Biomedical subjects

C Richert

Publications and source records attributed to C Richert.

22 records · Page 2Linked to original sources

PubMed1994

Photodynamic antitumor agents: beta-methoxyethyl groups give access to functionalized porphycenes and enhance cellular uptake and activity.

Porphycene photosensitizers bearing two or four methoxyethyl side chains were synthesized in nine steps from commercially available starting materials. Ether cleavage led to (hydroxyethyl)- and (bromoethyl)porphycenes that were converted to vinyl and benzo derivatives. Five of the side chain-functionalized porphycenes were biologically studied in comparison with two tetra-n-propylporphycenes. Porphycenes were incorporated in small unilamellar liposomes and incubated with cultivated SSK2 murine fibrosarcoma cells. Cellular uptake and phototoxicity 24 h after 5 J/cm2 laser light treatment were determined. The porphycenes tested were between 17 and 220 times more photodynamically active than the currently clinically used sensitizer Photofrin, although extinction coefficients of the porphycenes' irradiated bands are only approximately 10-fold higher. The LD50 concentration for SSK2 cells in the incubation medium was as low as (8.5 +/- 2.8) x 10(-9) M for tetrakis(methoxyethyl)porphycene. Two methoxy or hydroxy groups enhanced cellular uptake, three or four methoxy groups both enhanced and accelerated cellular uptake of tetraalkylporphycenes. Half-life times of the uptake processes varied between (0.14 +/- 0.04) and (14 +/- 4) h and cellular saturation levels between (1.2 +/- 0.2) and (26 +/- 3) pmol/10(5) cells. When individual uptake rates were accounted for, all porphycenes had a similar "cellular" phototoxicity, pointing toward a common mechanism of action. Evidence is presented for the assumption that cell membranes are the primary targets of the tested porphycenes and that membrane solubility may play a critical role in their photodynamic efficiency. The results show that nonionic polar side chain functionalities can strongly enhance cellular uptake and antitumor activity of lipophilic porphyrinoids and thus that the known lipophilicity/activity relationship can be reversed for very hydrophobic sensitizers.

Animals↗

PubMed1993

Tumour localisation kinetics of photofrin and three synthetic porphyrinoids in an amelanotic melanoma of the hamster.

In this study the localisation of porphyrinoid photosensitizers in tumours was investigated. To determine if tumour selectivity results from a preferential uptake or prolonged retention of photosensitizers, intravital fluorescence microscopy and chemical extraction were used. Amelanotic melanoma (A-Mel-3) were implanted in a skin fold chamber in Syrian Golden hamsters. Distribution of the porphyrin mixture Photofrin and three porphycenes, pure porphyrinoid model compounds, was studied quantitatively by intravital fluorescence microscopy. Extraction of tissue and blood samples was performed to verify and supplement intravital microscopic results. Photofrin accumulated in melanomas reaching a maximum tumour:skin tissue ratio of 1.7:1. Localisation of the different porphycenes was found to be highly tumour selective (3.2:1), anti-tumour selective (0.2:1), and non-selective (1:1) with increasing polarity of the porphycenes. The two non-tumour selective porphycenes had distinctly accelerated serum and tissue kinetics; serum halflife times being as short as 1 min. The specific localisation of the slowly distributed, tumour selective photosensitizers, occurred exclusively during the distribution from serum and uptake into tissues. For the most selective porphycene, the tumour selection process had a halflife of 260 +/- 150 min and led to a strongly fluorescent tumour edge edema. Accumulation of porphyrines by the amelanotic melanoma (A-Mel-3) can be attributed to an enhanced uptake rate for lipophilic molecules in this subcutaneously growing neoplasm. The slow distribution of the two tumour specific photosensitizers and the strong fluorescence of these hydrophobic molecules in the tumour compartment with a high water content indicate a carrier role of serum proteins in the selection process. Enhanced permeability of the tumour vasculature to macromolecules appears to be the most probable reason for the tumour selectivity of these two sensitisers.

Animals↗

PubMedDate not supplied

Monitored selection of DNA-hybrids forming duplexes with capped terminal C:G base pairs.

Reported here are the results of a search for modified oligodeoxynucleotides with a 5'-terminal cytidine residue whose affinity for target strands is enhanced by 5'-acylamido groups. These acylamido groups were envisioned to act as molecular caps that bind to the exposed terminal base pair of the duplex with the target strand. A total of 52 capped oligonucleotides of the sequence R-CGGTTGAC, where R denotes the 5'-appendage and C a 5'-amino-2',5'-dideoxycytidine residue, were tested. Among the building blocks employed to modify the 5'-amino group of the DNA strand were carboxylic acid residues, either appended directly or via an amino acid residue, and aromatic aldehydes, coupled via reductive amination. The carboxylic acids employed ranged from Fmoc-glycine to (Fmoc)(2)-vancomycin and included a number of aromatic acids and bile acids. Small libraries were subjected to MALDI-monitored nuclease selection experiments, and selected compounds were tested in UV-melting assays with target strands. Cholic acid appendages stabilized terminal C:G base pairs to the greatest extent, with melting point increases of up to 10 degrees C. Further, the cholic acid residue enhanced base pairing fidelity at the terminus, as determined in melting analyses with target strands containing a mismatched nucleobase at the 3'-terminus.

Base Pairing↗

PubMedDate not supplied

Selection of modified oligonucleotides with increased target affinity via MALDI-monitored nuclease survival assays.

Reported here is how modified oligonucleotides with increased affinity for DNA or RNA target strands can be selected from small combinatorial libraries via spectrometrically monitored selection experiments (SMOSE). The extent to which target strands retard the degradation of 5'-acyl-, 5'-aminoacyl-, and 5'-dipeptidyl-oligodeoxyribonucleotides by phosphodiesterase I (EC 3.1.4.1) was measured via quantitative MALDI-TOF mass spectrometry. Oligonucleotide hybrids were prepared on solid support, and nuclease selections were performed with up to 10 modified oligonucleotides in one solution. The mass spectrometrically monitored experiments required between 120 and 300 pmol of each modified oligonucleotide, depending on whether HPLC-purified or crude compounds were employed. Data acquisition and analysis were optimized to proceed in semiautomated fashion, and functions correcting for incomplete degradation during the monitoring time were developed. Integration of the degradation kinetics provided "protection factors" that correlate well with melting points obtained with traditional UV melting curves employing single, pure compounds. Among the components of the five libraries tested, three were found to contain 5'-substituents that strongly stabilize Watson--Crick duplexes. Selecting and optimizing modified oligonucleotides via monitored nuclease assays may offer a more efficient way to search for new antisense agents, hybridization probes, and biochemical tools.

Automation↗