Almost ever since their invention, monoclonal antibodies have held the promise of cancer-specific drug targeting--Paul Ehrlich's "magic bullet"--but only during the past decade have a modest number of anti-cancer antibodies received approval for clinical use. These, however, have proven largely successful, with very different kinds of conventional or recombinant, murine, humanized, recombinant fully human and fusion constructs, and mechanisms of action as diverse as complement or antibody dependent cytotoxicity, anti-angiogenesis, and growth factor inhibition. In these latter two mechanisms of action, antibodies compete with novel small-molecule drugs. This review tries to elucidate current trends in those antibody-based therapeutics that are currently in clinical development. With more than 400 such molecules registered for clinical trials, it is far from a chance to be complete. Still, from those antibodies selected for a closer view, two large trends can be distilled: The movement towards increasingly molecularly defined recombinant constructs, and away from classical antibody effector functions in immune activation towards additional mechanisms of action--either by stimulation or (more often) inhibition of a molecular target function, or by additional functional moieties attached to the antibody scaffold. While these trends probably mark the future of antibody development for cancer therapy and clinical applications in general, a considerable number of more conventional--hybridoma generated or recombinantly chimerized or humanized--Fc receptor-activating antibodies, originally generated a decade or longer ago, continue to make their way through clinical trials, some with remarkable success.

The cure of cancer is still a formidable challenge in medical science. Long-known modalities including surgery, chemotherapy and radiotherapy are successful in a number of cases; however, invasive, metastasized and inaccessible tumors still pose an unresolved and ongoing problem. Targeted therapies designed to locate, detect and specifically kill tumor cells have been developed in the past three decades as an alternative to treat troublesome cancers. Most of these therapies are either based on antibody-dependent cellular cytotoxicity, targeted delivery of cytotoxic drugs or tumor site-specific activation of prodrugs. The latter is a two-step procedure. In the first step, a selected enzyme is accumulated in the tumor by guiding the enzyme or its gene to the neoplastic cells. In the second step, a harmless prodrug is applied and specifically converted by this enzyme into a cytotoxic drug only at the tumor site. A number of targeting systems, enzymes and prodrugs were investigated and improved since the concept was first envisioned in 1974. This review presents a concise overview on the history and latest developments in targeted therapies for cancer treatment. We cover the relevant technologies such as antibody-directed enzyme prodrug therapy (ADEPT), gene-directed enzyme prodrug therapy (GDEPT) as well as related therapies such as clostridial- (CDEPT) and polymer-directed enzyme prodrug therapy (PDEPT) with emphasis on prodrug-converting enzymes, prodrugs and drugs.

A recombinant fusion protein of colon carcinoma binding A33 single chain antibody with cytosine deaminase displayed specific antigen binding and enzyme activity in surface plasmon resonance and is catalytic activity assay. In vitro, it selectively increased the toxicity of 5-FC to A33 antigen-positive cells by 300-fold, demonstrating the potency of this ADEPT strategy.

Abstract. Antibody-directed enzyme-prodrug therapy (ADEPT) aims at improving the specificity of conventional chemotherapy by employing artificial antibody-enzyme constructs to convert a non-toxic prodrug into a cytotoxic agent specifically localized to the tumor site. The gpA33 antigen is a promising target for ADEPT in colon cancer, as it is expressed by >95% of human colon cancers, but is absent in all non-gastrointestinal tissues. We designed a recombinant fusion construct of a phage display-generated anti-gpA33 single chain fragment, A33scFv, with cytosine deaminase from yeast (CDy), which converts 5-fluorocytosine (5-FC) into 5-fluorouracil (5-FU). The resulting construct, A33scFv::CDy, was overexpressed in Pichia pastoris and secreted into culture supernatant. The fusion protein was purified by affinity chromatography on protein L. Silver-staining after SDSpolyacrylamide gel electrophoresis confirmed molecular mass and purity. Antibody binding and specificity were quantified by flow cytometry. The complete ADEPT system was applied in vitro on gpA33-positive LIM1215 cells, assessing cell survival by a fluorescein diacetate assay. Cytotoxicity of the prodrug 5-FC after A33scFv::CDy binding was equimolar to that of 5-FU, and this effect depended specifically on both antibody and enzyme function. These results demonstrate bifunctional activity of the heterogeneous Pichia-produced A33scFv::CDy fusion protein and proof of principle for the ADEPT system proposed herein. IntroductionMonoclonal antibodies have become an accepted modality of cancer therapy. Recombinant antibodies and antibody-based fusion proteins hold the promise of further extending the therapeutic possibilities of this modality. Single chain variable fragments (scFv) consist of the variable regions of an antibody's heavy and light chains fused together via a flexible linker, whose length determines the quaternary structure. Thus, they carry the complete antigen binding site in a single polypeptide chain of only about 30 kDa. In tumor targeting, scFv have demonstrated excellent tumor penetration, high ratios of tumor to normal tissue concentration, and low background (1,2). This makes them attractive targeting components of bifunctional fusion proteins such as those needed for antibodydirected enzyme-prodrug therapy (ADEPT). In ADEPT, after binding of an antibody-enzyme construct to the cognate tumor antigen, the enzyme component converts a prodrug into a cytotoxic drug, thus generating drug activity specifically in tumor tissue (3,4).Several ADEPT systems have shown promising in vivo efficacy in a number of tumor models (5) and in several xenograft systems in nude mice (4,6-8), demonstrating in principle that ADEPT can target tumor tissue with high selectivity and deliver chemotherapeutic drugs with high intratumoral concentrations.Senter's group first used bacterial cytosine deaminase for ADEPT to catalyze the deamination of 5-fluorocytosine (5-FC), which is non-toxic in mammals, into 5-fluorouracil (5-FU) (9). Clinical studies on ADEPT have pro...

Invasive pulmonary aspergillosis (IPA) is one of the major complications in immunocompromised patients. The mainstay of diagnostic imaging is non-enhanced chest-computed-tomography (CT), for which various non-specific signs for IPA have been described. However, contrast-enhanced CT pulmonary angiography (CTPA) has shown promising results, as the vessel occlusion sign (VOS) seems to be more sensitive and specific for IPA in hematologic patients. The aim of this study was to evaluate the diagnostic accuracy of CTPA in a larger cohort including non-hematologic immunocompromised patients. CTPA studies of 78 consecutive immunocompromised patients with proven/probable IPA were analyzed. 45 immunocompromised patients without IPA served as a control group. Diagnostic performance of CTPA-detected VOS and of radiological signs that do not require contrast-media were analyzed. Of 12 evaluable radiological signs, five were found to be significantly associated with IPA. The VOS showed the highest diagnostic performance with a sensitivity of 0.94, specificity of 0.71 and a diagnostic odds-ratio of 36.8. Regression analysis revealed the two strongest independent radiological predictors for IPA to be the VOS and the halo sign. The VOS is highly suggestive for IPA in immunocompromised patients in general. Thus, contrast-enhanced CTPA superior over non-contrast_enhanced chest-CT in patients with suspected IPA.