title: A flow cytometry method to quantitate internalized immunotoxins shows that taxol synergistically increases cellular immunotoxins uptake.
abstract: Tumor microenvironments present significant barriers to penetration by antibodies, immunoconjugates, and other immunotoxins. In this report, we illustrate a novel strategy to increase tumor cell uptake of immunotoxin by combination with Taxol. SS1P is an immunotoxin composed of the Fv portion of a mesothelin-specific antibody fused to a bacterial toxin that is presently undergoing phase II testing in mesothelioma. Using novel flow cytometry and gel filtration methods, we quantified SS1P uptake in individual tumor cells along with levels of shed mesothelin (sMSLN), a barrier of SS1P therapy. The validity of our flow cytometric method was confirmed by the ability to similarly quantitate tumor cell uptake of Herceptin and an immunotoxin targeting HER2/neu. SS1P uptake peaked several hours after SS1P was cleared from the blood, reflecting an intratumor distribution process of SS1P that is independent of blood supply. Using the methods developed, we demonstrated that Taxol could improve SS1P penetration into tumors in parallel with an associated reduction of sMSLN in tumor extracellular fluid. Our findings offer a mechanistic rationale to combine SS1P with Taxol or another cytotoxic drug as a strategy to increase immunotoxin uptake by tumor cells. Further, we suggest one basis to understand why chemotherapy and antibody-based therapies cooperate when combined in cancer treatment.
authors.affiliation: Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland 20892-4264, USA.
authors.forename: Yujian
authors.initials: Y
authors.lastname: Zhang
authors.forename: Johanna K
authors.initials: JK
authors.lastname: Hansen
authors.forename: Laiman
authors.initials: L
authors.lastname: Xiang
authors.forename: Seiji
authors.initials: S
authors.lastname: Kawa
authors.forename: Masanori
authors.initials: M
authors.lastname: Onda
authors.forename: Mitchell
authors.initials: M
authors.lastname: Ho
authors.forename: Raffit
authors.initials: R
authors.lastname: Hassan
authors.forename: Ira
authors.initials: I
authors.lastname: Pastan
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism / Q000493:pharmacokinetics*; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D045744:Cell Line, Tumor; D004705:Endocytosis / Q000187:drug effects; D005434:Flow Cytometry / Q000379:methods*; D058851:GPI-Linked Proteins; D006801:Humans; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D000090204:Mesothelin; D051379:Mice; D041681:NIH 3T3 Cells; D009369:Neoplasms / Q000378:metabolism / Q000473:pathology; D017239:Paclitaxel / Q000494:pharmacology*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D000972:Antineoplastic Agents, Phytogenic; D058851:GPI-Linked Proteins; D008562:Membrane Glycoproteins; D000090204:Mesothelin; D017239:Paclitaxel; D018719:Erb-b2 Receptor Tyrosine Kinases; C000715528:Msln protein, mouse; C474515:SS1(dsFv)PE38