Potential Dual Targeting of EGFR and VEGFR2 by Repurposed Drugs for Cholangiocarcinoma: A Molecular Docking Study

Öz

Objective: Cholangiocarcinoma is an aggressive hepatobiliary malignancy characterized by high mortality, late-stage diagnosis, and limited systemic treatment options. The epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor 2 (VEGFR2) signaling pathways play central roles in tumor cell proliferation, survival, and tumor angiogenesis. This study investigated the potential inhibitory interactions of three commonly used non-oncologic drugs, doxycycline, propranolol, and metformin, with EGFR and VEGFR2 using molecular docking analysis, and compared their binding profiles with that of sorafenib, a reference multi-target kinase inhibitor.

Materials and Methods: The crystal structures of EGFR (PDB ID: 1M17) and VEGFR2 (PDB ID: 4ASD) kinase domains were retrieved from the Protein Data Bank and prepared using OpenBabel. Three-dimensional ligand structures were generated from SMILES representations using RDKit, energy-minimized with the MMFF94 force field, and converted to PDBQT format using Meeko. Docking simulations were performed with AutoDock Vina (version 1.2). The docking protocol was validated by multi-pose redocking of the co-crystallized ligands (erlotinib and sorafenib), with the lowest root-mean-square deviation (RMSD) value reported. Ligand-protein interactions were evaluated using an in-house geometric classification algorithm and independently verified using the Receptor-Ligand Interactions module of BIOVIA Discovery Studio Visualizer. Drug-likeness and physicochemical properties were assessed using RDKit according to Lipinski's rule of five.

Results: Docking protocol validation yielded RMSD values of 1.32 Å for EGFR and 0.30 Å for VEGFR2, confirming the reliability of the docking workflow. Among the repurposing candidates, doxycycline exhibited the strongest binding affinity toward both EGFR (-8.82 kcal/mol) and VEGFR2 (-8.17 kcal/mol), with binding energies approaching those of the reference compound sorafenib (EGFR: -9.01 kcal/mol; VEGFR2: -10.54 kcal/mol). Propranolol showed moderate binding affinity, whereas metformin displayed the weakest interactions with both targets. Doxycycline formed conventional hydrogen bonds with the catalytic Asp831 (DFG motif) and hinge-region Thr766 residues of EGFR, and with Asp1046 (DFG motif) and Asp814 residues of VEGFR2. All investigated compounds satisfied Lipinski's rule of five.

Conclusion: These findings highlight doxycycline as a noteworthy in silico dual-target candidate against EGFR and VEGFR2 in cholangiocarcinoma, warranting further experimental evaluation. The validated docking workflow and consistent interaction patterns support this observation; however, these findings remain hypothesis-generating and require confirmation through experimental studies before any biological or therapeutic conclusions can be drawn.

Anahtar Kelimeler

• Cholangiocarcinoma
• EGFR
• VEGFR2
• drug repurposing
• molecular docking
• doxycycline
• AutoDock Vina

Kolanjiyokarsinom İçin Yeniden Konumlandırılan İlaçların EGFR ve VEGFR2'yi Potansiyel Çift Hedeflemesi: Bir Moleküler Docking Çalışması

Öz

Amaç: Kolanjiokarsinom; yüksek mortalite, geç evrede tanı ve sınırlı sistemik tedavi seçenekleriyle karakterize agresif bir hepatobiliyer malignitedir. Epidermal büyüme faktörü reseptörü (EGFR) ve vasküler endotelyal büyüme faktörü reseptörü 2 (VEGFR2) sinyal yolakları, tümör hücre proliferasyonu, sağkalımı ve tümör anjiyogenezinde merkezi rol oynamaktadır. Bu çalışmada, yaygın kullanılan üç onkolojik olmayan ilacın doksisiklin, propranolol ve metformin EGFR ve VEGFR2 ile potansiyel inhibitör etkileşimleri moleküler docking analiziyle araştırılmış ve bağlanma profilleri referans çok-hedefli bir kinaz inhibitörü olan sorafenib ile karşılaştırılmıştır.

Gereç ve Yöntemler: EGFR (PDB ID: 1M17) ve VEGFR2 (PDB ID: 4ASD) kinaz domainlerinin kristal yapıları Protein Veri Bankası’ndan temin edilmiş ve OpenBabel ile hazırlanmıştır. Üç boyutlu ligand yapıları RDKit kullanılarak SMILES gösterimlerinden üretilmiş, MMFF94 kuvvet alanı ile enerjisi minimize edilmiş ve Meeko ile PDBQT formatına dönüştürülmüştür. Docking simülasyonları AutoDock Vina (versiyon 1.2) ile gerçekleştirilmiştir. Docking protokolü, ko-kristalize ligandların (erlotinib ve sorafenib) çoklu-poz redocking’i ile doğrulanmış, en düşük RMSD (kök ortalama kare sapması) değeri raporlanmıştır. Ligand-protein etkileşimleri özgün bir geometrik sınıflandırma algoritmasıyla değerlendirilmiş ve BIOVIA Discovery Studio Visualizer’ın Receptor-Ligand Interactions modülüyle bağımsız olarak doğrulanmıştır. İlaç benzerliği ve fizikokimyasal özellikler, Lipinski’nin beşli kuralına göre RDKit kullanılarak değerlendirilmiştir.

Bulgular: Docking protokolü doğrulaması, EGFR için 1.32 Å ve VEGFR2 için 0.30 Å RMSD değerleri vermiş, docking iş akışının güvenilirliğini teyit etmiştir. Repozisyon adayları arasında doksisiklin, hem EGFR’ye (-8.82 kcal/mol) hem de VEGFR2’ye (-8.17 kcal/mol) karşı en güçlü bağlanma afinitesini göstermiş olup, bu değerler referans bileşik sorafenibinkine (EGFR: -9.01 kcal/mol; VEGFR2: -10.54 kcal/mol) yaklaşmaktadır. Propranolol orta düzeyde bağlanma afinitesi gösterirken, metformin her iki hedefle de en zayıf etkileşimi sergilemiştir. Doksisiklin, EGFR’nin katalitik Asp831 (DFG motifi) ve menteşe bölgesi Thr766 kalıntılarıyla, VEGFR2’nin ise Asp1046 (DFG motifi) ve Asp814 kalıntılarıyla konvansiyonel hidrojen bağları oluşturmuştur. İncelenen tüm bileşikler Lipinski’nin beşli kuralına uymuştur.

Sonuç: Bu bulgular, doksisiklini kolanjiokarsinomda EGFR ve VEGFR2’ye karşı dikkat çekici bir in silico çift-hedefli aday olarak öne çıkarmakta olup, daha ileri deneysel değerlendirmeyi gerekli kılmaktadır. Doğrulanmış docking iş akışı ve tutarlı etkileşim örüntüleri bu gözlemi desteklemektedir; ancak bu bulgular hipotez üretici niteliktedir ve herhangi bir biyolojik veya terapötik sonuca varılmadan önce deneysel çalışmalarla doğrulanması gerekmektedir.

Anahtar Kelimeler

• Kolanjiyokarsinom
• EGFR
• VEGFR2
• ilaç yeniden konumlandırma
• moleküler docking
• doksisiklin
• AutoDock Vina

Kaynakça

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