RESUMO
BACKGROUND: GTPase-activating proteins (GAPs) with a TBC (Tre-2/Bub2/Cdc16) domain architecture serve as negative regulators of Rab GTPases. The related crystal structure has been studied and reported by other members of our research group in 2017 (Chen et al. in Protein Sci 26(4):834-846, 2017). The protein crystal structure and sequencing data accession numbers in Protein structure database (PDB) are 5TUB (Shark TBC1D15 GAP) and 5TUC (Sus TBC1D15 GAP), respectively. In this paper, we analyzed the Rab-GAP specificity of TBC1D15 in the evolution and influence of key amino acid residue mutations on Rab-GAP activity. RESULTS: Sequence alignment showed that five arginine residues of the TBC1D15-GAP domain are conserved among the species Sus/Mus/Homo but have been replaced by glycine or lysine in Shark. A fragment activity assay was conducted by altering the five residues of Shark TBC1D15-GAP to arginine, and the corresponding arginine in TBC1D15 GAP domains from Sus and Homo species were mutated to resemble Shark TBC1D15-GAP. Our data revealed that the residues of G28, K45, K119, K122 and K221 in the Shark TBC1D15-GAP domain had a key role in determining the specificity for Rab7 and Rab11. Mutation of the five residues significantly altered the Shark TBC1D15-GAP activity. CONCLUSIONS: These results revealed that the substrate specificity of TBC1D15 has had different mechanisms across the evolution of species from lower-cartilaginous fish to higher mammals. Collectively, the data support a different mechanism of Shark TBC1D15-GAP in substrate selection, which provides a new idea for the development of Marine drugs.
Assuntos
Sequência Conservada , Evolução Molecular , Proteínas Ativadoras de GTPase/química , Tubarões/metabolismo , Proteínas rab de Ligação ao GTP/química , Sequência de Aminoácidos , Animais , Arginina/química , Arginina/genética , Cristalografia por Raios X , Glicina/química , Glicina/genética , Humanos , Lisina/química , Lisina/genética , Camundongos , Mutação , Domínios Proteicos , Alinhamento de Sequência , Tubarões/genética , Especificidade por Substrato , Suínos , Proteínas rab de Ligação ao GTP/genéticaRESUMO
BACKGROUND: The efficacy of adding panitumumab to chemotherapy remains controversial in the treatment of metastatic colorectal cancer (mCRC). Thus, we conducted this meta-analysis to assess the efficacy and safety of this combination regimen in patients with mCRC. METHODS: The PubMed, Embase, and Web of Science databases were comprehensively searched. Eligible studies included randomized controlled trials (RCTs) that estimated the efficacy of panitumumab with or without chemotherapy in the treatment of patients with mCRC. Hazard ratio (HR), risk ratio (RR), and 95% confidence intervals (CIs) were calculated, and heterogeneity was tested using I (2) statistics. RESULTS: Four studies involving a total of 3,066 patients were included in this meta-analysis. The addition of panitumumab to chemotherapy significantly improved progression-free survival (PFS) (HR =0.84, 95% CI =0.78-0.91, P=0.000) and the objective response rate (ORR) (RR =2.18, 95% CI =1.13-4.22, P=0.021) compared to chemotherapy alone, but no effect was noted on overall survival (OS) (HR =0.97, 95% CI =0.89-1.05, P=0.402). Subgroup analysis based on KRAS gene status revealed that the combined therapy significantly improved PFS (HR =0.71, 95% CI =0.57-0.88, P=0.002) and ORR (RR =2.43, 95% CI =1.21-4.90, P=0.013) in patients with wild-type KRAS tumors. Irinotecan-based chemotherapy plus panitumumab significantly prolonged PFS in patients with mCRC (HR =0.84, 95% CI =0.76-0.94, P=0.002). The combined treatment also increased the incidence of grade 3/4 adverse events. CONCLUSION: This meta-analysis indicates that the combination of panitumumab and chemotherapy effectively improved PFS and ORR, but it did not prolong OS. However, as the number of studies in the meta-analysis was limited, more large-scale, better-designed RCTs are needed to assess the combination of panitumumab and chemotherapy.
