Tumor associated monoclonal antibodies (mAb's) are therapeutic agents when used as selective carriers of cytotoxic agents to malignancies. This hypothesis is tested in animal model systems with mAbs directed toward antigens associated with human disease. The cytocidal agents employed are particle emitting radionuclides. The relative efficacy is evaluated in the appropriately validated murine tumor xenograft model system. The radionuclides chosen for study span the range of radionuclidic properties available permitting an assay of the effects of emission energy, half-life, and type of emission. Current research focuses on performing extensive pre-clinical studies with the alpha-particle emitting radionuclide Pb-212, with decreased interest in Bi-213, and full activation of parallel studies with the alpha emitter, At-211. Ongoing collaborative clinical trials employ the second generation bifunctional chelating agent 1B4M-DTPA (aka MX-DTPA or tiuxetan) for sequestering Y-90, a high energy pure beta emitting radionuclide well established in clinical applications and in the commercial product, Zevalin. Note that the chemistry that makes Zevalin, the 1st FDA approved radiolabeled antibody therapeutic, possible was developed by the Chemistry Section. Nearly all of those studies ongoing and planned by the Section now employ the 3rd generation bifunctional chelating agent, CHX-A''DTPA for sequestering In-111, Y-90, Bi-213, and Lu-177. Current studies initiated and ongoing continue to validate use of CHX-A''DTPA in PET imaging with the cyclotron produced (refined and purified by the Chemistry Section) Y-86. There have been a number of PET imaging studies recently reported by the Chemistry Section regarding the application of Y-86 for PET imaging targeting HER2 and HER1(EGFR) for visualizing a variety of diseases such ovarian, colorectal, pancreatic, prostate cancer;an unpublished study on Y-86 for PET imaging targeting HER1(EGFR) for imaging mesothelioma is in draft. Pre-clinical evaluation of novel bifunctional chelating agents and linkers for targeted radiotherapy with isotopes of interest continues primarily to refine conjugation chemistry functional group options and radiolabeling improvements. These refinements stem from the provision of agents for peptide chemistry as well as for site-specific conjugation strategies amenable for use with both radio-lanthanides and alpha-particle emitting radionuclides. Additionally, novel linking chemistry for extant bifunctional chelating agents has been developed for peptide usage and in use with peptide synthesizer instrumentation. Many of the established agents of the Chemistry Section are now being extended to peptides and other small delivery vectors targeting receptors of interest. This same novel chemistry and the discoveries therein are frequently found to be appropriate to the other project of the Chemistry Section. The Section has created a number of novel linkage chemistry agents for use in site-specific linkage strategies such as click chemistry and specific carbohydrate modification strategies being developed by the Qasba laboratory in Frederick. Studies with At-211 have been fully activated pending validation of the cyclotron target processing facility and instrumentation within the Chemistry Section that has originated from the upgrade in the production facilities for this radionuclide, with input obtained directly from the Zalutsky laboratory at Duke U. The Chemistry section previously reported on validation of the most stable At-211 linker reagent, N-Me-SAPS, and a recent entire re-synthesis of the agent will facilitate a parallel investigation to those ongoing with other alpha emitters. The recent addition of new personnel to the Section is anticipated in spurring this aspect of this project forward at an accelerated pace. The highly extensive and focused pre-clinical investigation into the use of Bi-213 and Pb-212 continues for the treatment of disseminated intraperitoneal disease, e.g., from either ovarian or pancreatic cancer. Bi-213 reached a point whereby evaluation of cost-effectiveness combined with failed national availability effectively forced termination of study of this radionuclide. Despite its significant efficacy, the full range of use and value of Bi-213 for therapy will probably never be defined. Development of Pb-212 continues to move forward towards a potential clinical trial;however, the planned trial with Pb-212 was relocated to UAB due to unresponsive leadership within the NCI. A pre-IND meeting with the FDA that defined relevant issues regarding required toxicity studies while GMP manufacturing was completed by an industrial CRADA partner. Murine toxicology experiments have been completed by the Section labs in support of the IND for this clinical trial to treat ovarian cancer. In addition to evaluation of the efficacy of radionuclides individually with specific mAbs, use of combined radiolabeled mAbs, and their combinations with chemotherapeutics continues to be systematically investigated. This investigation rests on the hypothesis that single doses of a single, targeted radionuclide lacks a rational basis for cancer therapy;combined modality therapies will achieve significant therapeutic enhancements. Results indicate that substantial increases in median life expectancy in murine models are possible with single doses of Bi-213 or Pb-212 conjugated to clinically relevant antibodies such as CC49(Delta)CH2, trastuzumab, cetuximab, and now panitumumab. Radiolabeling difficulties have eliminated cetuximab from further study. Use of Pb-212 labeled trastuzumab in combination with gemcitabine showed impressive enhanced therapeutic efficacy;multi-dosing of both Pb-212 and gemcitabine combined provided significant evidence that optimization of both drug combination and scheduling will extend survival. Studies combining administration of Bi-213 or Pb-212 with paclitaxel demonstrated significant extension of survival with a very strong dependence on administration scheduling. Similarly, combination with carboplatin or cisplatin has also been performed and will be the subject of a publication. The delivery of radiation to multiple molecular targets on tumors and to overcome antigen heterogeneity was also reported this year by the Chemistry Section. In sum, the use of two antibodies, CC49(Delta)CH2 and trastuzumab, both radiolabeled and administered in a variety of permutations demonstrated again the requirement for empirical determination of administration order to achieve optimal therapeutic efficacy as opposed to the reliance on in vitro cell culture studies that pose no relationship or predictability to in vivo tumor environments. Results have indicated superiority in therapeutic response to Pb-212 over Bi-213. Studies initiated in collaboration with the Gius lab to investigate and define the biological mechanisms at the cellular level of both damage response and repair as well as genetic regulation of the cell biology in response to high-LET radiation are being continued by new personnel in the Section. Studies with the Camphausen lab to evaluate the impact of tumor growth environment on genotype are being initiated. Additionally, studies with the Citrin lab are being initiated to assess impact of targeted radiation combined with external beam radiation. Studies to expand the qualities of trifunctional imaging agents combining radionuclidic imaging (SPECT or PET) and NIR dye (Optical imaging) incorporated an element of PEG. Critical discoveries were made regarding self-aggregation and signal quenching properties of dye-antibody conjugates puting a significant body of literature in doubt. This advance provides actual understanding of the fundamental chemistry for the creation of directly quantitative Optical-radionuclidic dual modality molecular imaging agents.
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