Novel Targeting Strategies Using Recombinant Antibodies for Early Diagnosis and Therapy of Ovarian Cancer

Editorial Novel targeting strategies using recombinant antibodies for early diagnosis and therapy of ovarian cancer “…carefully designed screening strategies of recombinant antibody libraries have the potential to lead to the identification of ideal targeting reagents for both in vitro and in vivo uses.” Despite major efforts to identify biomarkers or lipid-based or paramagnetic nanoparticles to biomarker panels performing better than CA125, highly versatile polymers and viral capsids, can which is the most studied ovarian cancer bio- be used to detect and characterize pathological marker, no diagnostic marker for the early detec- abnormalities [10–14]. The extra- or intra cellular Nathalie Scholler tion of ovarian cancer has been approved to date. localization of the macromolecules can be con- Department of Obstetrics Almost 30 years after the discovery of CA125 [1], trolled by coupling them to cell-penetrating & Gynecology, Center for Research on Reproduction & Women’s Health, ovarian cancer continues to be generally diag- peptides and transduction domains [15,16], while Penn Ovarian Cancer Research Center, nosed at an advanced stage where the case:fatality sensitivity and specificity can be increased by University of Pennsylvania, ratio is high, thus remaining the most lethal of strategies such as attachment to proteins and Philadelphia, PA 19104-6160, USA Tel.: +1 215 898 0164 all gynecologic malignancies among US women. particles, encapsulation into micelles and caged Fax: +1 215 573 5129 Attempts at screening with conventional imaging structures, biochemical reporters that are sensi- [email protected] strategies have been hampered by high levels of tive to pH, proteins, temperature or oxygen [17], false-positives with only 13–21% of the women and targeting to receptors. The conjugation of that undergo surgery to remove adnexal masses targeting platforms to nanoparticles optimizes actually having ovarian cancer [2]. As a result, specific local delivery. Any moiety with specific while the case for early detection of ovarian can- binding properties, such as monoclonal anti- cer is undeniable [3], the current opinions regard- bodies [18], or molecules that target cell recep- ing screening for ovarian cancer in the general tors [19–21] can be used as targeting reagents. For population are divided between negative [4–6] example, the folate receptor-a that is highly and mildly positive [7,8] recommendations that expressed by ovarian cancer cells can be targeted emphasize stable markers in healthy controls by folate or by an antifolate receptor-a antibody. over time. Clearly, to become a standard-of-care Yet, since antibodies can specifically bind to a screening, modalities must be improved, but unique antifolate receptor while folate binds doubts about the achievability of the task haunt to any folate receptors, the exquisite epitope the field. When necessary to defy apparently recog nition of antibody conceptually guaran- grim probabilities, the quote of my compatriots, tees a much higher specificity than natural mol- “Impossible n’est pas français,” naturally comes ecules. However, multiple hurdles – for which to mind. (“Impossible n’est pas français” can be cost and time are not the least factors – exist on translated in English as “There is no such word the road of production of monoclonal antibodies as can’t”. This sentence attributed to Napoleon tolerated by the human immune system. Even is actually a pun meaning both ‘impossible is not if technical obstacles can be overcome given French’ and ‘impossible is not idiomatically cor- enough time and work, impediments linked to rect’.) Indeed, with the accelerated development the immunization process cannot. of targeted macromolecules engineered for cargo Antibodies are naturally produced by organ- delivery, early diagnosis and therapy of ovarian isms in contact with foreign and immunogenic cancer now appear as a tangible future. molecules, two characteristics that most tumor- Macromolecules preferentially accumulate associated antigens lack. Tumor-associated anti- in tumors due to the enhanced permeability gens are mainly endogenous molecules over- and retention effect that arises from the dif- expressed by tumors rather than being tumor ference in clearance rate between solid tumors specific, and are often conserved between and normal tissues [9]. Various imaging and humans and animals used for immunization. In therapeutic reagents or reporters for molecular addition, the ideal epitopes for in vivo targeting and functional imaging studies, ranging from cell-surfaced, overexpressed molecules are most 10.2217/THY.10.19 © 2010 Future Medicine Ltd Therapy (2010) 77(3),(3), 209–212 ISSN 1475-0708 209 Editorial Scholler Strategies using recombinant antibodies for early diagnosis & therapy of ovarian cancer Editorial likely conformational, but the engineering of cell lines or tissues, in vitro or in vivo [30]. As recombinant molecules bearing conformational the screening of recombinant antibody libraries epitopes remains challenging. Finally, most identifies the best specific binders from a pool of tumor-associated antigens expressed by tumor existing molecules without any de novo genera- cells display altered post-translational modifi- tion through immun ization, the successful isola- cations, including glycosylation [22], which are tion of effective affinity reagents only depends both difficult to reproduce in vitro and poorly on the library size and diversity. The screening immuno genic. Consequently, instead of mount- process can involve several rounds of selection, ing an immune response against conformational including the use of both positive and nega- epitopes or post-translational modifications tive selection, to permit the isolation of affin- uniquely expressed by tumor-associated anti- ity reagents highly specific for tumor-associated gens, immunized animals preferentially develop antigens. For example, the positive screening of antibodies directed against the nonconserved recombinant antibodies that bind to tumor cells regions of the recombinant proteins or linear expressing tumor-associated antigens and that peptides used for immunization. This can yield cross-react between mouse and human antigens production of antibodies that do not cross-react can be followed by the negative screening of with the corresponding tumor-associated anti- the selected recombinant antibodies that bind gen despite their high affinity for the immuniz- to similar antigens presented by normal cells. ing agent. Thus, animal immunization does It is worth noting that recombinant antibod- not permit the production of affinity reagents ies derived from human B lymphocytes do not specifically optimized for cancer targeting. require the additional step of humanization for Furthermore, antibodies produced in animals are safe use in the clinic, as they display little or no often immuno genic themselves in humans and, immunogenicity. Altogether, carefully designed thus, require additional molecular re-engineering screening strategies of recombinant antibody (‘humanization’) for safe clinical use. Finally, the libraries have the potential to lead to the identifi- absence of mouse antibody cross-reactivity with cation of ideal targeting reagents for both in vitro mouse antigens requires performing preclinical and in vivo uses. experiments in human xenograft mouse models, where it is impossible to assess the poten- “Alternative methods of coupling need to be tially harmful side effects caused by antibody developed for in vivo usage. We believe that offsite (i.e., non tumor) binding. In conclusion, enzymatic cross-linking using although natural antibodies have proved invalu- transglutaminases offers a powerful able in the diagnosis and treatment of infectious potential for this specific application. and auto immune diseases, their use for cancer ” diagnosis or treatment may generate detection assays unable to distinguish between cases and Recombinant antibodies can be further tai- controls, and produce offsite effects that cannot lored for specific applications using random be addressed with the current preclinical models. evolution to increase affinity, molecular design The identification of targeting reagents by to optimize in vivo stability by adding disulfide screening recombinant antibody libraries circum- bonds or constant domains, or fusion with vari- vents caveats linked to immun ization. Libraries ous structures for functionalization. For exam- of recombinant antibodies are produced by ran- ple, T cells transduced with viruses encoding domly recombining the hypervariable regions of cell-surface-expressed recombinant antibodies the heavy and light immunoglobulin chains nat- fused to the CD28/TCR-z chimeric signal- urally produced by B lymphocytes or synthesized ing domain can be specifically activated after in vitro [23,24], and linking them together with [25] antigen binding and can kill tumor cells [31]. or without addition of constant domains to build Alternatively, recombinant antibodies can be Fab or scFv, respectively. More recently, novel genetically or chemically fused with bacterial or classes of very small recombinant anti bodies have plant toxins for use as immunotoxins [32]. The been developed using only one domain derived fusion of recombinant antibodies to detection from camelid immunoglobulin hypervariable reagents, toxins or nanoparticles is crucial for regions [26], or from synthetic hypervariable both in vitro diagnostic tests as well as for in vivo regions [27].

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