The tails were dilated with warm water and animal were injected iv. to a 2-arylaminobenzothiazole moiety, resulting in an analog with improved physicochemical properties, solubility and kidney:tumor ratio while maintaining potency (6; IC50 KP372-1 = 53 pM). The results presented herein utilized heterocyclic and solid-phase chemistry, cell adhesion assay, and optical imaging using the cyanine dye Cy5.5 conjugate. Introduction Many of the current cancer chemotherapeutic agents clinically deployed today are designed to be indiscriminately cytotoxic through DNA alkylation, unnatural base-pair incorporation, inhibition of topoisomerases, and microtubule stabilization mechanisms. Cancer chemotherapy, often administered near its maximum tolerated dose (MTD), aims to annihilate tumors without systemic toxicity. Several of these agents exemplified in Figure 1 possess a narrow therapeutic index that limits effectiveness. As a consequence, under-dosing at the tumor site is problematic with patients suffering from intolerable side effects including nausea, vomiting, diarrhea, malnutrition, hair and memory loss, anemia, immunosuppression, hemorrhaging, chronic pain, and various organ KP372-1 toxicities. Tremendous success has been achieved through lengthy syntheses of ornate cytotoxic natural products with significantly less attention being granted towards chemotherapeutics that would result in decreased off-target binding and ensuing side effects. Open in a separate window Figure 1 Structures of commonly administered indiscriminating cytotoxic chemotherapeutic agents. To achieve target selectivity, therapeutic compounds must be able to differentiate cancer cells from normal cells. In T- and B-cell lymphomas, targeting the activated form of cell surface receptors expressed on cancer cells allows for differentiation, as normal or inactivated versions remain untargeted. Specifically, the cell surface receptor 41 integrin regulates lymphocyte trafficking1 and homing in normal adult cells.2,3 A -subunit conformational change4 activates 41, which regulates tumor growth, metastasis, and angiogenesis, in addition to promoting the dissemination of tumor cells to distal organs.5 The ligand LLP2A (1; see Figure 2) recognizes this change and shows potential as a non-invasive imaging and therapeutic agent despite kidney uptake observed in xenograft models.6 This prompted creation of a KP372-1 water soluble benzimidazole analog KLCA4 (2)7 that would be dianionic8 at physiological pH (bisarylamino NH + CO2H) thereby improving solubility and decreasing kidney uptake based on electronic factors.7,9,10 While 2 has picomolar potency, it is still 10-fold less potent than the bisaryl urea 1. Herein, we report the design of an equipotent (to 1 1), comparably soluble (to 2) benzothiazole analog 6 that, when optically conjugated using Cy5.5, demonstrates excellent HSPA1 tumor uptake with preliminary evidence showing improved kidney:tumor ratios in xenograft models. Key to this approach is the heterocyclic design, which, in a condensed fashion, improves the ligand’s physicochemical properties without PEGylation or a poly-charged tail. Open in a separate window Figure 2 Evolution of ligand analogs using function-oriented synthesis (FOS): (a) Structure of 1 1 (LLP2A) which can be optically KP372-1 or radio conjugated and shows potential as an imaging or therapeutic agent for lypmphoma;6 (c) water soluble benzimidazole 2 analog (IC50 = 305 pM);7 (d) heterocyclic analogs 3-11 and requisite precursor heterocyclic acids 12-21 and aryl isothiocyanates 22a-d. Results and Discussion Our previous benzimidazole ligand showed excellent binding to human 41 integrin; however, it still did not bind as efficiently as LLP2A (1). In an attempt to regain the binding affinity and retain the desirable pharmacokinetics, systematic modifications to the heterocycle within the ring structure and the side chain were created. As delineated in Scheme 1, biological activity before radio studies. Scheme 2 delineates the synthesis of 6-Cy5.5 starting with Rink amide resin, followed by a series of Fmoc-deprotection and specificity and uptake measurements of pertinent organs and tumors for both agents at similar doses. Tumor uptake was observed as early as 5 min post injection and persisted for up to 120 h. To determine organ:tumor ratios, regions of interest were drawn around the tumor and each organ in the images and mean signal intensity was obtained by subtracting the lowest intensity background signal (heart) from each intensity value. Preliminary evidence indicates 6-Cy5.5 is comparable to 1-Cy5.5 in terms of tumor specificity and.