Robert Whalen - Academia.edu (original) (raw)

Papers by Robert Whalen

Research paper thumbnail of Influence of the dwarf mouse mutation on skeletal and cardiac myosin isoforms. Effect of one injection of thyroxine on skeletal and cardiac muscle phenotype

Journal of Biological Chemistry, 1987

Research paper thumbnail of Probing an HIV-1 virion library containing randomly recombined clade B envelope glycoproteins reveals a surprising diversity of phenotypes

Research paper thumbnail of Neonatal and adult myosin heavy chain isoforms in a nerve-muscle culture system

The Journal of Cell Biology, 1986

When adult mouse muscle fibers are co-cultured with embryonic mouse spinal cord, the muscle regen... more When adult mouse muscle fibers are co-cultured with embryonic mouse spinal cord, the muscle regenerates to form myotubes that develop cross-striations and contractions. We have investigated the myosin heavy chain (MHC) isoforms present in these cultures using polyclonal antibodies to the neonatal, adult fast, and slow MHC isoforms of rat (all of which were shown to react specifically with the analogous mouse isoforms) in an immunocytochemical assay. The adult fast MHC was absent in newly formed myotubes but was found at later times, although it was absent when the myotubes myotubes were cultured without spinal cord tissue. When nerve-induced muscle contractions were blocked by the continuous presence of alpha-bungarotoxin, there was no decrease in the proportion of fibers that contained adult fast MHC. Neonatal and slow MHC were found at all times in culture, even in the absence of the spinal cord, and so their expression was not thought to be nerve-dependent. Thus, in this culture ...

Research paper thumbnail of A Directed Molecular Evolution Approach to Improved Immunogenicity of the HIV-1 Envelope Glycoprotein

PLoS ONE, 2011

A prophylactic vaccine is needed to slow the spread of HIV-1 infection. Optimization of the wild-... more A prophylactic vaccine is needed to slow the spread of HIV-1 infection. Optimization of the wild-type envelope glycoproteins to create immunogens that can elicit effective neutralizing antibodies is a high priority. Starting with ten genes encoding subtype B HIV-1 gp120 envelope glycoproteins and using in vitro homologous DNA recombination, we created chimeric gp120 variants that were screened for their ability to bind neutralizing monoclonal antibodies. Hundreds of variants were identified with novel antigenic phenotypes that exhibit considerable sequence diversity. Immunization of rabbits with these gp120 variants demonstrated that the majority can induce neutralizing antibodies to HIV-1. One novel variant, called ST-008, induced significantly improved neutralizing antibody responses when assayed against a large panel of primary HIV-1 isolates. Further study of various deletion constructs of ST-008 showed that the enhanced immunogenicity results from a combination of effective DNA priming, an enhanced V3-based response, and an improved response to the constant backbone sequences.

Research paper thumbnail of Individual Analysis of Mice Vaccinated against a Weakly Immunogenic Self Tumor-Specific Antigen Reveals a Correlation between CD8 T Cell Response and Antitumor Efficacy

The Journal of Immunology, 2003

The weakly immunogenic murine P1A Ag is a useful experimental model for the development of new va... more The weakly immunogenic murine P1A Ag is a useful experimental model for the development of new vaccination strategies that could potentially be used against human tumors. An i.m. DNA-based immunization procedure, consisting of three inoculations with the P1A-coding pBKCMV-P1A plasmid at 10-day intervals, resulted in CTL generation in all treated BALB/c mice. Surprisingly, gene gun skin bombardment with the pBKCMV-P1A vector did not induce CTL, nor was it protective against a lethal challenge with the syngeneic P1A-positive J558 tumor cell line. To speed up the immunization procedure, we pretreated the tibialis anterior muscles with cardiotoxin, which induces degeneration of myocytes while sparing immature satellite cells. The high muscle-regenerative activity observable after cardiotoxin inoculation was associated with infiltration of inflammatory cells and expression of proinflammatory cytokines. A single pBKCMV-P1A plasmid inoculation in cardiotoxin-treated BALB/c mice allowed for...

Research paper thumbnail of Effects of hypothyroidism on myosin isozyme transitions in developing rat muscle

FEBS Letters, 1984

Hypothyroidism was induced in young rats by methylthiouracil treatment of pregnant mothers from 1... more Hypothyroidism was induced in young rats by methylthiouracil treatment of pregnant mothers from 18 days of gestation to 4 weeks after birth. Electrophoretic analysis of native myosin isozymes revealed a persistence of neonatal and embryonic myosin in developing fast and slow muscles up to at least 28 days after birth. The appearance of adult fast myosin was inhibited in 28-day old animals, however adult slow myosin was found in the soleus muscle. Immunocytochemical results on the soleus demonstrate a cellular heterogeneity in the response to hypothyroidism. About half fibers have a normal complement of slow myosin and do not contain neonatal myosin. Only the remaining fibers contain the large amounts of neonatal myosin demonstrated by electrophoresis. Immunocytochemistry Native myosin isozyme Methylthiouracil Soleus Gastrocnemius

Research paper thumbnail of Myosin isozyme changes in the heart following constriction of the ascending aorta of a 25-day old rat

FEBS Letters, 1983

When a constricting band is placed around the ascending aorta of young (25-day old) rats, all cha... more When a constricting band is placed around the ascending aorta of young (25-day old) rats, all chambers of the heart eventually produce hypertrophy. Both the left and right ventricles show strong shifts toward an isozyme pattern in which V3 is predominant, similar to that seen in models where hypertrophy is induced in adult rats. The hypertrophied atria however, show no detectable change in the native myosin isozymes or the light chain subunits. Myosin isozyme Light chain Cardiac hypertrophy Adaptation

Research paper thumbnail of DNA Vaccines for Emerging Infectious Diseases: What If?

Emerging Infectious Diseases, 1996

Research paper thumbnail of Effects of partially dismantling the CD4 binding site glycan fence of HIV-1 Envelope glycoprotein trimers on neutralizing antibody induction

Virology, 2017

Previously, VLPs bearing JR-FL strain HIV-1 Envelope trimers elicited potent neutralizing antibod... more Previously, VLPs bearing JR-FL strain HIV-1 Envelope trimers elicited potent neutralizing antibodies (nAbs) in 2/8 rabbits (PLoS Pathog 11(5): e1004932) by taking advantage of a naturally absent glycan at position 197 that borders the CD4 binding site (CD4bs). In new immunizations, we attempted to improve nAb responses by removing the N362 glycan that also lines the CD4bs. All 4 rabbits developed nAbs. One targeted the N197 glycan hole like our previous sera. Two sera depended on the N463 glycan, again suggesting CD4bs overlap. Heterologous boosts appeared to reduce nAb clashes with the N362 glycan. The fourth serum targeted a N362 glycan-sensitive epitope. VLP manufacture challenges prevented us from immunizing larger rabbit numbers to empower a robust statistical analysis. Nevertheless, trends suggest that targeted glycan removal may improve nAb induction by exposing new epitopes and that it may be possible to modify nAb specificity using rational heterologous boosts.

Research paper thumbnail of NASA symposium The influence of gravity and activity on muscle and bone January 29?30, 1990

Research paper thumbnail of Modeling the epitope footprints of neutralizing sera 613 and 647

Research paper thumbnail of Overview of human plasmas and animal sera

Research paper thumbnail of Direct gene transfer in skeletal muscle: plasmid DNA-based immunization against the hepatitis B virus surface antigen

Vaccine, 1994

9ene transfer by intramuscular injection of plasmid DNA encodin9 an antioenic protein may be used... more 9ene transfer by intramuscular injection of plasmid DNA encodin9 an antioenic protein may be used for the purpose of immunization. DNA-based immunization may be of value for basic immunological research and vaccine development. Several factors influence the uptake and expression of plasmid DNA in skeletal muscle, which in turn influence the immune response to the expressed protein. Physical barriers and other factors may impede diffusion of the DNA within the muscle tissue or its entry into the muscle fibres. Although the efficiency of 9ene transfer in normal mouse muscle is low (< 100 fibres per injection site), a humoral response to the hepatitis B surface antigen (HBsAg) is obtained after expression of a transferred 9ene. Direct 9ene transfer is ten times more efficient in regeneratin9 than in normal mouse muscle. DNA-based immunization in such regeneratin9 muscles results in an earlier and stronger humoral response to HBsA 9 than is seen in normal mature muscle. A needleless jet injection system (Biojector ®) isable to deliver DNA into normal muscle in rats and rabbits such that a substantial immune response is obtained.

Research paper thumbnail of The expression of slow myosin during mammalian somitogenesis and limb bud differentiation

The Journal of Cell Biology, Dec 1, 1988

The developmental pattern of slow myosin expression has been studied in mouse embryos from the so... more The developmental pattern of slow myosin expression has been studied in mouse embryos from the somitic stage to the period of secondary fiber formation and in myogenic cells, cultured from the same developmental stages.

Research paper thumbnail of The phosphorylation of diphosphoglycerate mutase

Journal of Biological Chemistry, Mar 10, 1973

Research paper thumbnail of Analysis of U.S. and Soviet efforts to maintain muscle and bone mass with exercise during prolonged bedrest and spaceflight

Research paper thumbnail of Hepatitis B Virus Vaccines

Research paper thumbnail of Myosin isozyme transitions ocurring during the postnatal development of the rat soleus muscle

Developmental Biology, 1984

Research paper thumbnail of Domain and residue-level mapping of potent rabbit sera

Research paper thumbnail of Androgen-induced receptivity

Research paper thumbnail of Influence of the dwarf mouse mutation on skeletal and cardiac myosin isoforms. Effect of one injection of thyroxine on skeletal and cardiac muscle phenotype

Journal of Biological Chemistry, 1987

Research paper thumbnail of Probing an HIV-1 virion library containing randomly recombined clade B envelope glycoproteins reveals a surprising diversity of phenotypes

Research paper thumbnail of Neonatal and adult myosin heavy chain isoforms in a nerve-muscle culture system

The Journal of Cell Biology, 1986

When adult mouse muscle fibers are co-cultured with embryonic mouse spinal cord, the muscle regen... more When adult mouse muscle fibers are co-cultured with embryonic mouse spinal cord, the muscle regenerates to form myotubes that develop cross-striations and contractions. We have investigated the myosin heavy chain (MHC) isoforms present in these cultures using polyclonal antibodies to the neonatal, adult fast, and slow MHC isoforms of rat (all of which were shown to react specifically with the analogous mouse isoforms) in an immunocytochemical assay. The adult fast MHC was absent in newly formed myotubes but was found at later times, although it was absent when the myotubes myotubes were cultured without spinal cord tissue. When nerve-induced muscle contractions were blocked by the continuous presence of alpha-bungarotoxin, there was no decrease in the proportion of fibers that contained adult fast MHC. Neonatal and slow MHC were found at all times in culture, even in the absence of the spinal cord, and so their expression was not thought to be nerve-dependent. Thus, in this culture ...

Research paper thumbnail of A Directed Molecular Evolution Approach to Improved Immunogenicity of the HIV-1 Envelope Glycoprotein

PLoS ONE, 2011

A prophylactic vaccine is needed to slow the spread of HIV-1 infection. Optimization of the wild-... more A prophylactic vaccine is needed to slow the spread of HIV-1 infection. Optimization of the wild-type envelope glycoproteins to create immunogens that can elicit effective neutralizing antibodies is a high priority. Starting with ten genes encoding subtype B HIV-1 gp120 envelope glycoproteins and using in vitro homologous DNA recombination, we created chimeric gp120 variants that were screened for their ability to bind neutralizing monoclonal antibodies. Hundreds of variants were identified with novel antigenic phenotypes that exhibit considerable sequence diversity. Immunization of rabbits with these gp120 variants demonstrated that the majority can induce neutralizing antibodies to HIV-1. One novel variant, called ST-008, induced significantly improved neutralizing antibody responses when assayed against a large panel of primary HIV-1 isolates. Further study of various deletion constructs of ST-008 showed that the enhanced immunogenicity results from a combination of effective DNA priming, an enhanced V3-based response, and an improved response to the constant backbone sequences.

Research paper thumbnail of Individual Analysis of Mice Vaccinated against a Weakly Immunogenic Self Tumor-Specific Antigen Reveals a Correlation between CD8 T Cell Response and Antitumor Efficacy

The Journal of Immunology, 2003

The weakly immunogenic murine P1A Ag is a useful experimental model for the development of new va... more The weakly immunogenic murine P1A Ag is a useful experimental model for the development of new vaccination strategies that could potentially be used against human tumors. An i.m. DNA-based immunization procedure, consisting of three inoculations with the P1A-coding pBKCMV-P1A plasmid at 10-day intervals, resulted in CTL generation in all treated BALB/c mice. Surprisingly, gene gun skin bombardment with the pBKCMV-P1A vector did not induce CTL, nor was it protective against a lethal challenge with the syngeneic P1A-positive J558 tumor cell line. To speed up the immunization procedure, we pretreated the tibialis anterior muscles with cardiotoxin, which induces degeneration of myocytes while sparing immature satellite cells. The high muscle-regenerative activity observable after cardiotoxin inoculation was associated with infiltration of inflammatory cells and expression of proinflammatory cytokines. A single pBKCMV-P1A plasmid inoculation in cardiotoxin-treated BALB/c mice allowed for...

Research paper thumbnail of Effects of hypothyroidism on myosin isozyme transitions in developing rat muscle

FEBS Letters, 1984

Hypothyroidism was induced in young rats by methylthiouracil treatment of pregnant mothers from 1... more Hypothyroidism was induced in young rats by methylthiouracil treatment of pregnant mothers from 18 days of gestation to 4 weeks after birth. Electrophoretic analysis of native myosin isozymes revealed a persistence of neonatal and embryonic myosin in developing fast and slow muscles up to at least 28 days after birth. The appearance of adult fast myosin was inhibited in 28-day old animals, however adult slow myosin was found in the soleus muscle. Immunocytochemical results on the soleus demonstrate a cellular heterogeneity in the response to hypothyroidism. About half fibers have a normal complement of slow myosin and do not contain neonatal myosin. Only the remaining fibers contain the large amounts of neonatal myosin demonstrated by electrophoresis. Immunocytochemistry Native myosin isozyme Methylthiouracil Soleus Gastrocnemius

Research paper thumbnail of Myosin isozyme changes in the heart following constriction of the ascending aorta of a 25-day old rat

FEBS Letters, 1983

When a constricting band is placed around the ascending aorta of young (25-day old) rats, all cha... more When a constricting band is placed around the ascending aorta of young (25-day old) rats, all chambers of the heart eventually produce hypertrophy. Both the left and right ventricles show strong shifts toward an isozyme pattern in which V3 is predominant, similar to that seen in models where hypertrophy is induced in adult rats. The hypertrophied atria however, show no detectable change in the native myosin isozymes or the light chain subunits. Myosin isozyme Light chain Cardiac hypertrophy Adaptation

Research paper thumbnail of DNA Vaccines for Emerging Infectious Diseases: What If?

Emerging Infectious Diseases, 1996

Research paper thumbnail of Effects of partially dismantling the CD4 binding site glycan fence of HIV-1 Envelope glycoprotein trimers on neutralizing antibody induction

Virology, 2017

Previously, VLPs bearing JR-FL strain HIV-1 Envelope trimers elicited potent neutralizing antibod... more Previously, VLPs bearing JR-FL strain HIV-1 Envelope trimers elicited potent neutralizing antibodies (nAbs) in 2/8 rabbits (PLoS Pathog 11(5): e1004932) by taking advantage of a naturally absent glycan at position 197 that borders the CD4 binding site (CD4bs). In new immunizations, we attempted to improve nAb responses by removing the N362 glycan that also lines the CD4bs. All 4 rabbits developed nAbs. One targeted the N197 glycan hole like our previous sera. Two sera depended on the N463 glycan, again suggesting CD4bs overlap. Heterologous boosts appeared to reduce nAb clashes with the N362 glycan. The fourth serum targeted a N362 glycan-sensitive epitope. VLP manufacture challenges prevented us from immunizing larger rabbit numbers to empower a robust statistical analysis. Nevertheless, trends suggest that targeted glycan removal may improve nAb induction by exposing new epitopes and that it may be possible to modify nAb specificity using rational heterologous boosts.

Research paper thumbnail of NASA symposium The influence of gravity and activity on muscle and bone January 29?30, 1990

Research paper thumbnail of Modeling the epitope footprints of neutralizing sera 613 and 647

Research paper thumbnail of Overview of human plasmas and animal sera

Research paper thumbnail of Direct gene transfer in skeletal muscle: plasmid DNA-based immunization against the hepatitis B virus surface antigen

Vaccine, 1994

9ene transfer by intramuscular injection of plasmid DNA encodin9 an antioenic protein may be used... more 9ene transfer by intramuscular injection of plasmid DNA encodin9 an antioenic protein may be used for the purpose of immunization. DNA-based immunization may be of value for basic immunological research and vaccine development. Several factors influence the uptake and expression of plasmid DNA in skeletal muscle, which in turn influence the immune response to the expressed protein. Physical barriers and other factors may impede diffusion of the DNA within the muscle tissue or its entry into the muscle fibres. Although the efficiency of 9ene transfer in normal mouse muscle is low (< 100 fibres per injection site), a humoral response to the hepatitis B surface antigen (HBsAg) is obtained after expression of a transferred 9ene. Direct 9ene transfer is ten times more efficient in regeneratin9 than in normal mouse muscle. DNA-based immunization in such regeneratin9 muscles results in an earlier and stronger humoral response to HBsA 9 than is seen in normal mature muscle. A needleless jet injection system (Biojector ®) isable to deliver DNA into normal muscle in rats and rabbits such that a substantial immune response is obtained.

Research paper thumbnail of The expression of slow myosin during mammalian somitogenesis and limb bud differentiation

The Journal of Cell Biology, Dec 1, 1988

The developmental pattern of slow myosin expression has been studied in mouse embryos from the so... more The developmental pattern of slow myosin expression has been studied in mouse embryos from the somitic stage to the period of secondary fiber formation and in myogenic cells, cultured from the same developmental stages.

Research paper thumbnail of The phosphorylation of diphosphoglycerate mutase

Journal of Biological Chemistry, Mar 10, 1973

Research paper thumbnail of Analysis of U.S. and Soviet efforts to maintain muscle and bone mass with exercise during prolonged bedrest and spaceflight

Research paper thumbnail of Hepatitis B Virus Vaccines

Research paper thumbnail of Myosin isozyme transitions ocurring during the postnatal development of the rat soleus muscle

Developmental Biology, 1984

Research paper thumbnail of Domain and residue-level mapping of potent rabbit sera

Research paper thumbnail of Androgen-induced receptivity