Anja Vervoorts

Taxol Inclusion Complexes with a Cyclodextrin Dimer: Possibilities to Detoxify Chemotherapeutics and to Target Drugs Specifically to Tumors?

The natural drug, paclitaxel (taxol), is highly effectiveas a tumor chemotherapeuticwith a low probability of inducing chemoresistance,but shows severe toxic side effectsat the therapeutic dose. How can this toxicitybe overcome? Here we report the synthesisof cyclodextrin dimers connected at thesecondary face by amide-bonded aliphatic spacers.The spacer length of one of the dimers referred to asdiβCD(2N-A4C5A4) or dimer 7cmatches the distance between the twobenzoic acid residues of paclitaxel. We investigated the physical inclusion of taxol into this dimer using the TNS-label competition method. Affinity constants with the dimer in comparison to free β-cyclodextrin are found to be of the order of 107 l/mole.When included into the cyclodextrin dimer, the drug shows a considerable time delay of incorporation into human tumor cell cultures (OAT SCLC cells) or a total exclusion from the cells. This is the prerequisite to avoid intoxication of other organs of a patient. Possibilities are discussed to detoxify chemotherapeutics and to target their inclusioncomplexes specifically to tumors using specific biological signals.

How many molecules of a photosensitizer are necessary to photosensitize a tumor cell

Diglycosylated porphyrinoids were shown to stack at the outer cell membrane of tumor cells in vitro. Similar phototoxicities are acquired as with the non-glycosylated drugs at amounts of only 1% of that normally taken up of the non-glycosylated drugs. This means that a strict positioning of sensitizers at sensitive compartments may reduce the amount of sensitizer necessary to sensitize a tumor cell to about 106 molecules per cell. Similar numbers were seen only in erythrocytes and with immunoconjugates in ovarian cancer. This low amount facilitates photosensitizer transport against intratumoral pressure gradients.

Carrier systems in PDT: on the way to novel antitumor drugs

A novel mode to apply photosensitizing drugs specifically to tumor tissue using the principles of polyphasic tumor therapies is outlined. Key compounds are tumor specific functionalized antibodies with reduced immunogenicity. These bind to drug inclusion complexes in a multiplicative manner. Drug inclusion complexes are designed on the basis of tethered functionalized (beta) -cyclodextrin dimers with maximum affinity to porphyrinoid photosensitizers forming monomeric chlathrates. To enhance porphyrin-cyclodextrin interaction peripheral groups of the porphyrin have to be chemically modified. The development of the method is not yet completed. First results are demonstrated.

Cyclodextrin Dimers Used To Prevent Side Effects of Photochemotherapy and General Tumor Chemotherapy

Recent developments of polyphasic tumor detection and tumor therapy including applications of dimeric cyclodextrin inclusion complexes of high affinity for Photodynamic Tumor Therapy (PTT) porphyrinoids are discussed in the light of future applications of similar methodology to general tumor chemotherapy. These approaches hopefully will end the martyrium of patients undergoing tumor chemotherapy today.

Infusion technique allowing long-term tumor targeting with antibodies on the hen's egg test (HET) system

The accumulation of antibodies or their fragments in tumor tissues is a basal requirement for immunotargeting of tumors in vivo. Immunotargeting is the first step for a polyphasic tumor therapy and diagnosis. Usually the high interstitial pressure and the insufficient blood supply of the tumor prevents the accumulation of tumor specific antibodies. To overcome these barriers the antibodies can be modified concerning their molecular weight. On the other hand, different injection techniques can influence the antibody distribution in-vivo. We developed a long term infusion technique for the testing of antibodies and antibody fragments in xenotransplanted tumors in the hens egg test system. This in-vivo model allows to develop new protocols for the antibody administration resulting in a tumor localization without targeting of non-tumor tissues. Macroscopic and microscopic findings in the HET-system may reflect the binding sites of antibodies in-situ. Detection of tumor markers has not yet been realized due to problems inherent to the system.

Specific labeling of microtumors in the hen's egg test system by dimethoxyhematoporphyrin

The accumulation of fluorescent dyes and anticancer drugs in tumors is a fundamental requirement for tumor-diagnosis and therapy. We tested various photosensitizers on vascularized microtumors in the hens egg test system according to their distribution and accumulation by in-situ videofluorescence microscopy and their influence on tumor blood flow by laser- Doppler spectroscopy. Only 2,4-di((alpha) -methoxyethyl)- hematoporphyrin showed strict tumor localization while diglucosamidyl-heptyl-chlorin labeled cell membranes exclusively. Results from laser-Doppler spectroscopy did not show any influence on tumor blood flow. Intelligent combining of the properties of these sensitizers with accumulation strategies should show in principle strict tumor localization without targeting of non-tumor tissues.

Laser Doppler flowmetry in photodynamic therapy on xenotransplanted tumors

Perfusion of tumor tissue is a necessary prerequisite for radiotherapy and Photodynamic Tumor Therapy (PDT). For PDT perfusion means oxygen supply to a light activated photosensitizing drug inside tumor cells to destroy these cells specifically by light activated singlet oxygen. These experiments are normally done in rodents, but can be much more easily performed on tumors transplanted to the extraembryonal painfree vessels developed during chick embryogenesis. We chose the yolk sac membrane (YSM) and the chorioallantoic membrane (CAM) for quantitative blood flow measurements. Near infrared light (830 nm) was used for measurements in vascularized xenotransplanted tumors on the extraembryonal membranes because at this wavelength blood flow can be detected also in vessels covered by tumor cells. We measured the influences on blood flow of different photosensitizers with and without therapeutic irradiation and single components of a polyphasic PDT System.

Quantitative data on blood flow during tumor PDT obtained by laser Doppler spectroscopy in the hen's egg test system

Oxygen supply is the most important requirement of type II photodynamic reactions. Prerequisite in photodynamic tumor therapy is an intact tumor blood flow during irradiation. Most photosensitizers destroy tumor vessels due to accumulation in endothelial cells. As a prerequisite to develop novel photosensitizing drugs an in-vivo test system is required to quantitatively assess for inertness of those sensitizers to the blood supply. We adapted and further developed a system capable of measuring the relative oxygen supply to heterotransplanted tumors on the yolk sac membrane (YSM) of fertilized chicken eggs.