Walter A. Henne

Synthesis and Biological Analysis of Prostate-Specific Membrane Antigen-Targeted Anticancer Prodrugs

Ligand-targeted therapeutics have increased in prominence because of their potential for improved potency and reduced toxicity. However, with the advent of personalized medicine, a need for greater versatility in ligand-targeted drug design has emerged, where each tumor-targeting ligand should be capable of delivering a variety of therapeutic agents to the same tumor, each therapeutic agent being selected for its activity on a specific patients cancer. In this report, we describe the use of a prostate-specific membrane antigen (PSMA)-targeting ligand to deliver multiple unrelated cytotoxic drugs to human prostate cancer (LNCaP) cells. We demonstrate that the PSMA-specific ligand, 2-[3-(1, 3-dicarboxy propyl)ureido] pentanedioic acid, is capable of mediating the targeted killing of LNCaP cells with many different therapeutic warheads. These results suggest that flexibility can be designed into ligand-targeted therapeutics, enabling adaptation of a single targeting ligand for the treatment of patients with different sensitivities to different chemotherapies.

Imaging Sites of Infection Using a 99mTc-Labeled Folate Conjugate Targeted to Folate Receptor Positive Macrophages

EC20, a folate-targeted (99m)Tc based radioimaging agent with a high folate receptor (FR) binding affinity, has been used for both the diagnosis and the staging of FR positive malignancies (currently in phase III trials) and also for the localization of inflamed lesions characterized by the accumulation of FR+ macrophages. Because recent evidence has suggested that FR+ macrophages might accumulate at sites of infectious disease, this study evaluated whether EC20 might prove similarly useful for imaging bacterial infection foci. Using gamma scintigraphic imaging, it was demonstrated that EC20 accumulated at sites of Staphylococcus aureus infection with a significant difference (P < 0.0001, n = 12) in enrichment noted between infected and noninfected limbs. Confirmation that the elevated uptake of EC20 in infected limbs was FR-mediated was supported by suppression of EC20 accumulation in the presence of a 200-fold excess of free folic acid (P < 0.0001, n = 12). This study establishes for the first time the use of EC20 to image and localize sites of infectious disease.

Synthesis and activity of a folate targeted monodisperse PEG camptothecin conjugate

A folate targeted camptothecin small molecule drug conjugate (SMDC) was synthesized using a monodisperse PEG spacer linked to folate via a releasable disulfide carbonate linker. Cell cytotoxicity in human KB cells exhibited an IC50 of 6nM. Importantly, activity of the prodrug was blocked by excess folate, demonstrating receptor-mediated celluar uptake of the PEG conjugate.

Synthesis and activity of folate conjugated didemnin B for potential treatment of inflammatory diseases.

A folate receptor targeted didemnin B conjugate was synthesized using a hydrophilic peptide spacer linked to folate via a releasable disulfide carbonate linker. Cell cytotoxicity and TNF-α inhibition in RAW264.7 macrophage-like cells exhibited IC(50)s of 13 and 5 nM, respectively. Folate didemnin B was found to be ∼50-100 fold more potent than didemnin B itself. More importantly, activity of the prodrug was blocked by excess folic acid, demonstrating receptor-mediated cellular uptake of the conjugate.

Sensing of Bacillus Subtilis Spores with Peptide Functionalized Microcantilevers

Short peptide ligands were synthesized to selectively capture spores using microcantilever arrays. Our initial studies focused on the capture of Bacillus subtilis (a stimulant of Bacillus anthracis) spores in liquids. A peptide-ligand-functionalized microcantilever array was mounted on a flow cell filled with a B. subtilis suspension while cantilever deflection was monitored. Also, fifth mode resonant frequency measurements were performed before and after dipping a peptide-ligand-functionalized microcantilever array in a static B. subtilis solution. Both techniques showed selective binding to functionalized cantilevers when compared to arrays functionalized with a similar control (non-binding) peptide. Conclusive confirmation of selective binding was obtained by subsequent examination of the microcantilever arrays under a dark field microscope. The concentration sensitivity, shelf-life of the binding ligands, and their binding efficiency were studied as well.