Anlotinib Enhances 177Lu-DOTA-TOC Antitumor Activity in an SSTR2-Overexpressing PC12 Pheochromocytoma Model Relevant to PPGL: Association with Cytoskeletal Remodeling and Clathrin-Sensitive Internalization

Background: Metastatic pheochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine malignancies with limited therapies. Peptide Receptor Radionuclide Therapy (PRRT) with radiolabeled somatostatin analogs is clinically relevant for SSTR-positive disease, since SSTR2 is highly expressed in patients with PPGL. Anlotinib, a multi-tyrosine kinase inhibitor approved in China for several cancers, remains investigational in PPGL. This study evaluated whether anlotinib enhances 177Lu-DOTA-TOC activity in an SSTR2-overexpressing PC12 pheochromocytoma model and examined internalization. Methods: PC12-SSTR2 cell and vector control were generated. PC12-SSTR2 cells were treated with anlotinib, 177Lu-DOTA-TOC, or their combination to assess viability, clonogenic growth, apoptosis, DNA damage, cytoskeletal morphology, and internalization. Endocytosis was probed with chlorpromazine, filipin, EIPA, and colchicine. BALB/c nude mice bearing PC12-SSTR2 xenografts received control, monotherapy, or combination with PRRT on Day 1, 4, or 7 of anlotinib. Results: Combination increased apoptosis versus monotherapy (both p < 0.0001). At Day 14, sequential combination groups had lower tumor volumes than control (both p < 0.001). In the primary six-group analysis, Day 4 versus Day 1 was only borderline significant (p = 0.0498), whereas Day 7 versus Day 1 was not significant (p = 0.1389). Anlotinib increased the relative internalized fraction of cell-associated 177Lu-DOTA-TOC from 20.63 ± 2.41% to 43.90 ± 3.57% (p < 0.0001). Chlorpromazine reduced this fraction in anlotinib-pretreated and untreated cells, consistent with clathrin-sensitive internalization. Conclusions: Anlotinib plus 177Lu-DOTA-TOC enhanced antitumor activity in this model. Cytoskeletal remodeling and increased clathrin-sensitive internalization suggest a testable mechanism; however, causality has not been established, and absolute intracellular activity, retention, optimal sequencing, and dosimetry require further study.