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Indole-containing natural products have long been an inspiration to synthetic chemists. Melanin pigments in human skin, hair, and eyes, among the most visible of natural products, are indole-based materials.  Eumelanin, the brown to black pigment in humans, is known to be composed of dihydroxyindole monomers; these are thought to form relatively short covalent oligomers that self-assemble into nanoparticles. The oligomers are composed of four to eight monomers and are heterogeneous in nature.  Such oligomers are within the range of modern organic synthesis, similar to oligoarene foldamers.  The ability to produce well-defined synthetic oligomers will advance our knowledge of this common but poorly understood biomaterial.  This grant focuses on developing the synthetic methods necessary to generate indole and dihydroxyindole oligomers of relevance to eumelanin.

Palladium catalyzed cross-coupling of aryl boronic acids with aryl halides, the Suzuki reaction, was one of the reactions highlighted in this year's Nobel Prize in Chemistry.  It has become a central method for oligoarene synthesis.  However, until recently indole-indole Suzuki couplings have been rare; thus, we began by examining indole dimerization.  Prior to the initiation of PRF funding, we had investigated Zhang and co-workers' method¹ for ligand-free Suzuki reactions.  In our hands, this method was inefficient for cross-coupling of indoles, but when the bromide partner was serendipitously left out of a reaction, we obtained a symmetrical dimer from homocoupling of 4-boronic acid indole.  Repeating the serendipitous reaction with an increased catalyst loading of 5 mol% Pd(OAc)₂ provided the homocoupling product 4-4' biindolyl in 69% yield.  At the beginning of the grant period, we focused on optimization of this homocoupling reaction.  In particular, following the work of Kabalka,² we found that including tosyl chloride (0.5 eq. relative to boron) in the reaction of 4-boronic acid indole yielded 4-4' biindolyl in 81% yield.  Similarly, 5-5' biindolyl and 6-6' biindolyl were obtained in quantitative yield from 5- and 6-boronic acid indole, respectively, under the same conditions.  The homocoupling reactions are fast and operationally simple, employing air, water, and room temperature.   A manuscript on indole boronic acid homocoupling with three undergraduate co-authors is in preparation.

Following our work on the homocoupling reaction, we have returned to Suzuki cross-coupling of indoles, following the successful work of Buchwald³ and Huleatt an Chai.⁴   Last year we began to develop a set of indole monomers by installing the requisite boronates and halogens regioselectively at appropriate positions on the indole ring.   In eumelanin, the key linkages between dihydroxyindoles are the 2, 4, and 7 positions.  Coincidentally, these are the same positions that are reactive to iridium-catalyzed C-H activation/borylation.⁵  We have used these regioselective reactions to install one or two boronate groups per indole.  These can be further transformed to bromides using a copper-mediated boron-to-bromide functional group exchange,⁶ which preserves the desired regiochemistry.  This year we have successfully utilized these monomers in Suzuki cross-coupling reactions to generate several indole trimers. These compounds have interesting optical properties, which we aim to investigate further, as well as their potential self-assembly and biological recognition properties.  A manuscript on the indole functionalization and trimer synthesis with one undergraduate co-author is in preparation.

Most excitingly, we extended these studies to produce out first trimer via an iterative route.  The key building block in this iterative synthesis is a 4-bromo-7-boronate indole where the boronic acid group is protected using Suginome's diaminonaphthlene protecting group (B-dan).⁷  To install this protecting group, we devised a novel one-pot pinacolboronate to B-dan exchange.  With an iterative synthetic strategy in hand, and set of monomers from borylation/halogenation and boronate exchange, we are in position to generate a range of indole oligomers with diverse linkages, and to begin to extend these studies to protected dihydroxyindole monomers.    

Working with both melanin and boronic acids, we became interested in using boronic acids as inhibitors of melanin formation via reversible covalent bond formation between boronic acids and the catechol-like functionality of dihydroxyindole intermediates.  Following early work from Mishima and co-workers,⁸ we developed a simple spectroscopic assay to study this inhibition with using commercially available aryl boronic acids.⁹  We are currently screening a broader range of heteroaryl boronic acids in our original assay, as well as investigating the interactions between boronic acids, the enzyme tyrosinase (which initiates eumelanin biosynthesis) and the tyrosinase substrate L-dopa (which is itself a catechol).  Several mechanisms of inhibition, including classical competitive inhibition (boronic acid binds tyrosinase) and substrate depletion (boronic acid binds L-dopa), are operative depending on the structure of the boronic acid.  Our next step is to investigate the most promising inhibitors in cell culture.  Inhibitors of melanin formation have materials, therapeutic, and cosmetic applications.

References:

1.  Lui, L.; Zhang, Y.; Xin, B. "Synthesis of Biaryls and Polyaryls by Ligand-Free Suzuki Reaction in Aqueous Phase"  J. Org. Chem. 2006, 71, 3994-3997.

2.  Kabalka, G. W.; Wang, L.  "Ligandless palladium chloride-catalyzes homo-coupling of arylboronic acids in aqueous media" Tetrahedron Lett. 2002, 43, 3067-3068.

3.  Billingsley, K.; Buchwald, S. L. "Highly Efficient Monophosphine-Based Catalyst for the Palladium-Catalyzed Suzuki-Miyaura Reaction of Heteroaryl Halides and Heteroaryl Boronic Acids and Esters" J. Am. Chem. Soc. 2007, 129, 3358-3366.

4.  Duong, H. A.; Chua, S.; Huleatt, P. B.; Chai, C. L. L.  "Synthesis of Biindolyls via Palladium-Catalyzed Reactions" J. Org. Chem. 2008, 73, 9177-9180.

5.   Lo, W. F.; Kaiser, H. M.; Spannenberg, A.; Beller, M.; Tse, M. K. "A highly selective Ir-catalyzed borylation of 2-substituted indoles: a new access to 2,7- and 2,4,7-substituted indoles" Tetrahedron Lett. 2007, 48, 371-375.

6. Thompson, A. L. S.; Kabalka, G. W.; Akula, M. R.; Huffman, J. W. "The Conversion of Phenols to the Corresponding Aryl Halides Under Mild Conditions" Synthesis 2004, 547-550.

7.  Noguchi, H.; Hojo, K.; Suginome, M.  "Boron-masking strategy of the selective synthesis of oligoarenes via iterative Suzuki-Miyaura coupling" J. Am. Chem. Soc. 2007, 129, 758-759.

8.  Mishima, Y.; Kondoh, H.  "Dual control of melanogenesis andd melanoma growth: overview molecular to clinical level and the reverse" Pigment Cell Res. 2000, Suppl. 8, 10-22.

9.  Belitsky, J. M. "Aryl boronic acid inhibition of synthetic melanin polymerization" Bioorg. Med. Chem. Lett. 2010, 20, 4475-78.