Light-Driven Boron-Carbon Bond Construction: Synthesis of Carboranyl Peptides Targeting BNCT

A study titled "Photocatalyzed Decarboxylative B−C Couplings for the Synthesis of Carboranyl Amino Acids and Peptides," published in the Journal of the American Chemical Society, was led by the research team of Feng Zhu, Bo Yang, and Lijuan Zhu from Shanghai Jiao Tong University. Addressing the urgent need for novel and efficient boron delivery agents in boron neutron capture therapy (BNCT), this work developed a new visible-light-driven decarboxylative B–C cross-coupling method. The strategy utilizes carborane carboxylic acids to generate boron-vertex-centered radicals under photocatalysis, which then undergo highly regioselective Giese-type addition with dehydroalanine (Dha) residues in peptide chains. This approach enables, for the first time, the synthesis of B-vertex-substituted carboranyl peptides under mild conditions. The method exhibits excellent functional group tolerance and broad substrate scope, allowing efficient synthesis of enantiomerically pure carboranyl alanines. It has been successfully applied to solid-phase peptide synthesis, DNA-encoded library construction, and bioconjugation with targeting aptamers. Preliminary cell experiments confirmed that the resulting aptamer-carborane conjugates are efficiently recognized and internalized by tumor cells, providing a powerful chemical platform for developing next-generation targeted BNCT drugs.

01 Research Background​

Boron Neutron Capture Therapy (BNCT) is a "binary" radiotherapy strategy capable of selectively killing cancer cells while sparing healthy tissues. Its efficacy highly depends on carriers that can selectively deliver sufficient boron-10 (¹⁰B) to tumor cells. Current clinically used boron delivery agents (e.g., BPA and BSH) have limitations in tumor selectivity, boron content, or metabolic stability, underscoring the need for new-generation drugs. Carboranes, with their extremely high boron content and excellent chemical and metabolic stability, are ideal boron carriers. However, existing strategies for incorporating carboranes into biomolecules (e.g., peptides) mainly rely on functionalization at the carbon (C) vertices, while selective modification at the boron (B) vertices—especially the construction of stable B–C bonds—remains highly difficult due to the chemical inertness of B–H bonds and the similar environments of multiple B atoms. B-vertex modification can impart electron-donating properties to carboranes, complementing the electron-withdrawing nature of C-vertex modification, thereby opening avenues for exploring new structure-activity relationships. Meanwhile, dehydroalanine (Dha), an electrophilic unnatural amino acid that can be efficiently incorporated into peptide chains, serves as a versatile handle for site-selective modification. However, its use for constructing B–C bonds with carboranes has not been reported. Therefore, developing a mild and efficient method for site-specific conjugation of carborane B-vertices with bioactive peptides is of great significance for expanding the chemical space of BNCT drugs.

02 Innovative Highlights​

First realization of photocatalyzed B-vertex radical-based B–C coupling with peptides

The team pioneered the use of visible-light photocatalysis (violet LED) to trigger decarboxylation of carborane carboxylic acids, producing highly reactive boron-vertex-centered carboranyl radicals in situ. These radicals efficiently and regioselectively undergo Giese-type addition with Dha residues in peptide chains to form B–C bonds. The reaction proceeds under mild conditions (45°C), without transition-metal catalysis, and exhibits excellent functional group tolerance, offering a new paradigm for carborane bioconjugation.

Site-selective and modular synthesis of B-vertex carboranyl peptides achieved

This strategy enables site-selective late-stage modification of peptide chains, with the position of Dha determining the site of carborane introduction. Using chiral Karady–Beckwith-type Dha derivatives, the first highly diastereoselective synthesis (d.r. > 20:1) of enantiomerically pure B-vertex-substituted L- and D-carboranyl alanines was achieved, paving the way for introducing carboranyl unnatural amino acids with defined chirality.

Successful integration from basic building blocks to complex bioconjugates

The study demonstrates the powerful synthetic capability of this platform: ① The resulting carboranyl alanines can serve as monomers and be successfully assembled via solid-phase peptide synthesis into defined-sequence peptides containing 5 to 15 amino acids; ② Through click chemistry, carborane–amino acid modules were conjugated with DNA tags, introducing boron cluster diversity into DNA-encoded compound libraries; ③ Carborane amino acids were conjugated with nucleic acid aptamers targeting the tumor surface protein EpCAM, constructing targeted delivery systems.

Targeting and cellular uptake ability of conjugates confirmed

Preliminary cellular studies demonstrated that the constructed fluorescently labeled aptamer-carborane conjugates effectively target and are internalized into EpCAM-high cancer cells (HT29 and MDA-MB-231). The intracellular boron content exceeded the therapeutic threshold for BNCT (>10⁹ atoms/cell), proving the practical value of this chemical platform in constructing bioactive bioconjugates.

03 Results and Discussion​

3.1 Reaction development and condition optimization​

Using meta-carborane carboxylic acid 13a​ and N-Boc-dehydroalanine 14a​ as model substrates, detailed condition screening established the optimal reaction system: photocatalyst Ir[dF(CF₃)ppy]₂(dtbbpy)PF₆ (PC-1), Cs₂CO₃ as base, acetonitrile as solvent, under irradiation with an 18W violet LED at 45°C for 24 hours under a nitrogen atmosphere, affording the target B–C coupling product 16​ in 86% isolated yield. Control experiments confirmed that light, photocatalyst, and base are all essential.

3.2 Substrate scope investigation​

Dha peptide substrate scope: The reaction shows broad applicability to dipeptides and triptides containing Dha. Regardless of whether Dha is located at the N-terminus, C-terminus, or within the peptide chain, and irrespective of neighboring amino acids being glycine, alanine, valine, phenylalanine, or amino acids with hydroxyl (serine), carboxyl (aspartic acid), amino (lysine), or thioether (methionine) side chains, the reaction proceeds smoothly, yielding the corresponding carboranyl peptides in moderate to excellent yields (49%–84%).

Carborane carboxylic acid substrate scope: A series of meta- and ortho-carborane carboxylic acids with different substituents (alkyl, aryl) on the carbon atoms successfully participated in the reaction, affording the corresponding products in good to excellent yields (up to 89%), demonstrating the generality of the method.

3.3 Synthetic applications​

Gram-scale preparation and derivatization: The reaction could be scaled up to 3 mmol with good yield (71%). The chiral product could be converted via deprotection and Fmoc protection into building block N-Fmoc-protected B-carboranyl alanine (51) suitable for solid-phase peptide synthesis.

Solid-phase and solution-phase peptide synthesis: Building block 51​ was successfully incorporated via standard Fmoc solid-phase synthesis strategies into peptide chains containing 5 and 15 amino acids. Simultaneously, the triptide Cba-Val-Phe was successfully synthesized in solution phase, demonstrating compatibility with conventional peptide synthesis techniques.

DNA-encoded library compatibility: The study conjugated the carborane–amino acid module to DNA tags via copper-catalyzed azide-alkyne cycloaddition (CuAAC), successfully constructing carborane–amino acid–DNA conjugates, proving the potential of this chemistry for DNA-encoded library technology.

3.4 Bioconjugation and preliminary cellular evaluation​

Azide-functionalized B-vertex carboranyl alanine derivative 56​ was conjugated via CuAAC to a DBCO-modified, EpCAM-targeting fluorescently labeled aptamer, yielding conjugate FAM-Epcam-B. Successful conjugation was confirmed by gel electrophoresis and mass spectrometry. Flow cytometry and confocal microscopy imaging showed that compared to unmodified aptamer, FAM-Epcam-B exhibited significantly enhanced internalization in EpCAM-high HT29 and MDA-MB-231 cells. ICP-MS quantitative analysis revealed that treated HT29 cells contained 7.7×10⁹ boron atoms per cell, exceeding the therapeutic threshold for BNCT.

3.5 Proposed reaction mechanism​

Based on experimental data and literature, a plausible photocatalytic cycle mechanism is proposed: upon photoexcitation, the photocatalyst oxidizes the carborane carboxylate anion, which undergoes decarboxylation to generate the key boron-vertex carboranyl radical; this radical adds to the double bond of Dha, producing a new radical intermediate; this intermediate is reduced by the reduced photocatalyst and protonated to yield the final product. Radical trapping experiments support the existence of the carboranyl radical intermediate.

04 Conclusion and Future Perspectives​

This research successfully developed a visible-light-photocatalyzed decarboxylative B–C cross-coupling reaction, achieving for the first time the efficient and site-selective linkage of carborane B-vertices with dehydroalanine residues in peptide chains. This work not only provides a general method for synthesizing structurally novel B-vertex-substituted carboranyl peptides but also successfully applies it to construct carborane–nucleic acid aptamer targeted conjugates and demonstrates its potential in DNA-encoded library technology.

The core breakthrough of this work lies in utilizing photocatalytic radical chemistry to overcome the challenge of selective functionalization at carborane B-vertices and seamlessly integrating this reaction with peptide and biomacromolecule modification. The established platform combines mild conditions, modularity, and versatility, providing a powerful chemical tool for carborane-based BNCT drugs, molecular probes, and even new drug discovery. Future directions may include: ① Extending this strategy to conjugate carborane modules to other, more complex biomacromolecules such as proteins and antibodies; ② Conducting systematic in vivo efficacy, pharmacokinetic, and safety evaluations of the constructed targeted carborane conjugates to advance their clinical translation; ③ Exploring applications of B-vertex carboranyl peptides in other biomedical fields, such as protein stabilization and inhibitor design. In summary, this research opens a promising new avenue at the intersection of boron cluster chemistry, chemical biology, and cancer therapeutics.