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Papers by adrienne manning

International Journal of Neonatal Screening, Nov 28, 2022

This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY

Biochemical Pharmacology, 1997

Oligodeoxynucleotide phosphorothioates (PS-oligos) are being studied as novel therapeutic agents ... more Oligodeoxynucleotide phosphorothioates (PS-oligos) are being studied as novel therapeutic agents based on their ability to inhibit gene expression. Preclinical studies produced unanticipated complement and coagulation effects in monkeys receiving high-dose PS-oligo. In the present in viao studies, PS-oligo inhibited normal human blood clotting as well as subsequent assays for prothrombin fragment PF,,, and hemolytic complement. PS-oligo treatment of normal donor plasma produced concentration-dependent prolongations of clotting times, with the activated partial thromboplastin time more sensitive than prothrombin time or thrombin clotting time. PS-oligo treatment of normal donor serum similarly reduced hemolytic complement activity in a concentration-dependent manner. Reduced hemolysis correlated with increased levels of complement fragment C4d. The anti-heparin drug protamine sulfate inhibited in vitro effects of PS-oligo in both complement and coagulation assays, suggesting that charged residues in intemucleotide linkages of PS-oligo mediated the observed activities. Therefore, oligonucleotides with varying intemucleotide linkages, nucleotide sequence, or secondary structure were compared. Both complement and coagulation effects appeared to be independent of nucleotide sequence but were strongly related to the nature of internucleotide linkages. Several of these modified oligonucleotides have been shown previously to retain potent antisense activity and thus may represent viable alternatives for antisense therapeutics.

We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and... more We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and 3′-5′-deoxyribonucleotide segments. Thermal melting studies of the phosphodiester MBOs (three 2′-5′ linkages at each end) with the complementary 3′-5′-DNA and-RNA target strands suggest that 2′-5′-ribonucleoside incorporation into 3′-5′-oligodeoxyribonucleotides reduces binding to the target strands compared with an all 3′-5′-oligodeoxyribonucleotide of the same sequence and length. Increasing the number of 2′-5′ linkages (from six to nine) further reduces binding to the DNA target strand more than the RNA target strand [Kandimalla,E.R. and Agrawal,S. (1996) Nucleic Acids Symp. Ser., 35, 125-126]. Phosphorothioate (PS) analogs of MBOs destabilize the duplex with the DNA target strand more than the duplex with the RNA target strand. Circular dichroism studies indicate that the duplexes of MBOs with the DNA and RNA target strands have spectral characteristics of both A-and B-type conformations. Compared with the control oligonucleotide, MBOs exhibit moderately higher stability against snake venom phosphodiesterase, S1 nuclease and in fetal calf serum. Although 2′-5′ modification does not evoke RNase H activity, this modification does not effect the RNase H activation property of the 3′-5′-deoxyribonucleotide segment adjacent to the modification. In vitro studies with MBOs suggest that they have lesser effects on cell proliferation, clotting prolongation and hemolytic complement lysis than do control PS oligodeoxyribonucleotides. PS analogs of MBOs show HIV-1 inhibition comparable with that of a control PS oligodeoxyribonucleotide with all 3′-5′ linkages. The current results suggest that a limited number of 2′-5′ linkages could be used in conjunction with PS oligonucleotides to further modulate the properties of antisense oligonucleotides as therapeutic agents.

Background: Methylotrophs are a diverse group of bacteria that can utilize single-carbon compound... more Background: Methylotrophs are a diverse group of bacteria that can utilize single-carbon compounds as a sole energy source, and are often catalase-positive. Known as environmental symbionts, they are emerging as disease-causing organisms in patients with CGD. Methods: We present a case of lymphadenitis due to Granulibacter bethesdensis, a facultative methylotroph, and review 8 other infections caused by methylotrophs in patients with CGD. Results: There have been 9 reported cases of infections due to methylotrophs in patients with CGD. Seven cases were due to G. bethesdensis, one was due to Acidomonas methanolica and one was due to a Methylobacter. In all cases, 16s rRNA gene sequencing was required for diagnosis. Conclusions: Methylotrophs are fastidious and difficult to identify. Although the mechanisms underlying susceptibility to infection with methylotrophs in CGD remain to be elucidated, these bacteria should be included in the spectrum of pathogens associated with infections ...

Nucleic Acids Research, 1995

Foldback triplex-forming oligonucleotides (FTFOs) that contain an abasic linker, [2-(4-aminobutyr... more Foldback triplex-forming oligonucleotides (FTFOs) that contain an abasic linker, [2-(4-aminobutyr-1-yl)-1,3-propanediol] (APD linker), in the Hoogsteen domain against pyrimidine bases of a C:G and a T:A base pair were studied for their relative stability and sequence specificity of triplex formation. In general, the APD linker has less destabilizing effect against a C:G base pair than a T:A base pair. Incorporation of three APD linker moieties resulted in decreased

Genetics in Medicine, 2012

Genetics in Medicine, 2011

To achieve clinical validation of cutoff values for newborn screening by tandem mass spectrometry... more To achieve clinical validation of cutoff values for newborn screening by tandem mass spectrometry through a worldwide collaborative effort. Methods: Cumulative percentiles of amino acids and acylcarnitines in dried blood spots of approximately 25-30 million normal newborns and 10,742 deidentified true positive cases are compared to assign clinical significance, which is achieved when the median of a disorder range is, and usually markedly outside, either the 99th or the 1st percentile of the normal population. The cutoff target ranges of analytes and ratios are then defined as the interval between selected percentiles of the two populations. When overlaps occur, adjustments are made to maximize sensitivity and specificity taking all available factors into consideration. Results: As of December 1, 2010, 130 sites in 45 countries have uploaded a total of 25,114 percentile data points, 565,232 analyte results of true positive cases with 64 conditions, and 5,341 cutoff values. The average rate of submission of true positive cases between December 1, 2008, and December 1, 2010, was 5.1 cases/day. This cumulative evidence generated 91 high and 23 low cutoff target ranges. The overall proportion of cutoff values within the respective See Acknowledgements for author affiliation.

JAMA, 2014

IMPORTANCE Newborn screening for severe combined immunodeficiency (SCID) using assays to detect T... more IMPORTANCE Newborn screening for severe combined immunodeficiency (SCID) using assays to detect T-cell receptor excision circles (TRECs) began in Wisconsin in 2008, and SCID was added to the national recommended uniform panel for newborn screened disorders in 2010. Currently 23 states, the District of Columbia, and the Navajo Nation conduct population-wide newborn screening for SCID. The incidence of SCID is estimated at 1 in 100,000 births. OBJECTIVES To present data from a spectrum of SCID newborn screening programs, establish population-based incidence for SCID and other conditions with T-cell lymphopenia, and document early institution of effective treatments. DESIGN Epidemiological and retrospective observational study. SETTING Representatives in states conducting SCID newborn screening were invited to submit their SCID screening algorithms, test performance data, and deidentified clinical and laboratory information regarding infants screened and cases with nonnormal results. I...

International Journal of Neonatal Screening

Newborn screening (NBS) laboratories cannot accurately compare mass spectrometry-derived results ... more Newborn screening (NBS) laboratories cannot accurately compare mass spectrometry-derived results and cutoff values due to differences in testing methodologies. The objective of this study was to assess harmonization of laboratory proficiency test (PT) results using quality control (QC) data. Newborn Screening Quality Assurance Program (NSQAP) QC and PT data reported from 302 laboratories in 2019 were used to compare results among laboratories. QC materials were provided as dried blood spot cards which included a base pool and the base pool enriched with specific concentrations of metabolites in a linear range. QC data reported by laboratories were regressed on QC data reported by the Centers for Disease Control and Prevention (CDC), and laboratory’s regression parameters were used to harmonize their PT result. In general, harmonization tended to reduce overall variation in PT data across laboratories. The metabolites glutarylcarnitine (C5DC), tyrosine, and phenylalanine were display...

The Journal of Pediatrics

OBJECTIVES To determine if children with congenital heart disease (CHD) have lower newborn T-cell... more OBJECTIVES To determine if children with congenital heart disease (CHD) have lower newborn T-cell receptor excision circles (TREC) levels than the general population and to evaluate if low TREC levels in newborns with CHD are associated with clinical complications such as hospitalization for infection. STUDY DESIGN The Connecticut Newborn Screening Program reported TREC levels for newborns with CHD delivered between October 2011 and September 2016 at 2 major Connecticut children's hospitals. TREC levels for children with CHD were compared with the general population. TREC levels and outcome measures, including hospitalization for infection, were compared. RESULTS We enrolled 575 participants with CHD in the study. The median TREC level for newborns with CHD was lower than the general population (180.1 copies/μL vs 312.5 copies/μL; P < .01). patients with CHD requiring hospitalization for infection had lower median TREC levels than their counterparts (143.0 copies/μL vs 186.7 copies/μL; P < .01). The combination of prematurity and low TREC level had a strong relationship to hospitalization for infection (area under the receiver operative characteristic curve of 0.89). There was no association between TREC level and CHD severity. CONCLUSIONS Newborns with CHD demonstrated lower TREC levels than the general population. Low TREC levels were associated with hospitalization for infection in preterm children with CHD. Study limitations include that this was a retrospective chart review. These findings may help to identify newborns with CHD at highest risk for infection, allowing for potential opportunities for intervention.

Journal of the American College of Cardiology

Routine childhood infections in patients with congenital heart disease (CHD) are a risk factor fo... more Routine childhood infections in patients with congenital heart disease (CHD) are a risk factor for morbidity and death. T-cell Receptor Excision Circles (TRECs) are an established biomarker of T-cell lymphopoiesis and low TREC levels are a marker for Severe Combined Immunodeficiency (SCID). False

J Biomol Struct Dyn, 1996

Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joi... more Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joined by a nucleotide loop (FTFOs) are studied for their binding affinity and specificity to the DNA and RNA single-stranded targets. Thermal denaturation studies reveal that FTFOs have high binding affinity for their targets than do antisense (duplex forming) and antigene (triplex forming) oligonucleotides, because of involvement of both the Watson-Crick

Journal of Clinical Immunology

Journal of Biomolecular Structure & Dynamics, 1995

Linear oligonucleotides that bind to single-stranded nucleic acid targets by formation of both Wa... more Linear oligonucleotides that bind to single-stranded nucleic acid targets by formation of both Watson-Crick (duplex) and Hoogsteen (triplex) hydrogen bonds simultaneously (foldback triplex-forming oligonucleotides; FTFOs) were studied for their ability to disrupt duplex DNA. Recently, we reported that FTFOs interfere with quadruplex forming ability of guanine rich RNA and DNA sequences and indicated that they might also disrupt duplex structures binding to the purine target strand by foldback triplex formation (Kandimalla and Agrawal, Nucleic Acids Res. (1995) 23, 1068–1074). We now obtained evidence for strand displacement of duplex DNA by FTFOs using nuclease assays and thermal melting studies. UV melting studies revealed that complementary strands of 16 to 31 bases long were completely displaced. Results of DNase I assays showed that the FTFOs bound to purine site by strand displacement probably by preassociating with the duplex DNA in the major groove via Hoogsteen hydrogen bonding and subsequently displacing the complementary strand. Experiments with SI nuclease, an enzyme specific for single-stranded nucleic acids, confirmed the strand displacement ability of the FTFOs.

Journal of Drug Targeting, 1998

Pharmacokinetic studies of phosphorothioate oligodeoxynucleotides (PS-oligonucleotides) in animal... more Pharmacokinetic studies of phosphorothioate oligodeoxynucleotides (PS-oligonucleotides) in animals show that following intravenous administration, PS-oligonucleotide clears out rapidly from the plasma and is distributed to majority of the organs. PS-oligonucleotides are bound to plasma proteins extensively. This study was aimed to determine the effect of aspirin, a commonly used drug, on pharmacokinetics of PS-oligonucleotides. In the present study, PS-oligonucleotide was administered to rats that had received aspirin by gavage. Pharmacokinetic study shows that if PS-oligonucleotide was administered following aspirin administration in rats, a) plasma pharmacokinetic parameters (t1/2alpha?, t1/2beta, AUC, etc.) had lower values, b) tissue disposition was different, and c) rate and route of elimination was affected in animals compared to rats receiving PS-oligonucleotide alone. This finding suggests that pharmacokinetics of PS-oligonucleotides can be affected with certain class of drugs, which may have direct impact on biological activity and safety.

We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and... more We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and 3′-5′-deoxyribonucleotide segments. Thermal melting studies of the phosphodiester MBOs (three 2′-5′ linkages at each end) with the complementary 3′-5′-DNA and -RNA target strands suggest that 2′-5′-ribonucleoside incorporation into 3′-5′-oligodeoxyribonucleotides reduces binding to the target strands compared with an all 3′-5′-oligodeoxyribonucleotide of the same sequence and length. Increasing the number of 2′-5′ linkages (from six to nine) further reduces binding to the DNA target strand more than the RNA target strand [Kandimalla,E.R. and Agrawal,S. (1996) Nucleic Acids Symp. Ser., 35, 125-126]. Phosphorothioate (PS) analogs of MBOs destabilize the duplex with the DNA target strand more than the duplex with the RNA target strand. Circular dichroism studies indicate that the duplexes of MBOs with the DNA and RNA target strands have spectral characteristics of both A-and B-type conformations. Compared with the control oligonucleotide, MBOs exhibit moderately higher stability against snake venom phosphodiesterase, S1 nuclease and in fetal calf serum. Although 2′-5′ modification does not evoke RNase H activity, this modification does not effect the RNase H activation property of the 3′-5′-deoxyribonucleotide segment adjacent to the modification. In vitro studies with MBOs suggest that they have lesser effects on cell proliferation, clotting prolongation and hemolytic complement lysis than do control PS oligodeoxyribonucleotides. PS analogs of MBOs show HIV-1 inhibition comparable with that of a control PS oligodeoxyribonucleotide with all 3′-5′ linkages. The current results suggest that a limited number of 2′-5′ linkages could be used in conjunction with PS oligonucleotides to further modulate the properties of antisense oligonucleotides as therapeutic agents.

Journal of Biomolecular Structure & Dynamics, 1996

Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joi... more Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joined by a nucleotide loop (FTFOs) are studied for their binding affinity and specificity to the DNA and RNA single-stranded targets. Thermal denaturation studies reveal that FTFOs have high binding affinity for their targets than do antisense (duplex forming) and antigene (triplex forming) oligonucleotides, because of involvement of both the Watson-Crick and Hoogsteen domains in the interaction. Studies with FTFOs containing different sizes and sequences of loops show that 4–6 bases long loops are optimum for binding; loop sequence does not have a dramatic effect on binding. The FTFOs have greater sequence specificity than do antisense and antigene oligonucleotides because they read the target sequence twice. S1-, PI- and mung bean nuclease protection assays show that the DNA FTFO forms a stable triplex with the DNA target strand, but a weak or no triplex with the RNA target strand. Gel mobility shift assay is used to determine binding of FTFOs to DNA and RNA targets. The circular dichroism (CD) spectrum of the foldback triplex formed with the DNA target strand resembles the B-DNA spectrum, suggesting that the triplex has a B- type of conformation.

Nucleic Acids Research, 1995

Short oligonucleotides that can bind to adjacent sites on target mRNA sequences are designed and ... more Short oligonucleotides that can bind to adjacent sites on target mRNA sequences are designed and evaluated for their binding affinity and biological activity. Sequence-specific binding of short tandem oligonucleotides is compared with a full-length single oligonucleotide (21 mer) that binds to the same target sequence. Two short oligonucleotides that bind without a base separation between their binding sites on the target bind cooperatively, while oligonucleotides that have a one or two base separation between the binding oligonucleotides do not. The binding affinity of the tandem oligonucleotides is improved by extending the ends of the two oligonucleotides with complementary sequences. These extended sequences form a duplex stem when both oligonucleotides bind to the target, resulting in a stable temary complex. RNase H studies reveal that the cooperative oligonucleotides bind to the target RNA with sequence specificity. A short oligonucleotide (9mer) with one or two mismatches does not bind at the intended site, while longer oligonucleotides (21 mers) with one or two mismatches still bind to the same site, as does a perfectly matched 21 mer, and evoke RNase H activity. HIV-1 Inhibition studies reveal an increase in activity of the cooperative oligonucleotide combinations as the length of the dimerization domain increases.

International Journal of Neonatal Screening, Nov 28, 2022

This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY

Biochemical Pharmacology, 1997

Oligodeoxynucleotide phosphorothioates (PS-oligos) are being studied as novel therapeutic agents ... more Oligodeoxynucleotide phosphorothioates (PS-oligos) are being studied as novel therapeutic agents based on their ability to inhibit gene expression. Preclinical studies produced unanticipated complement and coagulation effects in monkeys receiving high-dose PS-oligo. In the present in viao studies, PS-oligo inhibited normal human blood clotting as well as subsequent assays for prothrombin fragment PF,,, and hemolytic complement. PS-oligo treatment of normal donor plasma produced concentration-dependent prolongations of clotting times, with the activated partial thromboplastin time more sensitive than prothrombin time or thrombin clotting time. PS-oligo treatment of normal donor serum similarly reduced hemolytic complement activity in a concentration-dependent manner. Reduced hemolysis correlated with increased levels of complement fragment C4d. The anti-heparin drug protamine sulfate inhibited in vitro effects of PS-oligo in both complement and coagulation assays, suggesting that charged residues in intemucleotide linkages of PS-oligo mediated the observed activities. Therefore, oligonucleotides with varying intemucleotide linkages, nucleotide sequence, or secondary structure were compared. Both complement and coagulation effects appeared to be independent of nucleotide sequence but were strongly related to the nature of internucleotide linkages. Several of these modified oligonucleotides have been shown previously to retain potent antisense activity and thus may represent viable alternatives for antisense therapeutics.

We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and... more We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and 3′-5′-deoxyribonucleotide segments. Thermal melting studies of the phosphodiester MBOs (three 2′-5′ linkages at each end) with the complementary 3′-5′-DNA and-RNA target strands suggest that 2′-5′-ribonucleoside incorporation into 3′-5′-oligodeoxyribonucleotides reduces binding to the target strands compared with an all 3′-5′-oligodeoxyribonucleotide of the same sequence and length. Increasing the number of 2′-5′ linkages (from six to nine) further reduces binding to the DNA target strand more than the RNA target strand [Kandimalla,E.R. and Agrawal,S. (1996) Nucleic Acids Symp. Ser., 35, 125-126]. Phosphorothioate (PS) analogs of MBOs destabilize the duplex with the DNA target strand more than the duplex with the RNA target strand. Circular dichroism studies indicate that the duplexes of MBOs with the DNA and RNA target strands have spectral characteristics of both A-and B-type conformations. Compared with the control oligonucleotide, MBOs exhibit moderately higher stability against snake venom phosphodiesterase, S1 nuclease and in fetal calf serum. Although 2′-5′ modification does not evoke RNase H activity, this modification does not effect the RNase H activation property of the 3′-5′-deoxyribonucleotide segment adjacent to the modification. In vitro studies with MBOs suggest that they have lesser effects on cell proliferation, clotting prolongation and hemolytic complement lysis than do control PS oligodeoxyribonucleotides. PS analogs of MBOs show HIV-1 inhibition comparable with that of a control PS oligodeoxyribonucleotide with all 3′-5′ linkages. The current results suggest that a limited number of 2′-5′ linkages could be used in conjunction with PS oligonucleotides to further modulate the properties of antisense oligonucleotides as therapeutic agents.

Background: Methylotrophs are a diverse group of bacteria that can utilize single-carbon compound... more Background: Methylotrophs are a diverse group of bacteria that can utilize single-carbon compounds as a sole energy source, and are often catalase-positive. Known as environmental symbionts, they are emerging as disease-causing organisms in patients with CGD. Methods: We present a case of lymphadenitis due to Granulibacter bethesdensis, a facultative methylotroph, and review 8 other infections caused by methylotrophs in patients with CGD. Results: There have been 9 reported cases of infections due to methylotrophs in patients with CGD. Seven cases were due to G. bethesdensis, one was due to Acidomonas methanolica and one was due to a Methylobacter. In all cases, 16s rRNA gene sequencing was required for diagnosis. Conclusions: Methylotrophs are fastidious and difficult to identify. Although the mechanisms underlying susceptibility to infection with methylotrophs in CGD remain to be elucidated, these bacteria should be included in the spectrum of pathogens associated with infections ...

Nucleic Acids Research, 1995

Foldback triplex-forming oligonucleotides (FTFOs) that contain an abasic linker, [2-(4-aminobutyr... more Foldback triplex-forming oligonucleotides (FTFOs) that contain an abasic linker, [2-(4-aminobutyr-1-yl)-1,3-propanediol] (APD linker), in the Hoogsteen domain against pyrimidine bases of a C:G and a T:A base pair were studied for their relative stability and sequence specificity of triplex formation. In general, the APD linker has less destabilizing effect against a C:G base pair than a T:A base pair. Incorporation of three APD linker moieties resulted in decreased

Genetics in Medicine, 2012

Genetics in Medicine, 2011

To achieve clinical validation of cutoff values for newborn screening by tandem mass spectrometry... more To achieve clinical validation of cutoff values for newborn screening by tandem mass spectrometry through a worldwide collaborative effort. Methods: Cumulative percentiles of amino acids and acylcarnitines in dried blood spots of approximately 25-30 million normal newborns and 10,742 deidentified true positive cases are compared to assign clinical significance, which is achieved when the median of a disorder range is, and usually markedly outside, either the 99th or the 1st percentile of the normal population. The cutoff target ranges of analytes and ratios are then defined as the interval between selected percentiles of the two populations. When overlaps occur, adjustments are made to maximize sensitivity and specificity taking all available factors into consideration. Results: As of December 1, 2010, 130 sites in 45 countries have uploaded a total of 25,114 percentile data points, 565,232 analyte results of true positive cases with 64 conditions, and 5,341 cutoff values. The average rate of submission of true positive cases between December 1, 2008, and December 1, 2010, was 5.1 cases/day. This cumulative evidence generated 91 high and 23 low cutoff target ranges. The overall proportion of cutoff values within the respective See Acknowledgements for author affiliation.

JAMA, 2014

IMPORTANCE Newborn screening for severe combined immunodeficiency (SCID) using assays to detect T... more IMPORTANCE Newborn screening for severe combined immunodeficiency (SCID) using assays to detect T-cell receptor excision circles (TRECs) began in Wisconsin in 2008, and SCID was added to the national recommended uniform panel for newborn screened disorders in 2010. Currently 23 states, the District of Columbia, and the Navajo Nation conduct population-wide newborn screening for SCID. The incidence of SCID is estimated at 1 in 100,000 births. OBJECTIVES To present data from a spectrum of SCID newborn screening programs, establish population-based incidence for SCID and other conditions with T-cell lymphopenia, and document early institution of effective treatments. DESIGN Epidemiological and retrospective observational study. SETTING Representatives in states conducting SCID newborn screening were invited to submit their SCID screening algorithms, test performance data, and deidentified clinical and laboratory information regarding infants screened and cases with nonnormal results. I...

International Journal of Neonatal Screening

Newborn screening (NBS) laboratories cannot accurately compare mass spectrometry-derived results ... more Newborn screening (NBS) laboratories cannot accurately compare mass spectrometry-derived results and cutoff values due to differences in testing methodologies. The objective of this study was to assess harmonization of laboratory proficiency test (PT) results using quality control (QC) data. Newborn Screening Quality Assurance Program (NSQAP) QC and PT data reported from 302 laboratories in 2019 were used to compare results among laboratories. QC materials were provided as dried blood spot cards which included a base pool and the base pool enriched with specific concentrations of metabolites in a linear range. QC data reported by laboratories were regressed on QC data reported by the Centers for Disease Control and Prevention (CDC), and laboratory’s regression parameters were used to harmonize their PT result. In general, harmonization tended to reduce overall variation in PT data across laboratories. The metabolites glutarylcarnitine (C5DC), tyrosine, and phenylalanine were display...

The Journal of Pediatrics

OBJECTIVES To determine if children with congenital heart disease (CHD) have lower newborn T-cell... more OBJECTIVES To determine if children with congenital heart disease (CHD) have lower newborn T-cell receptor excision circles (TREC) levels than the general population and to evaluate if low TREC levels in newborns with CHD are associated with clinical complications such as hospitalization for infection. STUDY DESIGN The Connecticut Newborn Screening Program reported TREC levels for newborns with CHD delivered between October 2011 and September 2016 at 2 major Connecticut children's hospitals. TREC levels for children with CHD were compared with the general population. TREC levels and outcome measures, including hospitalization for infection, were compared. RESULTS We enrolled 575 participants with CHD in the study. The median TREC level for newborns with CHD was lower than the general population (180.1 copies/μL vs 312.5 copies/μL; P < .01). patients with CHD requiring hospitalization for infection had lower median TREC levels than their counterparts (143.0 copies/μL vs 186.7 copies/μL; P < .01). The combination of prematurity and low TREC level had a strong relationship to hospitalization for infection (area under the receiver operative characteristic curve of 0.89). There was no association between TREC level and CHD severity. CONCLUSIONS Newborns with CHD demonstrated lower TREC levels than the general population. Low TREC levels were associated with hospitalization for infection in preterm children with CHD. Study limitations include that this was a retrospective chart review. These findings may help to identify newborns with CHD at highest risk for infection, allowing for potential opportunities for intervention.

Journal of the American College of Cardiology

Routine childhood infections in patients with congenital heart disease (CHD) are a risk factor fo... more Routine childhood infections in patients with congenital heart disease (CHD) are a risk factor for morbidity and death. T-cell Receptor Excision Circles (TRECs) are an established biomarker of T-cell lymphopoiesis and low TREC levels are a marker for Severe Combined Immunodeficiency (SCID). False

J Biomol Struct Dyn, 1996

Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joi... more Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joined by a nucleotide loop (FTFOs) are studied for their binding affinity and specificity to the DNA and RNA single-stranded targets. Thermal denaturation studies reveal that FTFOs have high binding affinity for their targets than do antisense (duplex forming) and antigene (triplex forming) oligonucleotides, because of involvement of both the Watson-Crick

Journal of Clinical Immunology

Journal of Biomolecular Structure & Dynamics, 1995

Linear oligonucleotides that bind to single-stranded nucleic acid targets by formation of both Wa... more Linear oligonucleotides that bind to single-stranded nucleic acid targets by formation of both Watson-Crick (duplex) and Hoogsteen (triplex) hydrogen bonds simultaneously (foldback triplex-forming oligonucleotides; FTFOs) were studied for their ability to disrupt duplex DNA. Recently, we reported that FTFOs interfere with quadruplex forming ability of guanine rich RNA and DNA sequences and indicated that they might also disrupt duplex structures binding to the purine target strand by foldback triplex formation (Kandimalla and Agrawal, Nucleic Acids Res. (1995) 23, 1068–1074). We now obtained evidence for strand displacement of duplex DNA by FTFOs using nuclease assays and thermal melting studies. UV melting studies revealed that complementary strands of 16 to 31 bases long were completely displaced. Results of DNase I assays showed that the FTFOs bound to purine site by strand displacement probably by preassociating with the duplex DNA in the major groove via Hoogsteen hydrogen bonding and subsequently displacing the complementary strand. Experiments with SI nuclease, an enzyme specific for single-stranded nucleic acids, confirmed the strand displacement ability of the FTFOs.

Journal of Drug Targeting, 1998

Pharmacokinetic studies of phosphorothioate oligodeoxynucleotides (PS-oligonucleotides) in animal... more Pharmacokinetic studies of phosphorothioate oligodeoxynucleotides (PS-oligonucleotides) in animals show that following intravenous administration, PS-oligonucleotide clears out rapidly from the plasma and is distributed to majority of the organs. PS-oligonucleotides are bound to plasma proteins extensively. This study was aimed to determine the effect of aspirin, a commonly used drug, on pharmacokinetics of PS-oligonucleotides. In the present study, PS-oligonucleotide was administered to rats that had received aspirin by gavage. Pharmacokinetic study shows that if PS-oligonucleotide was administered following aspirin administration in rats, a) plasma pharmacokinetic parameters (t1/2alpha?, t1/2beta, AUC, etc.) had lower values, b) tissue disposition was different, and c) rate and route of elimination was affected in animals compared to rats receiving PS-oligonucleotide alone. This finding suggests that pharmacokinetics of PS-oligonucleotides can be affected with certain class of drugs, which may have direct impact on biological activity and safety.

We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and... more We have designed and synthesized mixed backbone oligonucleotides (MBOs) containing 2′-5′-ribo-and 3′-5′-deoxyribonucleotide segments. Thermal melting studies of the phosphodiester MBOs (three 2′-5′ linkages at each end) with the complementary 3′-5′-DNA and -RNA target strands suggest that 2′-5′-ribonucleoside incorporation into 3′-5′-oligodeoxyribonucleotides reduces binding to the target strands compared with an all 3′-5′-oligodeoxyribonucleotide of the same sequence and length. Increasing the number of 2′-5′ linkages (from six to nine) further reduces binding to the DNA target strand more than the RNA target strand [Kandimalla,E.R. and Agrawal,S. (1996) Nucleic Acids Symp. Ser., 35, 125-126]. Phosphorothioate (PS) analogs of MBOs destabilize the duplex with the DNA target strand more than the duplex with the RNA target strand. Circular dichroism studies indicate that the duplexes of MBOs with the DNA and RNA target strands have spectral characteristics of both A-and B-type conformations. Compared with the control oligonucleotide, MBOs exhibit moderately higher stability against snake venom phosphodiesterase, S1 nuclease and in fetal calf serum. Although 2′-5′ modification does not evoke RNase H activity, this modification does not effect the RNase H activation property of the 3′-5′-deoxyribonucleotide segment adjacent to the modification. In vitro studies with MBOs suggest that they have lesser effects on cell proliferation, clotting prolongation and hemolytic complement lysis than do control PS oligodeoxyribonucleotides. PS analogs of MBOs show HIV-1 inhibition comparable with that of a control PS oligodeoxyribonucleotide with all 3′-5′ linkages. The current results suggest that a limited number of 2′-5′ linkages could be used in conjunction with PS oligonucleotides to further modulate the properties of antisense oligonucleotides as therapeutic agents.

Journal of Biomolecular Structure & Dynamics, 1996

Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joi... more Oligodeoxyribonucleotides containing both Watson-Crick and Hoogsteen hydrogen bonding domains joined by a nucleotide loop (FTFOs) are studied for their binding affinity and specificity to the DNA and RNA single-stranded targets. Thermal denaturation studies reveal that FTFOs have high binding affinity for their targets than do antisense (duplex forming) and antigene (triplex forming) oligonucleotides, because of involvement of both the Watson-Crick and Hoogsteen domains in the interaction. Studies with FTFOs containing different sizes and sequences of loops show that 4–6 bases long loops are optimum for binding; loop sequence does not have a dramatic effect on binding. The FTFOs have greater sequence specificity than do antisense and antigene oligonucleotides because they read the target sequence twice. S1-, PI- and mung bean nuclease protection assays show that the DNA FTFO forms a stable triplex with the DNA target strand, but a weak or no triplex with the RNA target strand. Gel mobility shift assay is used to determine binding of FTFOs to DNA and RNA targets. The circular dichroism (CD) spectrum of the foldback triplex formed with the DNA target strand resembles the B-DNA spectrum, suggesting that the triplex has a B- type of conformation.

Nucleic Acids Research, 1995

Short oligonucleotides that can bind to adjacent sites on target mRNA sequences are designed and ... more Short oligonucleotides that can bind to adjacent sites on target mRNA sequences are designed and evaluated for their binding affinity and biological activity. Sequence-specific binding of short tandem oligonucleotides is compared with a full-length single oligonucleotide (21 mer) that binds to the same target sequence. Two short oligonucleotides that bind without a base separation between their binding sites on the target bind cooperatively, while oligonucleotides that have a one or two base separation between the binding oligonucleotides do not. The binding affinity of the tandem oligonucleotides is improved by extending the ends of the two oligonucleotides with complementary sequences. These extended sequences form a duplex stem when both oligonucleotides bind to the target, resulting in a stable temary complex. RNase H studies reveal that the cooperative oligonucleotides bind to the target RNA with sequence specificity. A short oligonucleotide (9mer) with one or two mismatches does not bind at the intended site, while longer oligonucleotides (21 mers) with one or two mismatches still bind to the same site, as does a perfectly matched 21 mer, and evoke RNase H activity. HIV-1 Inhibition studies reveal an increase in activity of the cooperative oligonucleotide combinations as the length of the dimerization domain increases.