{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T16:54:45Z","timestamp":1785430485925,"version":"3.56.0"},"reference-count":23,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2026,4,11]],"date-time":"2026-04-11T00:00:00Z","timestamp":1775865600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Liaoning Normal University","award":["H2024041"],"award-info":[{"award-number":["H2024041"]}]},{"award":["H2024041"],"award-info":[{"award-number":["H2024041"]}],"id":[{"id":"https:\/\/ror.org\/04c3cgg32","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>Although the parental recombinant protein rLj-RGD3 exhibits antitumor activity, it carries immunogenicity risks owing to its large molecular size (13.5 kDa). We generated a genetically truncated mutant, rLj-RGD4 (6.27 kDa, four RGD motifs), which inhibited B16 melanoma cell proliferation, migration, and invasion in vitro. However, the in vivo efficacy and mechanisms of action remain unclear. Here, B16 xenograft mice were treated with rLj-RGD4 (5, 10, and 20 \u03bcg\/kg i.p. daily for 14 days). Tumor growth was measured, and histopathology\/apoptosis was evaluated using hematoxylin and eosin (HE), Masson\u2019s dye, Hoechst, and TUNEL staining. Apoptotic pathways (mitochondrial, death receptor, and MAPK) were analyzed via Western blotting, whereas endocytosis mechanisms were explored using inhibitors (filipin III, NaN3, cytochalasin D), and EGFR (epidermal growth factor receptor) interactions via fluorescence co-localization and phosphoprotein assays. The results demonstrated dose-dependent tumor growth inhibition (21.60\u201389.26% volume reduction, 41.03\u201386.51% weight reduction), with histological evidence of tissue loosening, fibrosis, and apoptosis. rLj-RGD4 induced apoptosis by activating the mitochondrial (Bax\/Bcl-2 upregulation), death receptor (caspase-8 activation), and MAPK (JNK\/p38 phosphorylation) pathways. Internalization was blocked by NaN3 and cytochalasin D, indicating actin-dependent macropinocytosis. Direct EGFR binding was confirmed, accompanied by reduced EGFR expression and the inhibition of FAK\/AKT\/Src signaling. In conclusion, rLj-RGD4 exerts potent in vivo antitumor activity via two mechanisms: induction of multi-pathway apoptosis and EGFR-targeted suppression of pro-survival signaling. RGD4 exerts its antitumor function in vivo by targeting and co-internalizing with EGFR.<\/jats:p>","DOI":"10.3390\/molecules31081266","type":"journal-article","created":{"date-parts":[[2026,4,13]],"date-time":"2026-04-13T07:29:29Z","timestamp":1776065369000},"page":"1266","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["A Genetically Truncated RGD-Containing Peptide rLj-RGD4 Exhibits Potent In Vivo Antitumor Activity via Induction of Multi-Pathway Apoptosis and EGFR-Targeted Signaling Suppression"],"prefix":"10.3390","volume":"31","author":[{"given":"Yuyao","family":"Song","sequence":"first","affiliation":[{"name":"School of Life Sciences, Liaoning Normal University, Dalian 116081, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huijie","family":"Yan","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Liaoning Normal University, Dalian 116081, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuebin","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Pharmacology, College of Pharmacy, Dalian Medical University, Dalian 116044, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jingyu","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Pharmacology, College of Pharmacy, Dalian Medical University, Dalian 116044, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1157-853X","authenticated-orcid":false,"given":"Li","family":"Lv","sequence":"additional","affiliation":[{"name":"Department of Pharmacology, College of Pharmacy, Dalian Medical University, Dalian 116044, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jihong","family":"Wang","sequence":"additional","affiliation":[{"name":"School of Life Sciences, Liaoning Normal University, Dalian 116081, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1615\/CritRevEukaryotGeneExpr.2020028454","article-title":"Malignant Melanoma: Skin Cancer-Diagnosis, Prevention, and Treatment","volume":"30","author":"Ahmed","year":"2020","journal-title":"Crit. 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