Biomedical subjects

M Caruthers

Publications and source records attributed to M Caruthers.

4 recordsLinked to original sources

PubMed1997

Comparative pharmacokinetics, tissue distribution, and tumor accumulation of phosphorothioate, phosphorodithioate, and methylphosphonate oligonucleotides in nude mice.

The goals of this study were to systematically compare the pharmacokinetics and tissue distribution of phosphorothioate (PS), methylphosphonate (MP), and phosphorodithioate (PS2) oligonucleotide analogs; 15-mers of sequence d-TAC GCC AAC AGC TCC (5'-3') complementary to the AUG region of K-ras were radiolabeled with carbon-14. Oligomers were administered as a single dose in the tail vein of nude mice harboring a K-ras-dependent human pancreatic tumor (CFPAC1). The kinetics of PS, PS2, and MP oligomer availability in the bloodstream was followed. Concentration versus time profiles for all oligomers were biphasic, indicative of a two-compartment model. A rapid distribution phase with t1/2 alpha values of 1 minute or less and an elimination phase with average t1/2 beta values of 24-35 minutes were observed. Volumes of distribution (Vd) were 3.2, 4.8, and 6.3 ml for PS2, MP, and PS, respectively, in comparison to 3.6 ml for sucrose, a fluid-phase marker. Relative tissue drug levels obtained at 1 and 24 hours after administration were kidney > liver > spleen > tumor > muscle. Total kidney and liver oligonucleotide accumulation was approximately 7%-15% of the initial dose, with tumor accumulating 2%-3%. Intact compound was recovered from all tissues, including tumor, as assessed by high-pressure reversed-phase HPLC coupled to radiometric detection. Integrity of the oligonucleotides ranged from 73% in blood to 43%-46% in kidney and liver. Kidney and liver appear to be the primary sites of metabolism. These results demonstrate widespread tissue availability of these compounds and suggest their development as potential antitumor agents.

Animals↗

PubMed1982

Synthesis of the nutL DNA segments and analysis of antitermination and termination functions in coliphage lambda.

The nut antiterminator sequence, when present between a promoter and a terminator, permits the N-mediated antitermination of transcription in phage lambda. The efficiency of nutL was determined by assaying the activity of gene galK placed on a plasmid downstream from the promoter, nutL and terminator modules. As a reference and an estimate of the plasmid copy number, we have used an improved and very reproducible assay for bla activity. Sequences consisting of the 17-bp nutL core flanked by two HindIII cohesive sites were synthesized by the phosphite coupling method, and cloned in proper orientation between the Pp promoter of pBR322 and lambda gene N followed by the tL1 terminator on a galK-expression plasmid. The antitermination efficiencies for two synthetic 17-bp nutL sequences, one wild type and one point mutant at the base of the nutL stem, are similar but substantially reduced in comparison with the native 25-bp nutL sequence cloned at the same site in the otherwise identical galK-expression plasmid. Multiple tandem insertions of the synthetic 17-bp nutL segment successively increase antitermination efficiency, but also to levels below those of comparable plasmids carrying multiple copies of the native 25-bp nutL sequence. Thus, several specific base pairs in the flanking sequences appear to be important for the efficient nut function. In an inverted orientation the 17-bp nutL sequence has lost its antitermination function. It also lost the termination activity exhibited by inversion of the longer 25-bp and 74-bp native nutL sequences.

Bacteriophage lambda↗

PubMed1982

lac repressor-lac operator interaction: NMR observations.

We show here the changes in the NMR spectra of the Escherichia coli lac repressor when bound to isolated lac operator DNA. The observations focus on the aromatic residues--four tyrosines and a single histidine--in the amino-terminal DNA binding domain of the lac repressor. There is a good correlation between chemical shift changes seen by 19F NMR when compared with 1 H NMR of otherwise identical repressor--DNA complexes. The results suggest that the tyrosines do not intercalate in the DNA. The NMR spectral changes with similarly sized DNA fragments, not containing the lac operator DNA sequence, are different. Thus, the amino-terminal domain of the lac repressor is independently capable of discriminating between lac operator and nonspecific DNA sequences. There can be two amino-terminal fragments per operator in the specific complex.

Amino Acid Sequence↗

PubMedDate not supplied

A microchemical facility for the analysis and synthesis of genes and proteins.

A series of automated instruments that use state-of-the-art chemical methods has been developed for high-sensitivity protein sequencing, DNA synthesis and peptide synthesis. These instruments have been integrated into a centralized microchemical facility in order to promote their use for the study of a variety of biologically interesting problems. This facility has as one of its major functions the development of new chemistries and instrumentation for the structural analysis and synthesis of genes and proteins.

Amino Acid Sequence↗