

The purpose of this study was to determine the potential causes of the lower lipolytic rates in obese Black American women compared to obese Caucasian women. Subcutaneous and omental adipose tissue were obtained from subjects during abdominal surgery, and hormone-sensitive lipase (HSL) mass, mRNA, and activity were determined. HSL mRNA levels did not differ between the Black American and Caucasian women in either subcutaneous or omental adipose tissue. However, HSL mass was approximately 35% lower (P <.05) in both subcutaneous and omental adipose tissue of the Black Americans. Because of these differences, we measured HSL activity in frozen subcutaneous and omental adipose tissue, and also measured basal and isoproterenol-stimulated lipolytic rates in tissue fragments. No racial differences were found in the activity of HSL in either subcutaneous or omental adipose tissue. However, basal lipolytic rates in the Black Americans were 53% and 44% lower (P <.05) in the subcutaneous and omental fat, respectively, compared to the Caucasian women, despite a lack of difference in cell size between the 2 groups. Interestingly, the degree of stimulation by isoproterenol was higher in both the subcutaneous and omental adipose tissue of the Black American than those of the Caucasian women, resulting in equal stimulation by isoproterenol in the 2 groups. These results indicate that despite the lower mass and lower basal HSL activity in the obese Black American women, stimulation of HSL results in equal activity of the enzyme in the 2 races. This suggests that the signaling pathway of HSL stimulation is more efficient in the Black American women.
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http://dx.doi.org/10.1053/meta.2002.35589DOI Listing
November 2002

Disruptive change. When trying harder is part of the problem.

Harv Bus Rev 2002 May;80(5):94-101, 134

Harvard Business School, Boston, USA.

When a company faces a major disruption in its markets, managers' perceptions of the disruption influence how they respond to it. If, for instance, they view the disruption as a threat to their core business, managers tend to overreact, committing too many resources too quickly. But if they see it as an opportunity, they're likely to commit insufficient resources to its development. Clark Gilbert and Joseph Bower explain why thinking in such stark terms--threat or opportunity--is dangerous. It's possible, they argue, to arrive at an organizational framing that makes good use of the adrenaline a threat creates as well as of the creativity an opportunity affords. The authors claim that the most successful companies frame the challenge differently at different times: When resources are being allocated, managers see the disruptive innovation as a threat. But when the hard strategic work of discovering and responding to new markets begins, the disruptive innovation is treated as an opportunity. The ability to reframe the disruptive technology as circumstances evolve is not an easy skill to master, the authors admit. In fact, it might not be possible without adjusting the organizational structure and the processes governing new business funding. Successful companies, the authors have determined, tend to do certain things: They establish a new venture separate from the core business; they fund the venture in stages as markets emerge; they don't rely on employees from the core organization to staff the new business; and they appoint an active integrator to manage the tensions between the two organizations, to name a few. This article will help executives frame innovations in more balanced ways--allowing them to recognize threats but also to seize opportunities.
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May 2002

Ethnic differences in postprandial triglyceride response to a fatty meal and lipoprotein lipase in lean and obese African American and Caucasian women.

Metabolism 2002 Feb;51(2):211-7

Department of Biochemistry, East Carolina University School of Medicine, Greenville, NC 27858, USA.

The purpose of this study was to determine if there were differences in the expression of lipoprotein lipase (LPL) in African American (AA) and Caucasian (CA) women. LPL mRNA and protein levels were determined in subcutaneous and omental fat of lean and obese subject from the 2 races (4 groups; 12 to 15 subjects/group). LPL mRNA levels of lean AA were not different from the lean CA women in either fat depot. LPL mRNA levels in the subcutaneous fat of the obese AA were higher than those of CA women (1.3 +/- 0.1 v 0.86 +/- 0.06, P.05), but not different in omental fat. LPL mass in subcutaneous fat of lean AA was higher (0.95 +/- 0.09 v 0.64 +/- 0.06, P.05), but not different in omental fat from the CA women. LPL mass in subcutaneous and omental fat was not different in the 2 obese groups. Differences in the activity of LPL were evaluated by (1) measuring the increments of triglycerides (TG) at 2, 4, 6, and 8 hours after a fat-rich meal and (2) by measuring postheparin plasma lipolytic activity. Plasma TG levels in the lean AA were lower than those of the lean CA women at basal and at 2, 4, 6, and 8 hours postprandially. The increase in TG levels at 2 hours tended to be lower in the AA than the CA women, was significantly lower at 4 hours (24 +/- 5 v 45 +/- 7, P.05), and was not different 8 hours postprandially. No differences were observed in either the absolute or the incremental concentrations of TG in the obese groups. Postheparin plasma LPL activity was higher in the lean AA than the lean CA women (4.8 +/- 0.4 v 3.4 +/- 0.4, P.05), but not different in the obese groups. These results indicate that the lower TG concentrations in the lean AA women may be partly due to enhanced expression, activity, and intravascular availability of LPL. Furthermore, it appears that the racial differences in expression and function of LPL are attenuated with obesity.
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http://dx.doi.org/10.1053/meta.2002.29991DOI Listing
February 2002