Co-delivery of doxorubicin and recombinant plasmid pHSP70-Plk1-shRNA by bacterial magnetosomes for osteosarcoma therapy.
To explore a novel combination of chemotherapy, gene therapy, and thermotherapy for osteosarcoma, a targeted heat-sensitive co-delivery system based on bacterial magnetosomes (BMs) was developed. The optimal culture conditions of magnetotactic bacteria (MTB) AMB-1 and characterization of BMs were achieved. A recombinant eukaryotic plasmid heat shock protein 70-polo-like kinase 1-short hairpin RNA (pHSP70-Plk1-shRNA) under transcriptional control of a thermosensitive promoter (human HSP70 promoter) was constructed for gene therapy. Doxorubicin (DOX) and pHSP70-Plk1-shRNA were included in the targeted thermosensitive co-delivery system, and in vitro DOX release activity, targeted gene silencing efficiency and in vitro antitumor efficacy were investigated. The results showed that the optimal culture conditions of MTB AMB-1 are an oxygen concentration of 4.0%, a pH value of 7.0, 20 μmol/L of ferrous sulfate, 800 mg/L of sodium nitrate, and 200 mg/L of succinic acid. The temperature of BMs reached 43°C within 3 minutes and could be maintained for 30 minutes by adjusting the magnitude of the alternating magnetic field (AMF). The diameters of BMs, BM-DOX, BM-recombinant eukaryotic plasmid pHSP70-Plk1-shRNA (shPlk1), and BM-DOX-shPlk1 were 43.7±4.6, 79.2±5.4, 88.9±7.8, and 133.5±11.4 nm, respectively. The zeta potentials of BMs, BM-DOX, BM-shPlk1, and BM-DOX-shPlk1 were -29.4±6.9, -9.5±5.6, -16.7±4.8, and -10.3±3.1 mV, respectively. Besides, the system exhibited good release behavior. DOX release rate from BM-DOX-shPlk1 was 54% after incubation with phosphate-buffered saline at 43°C and 37% after incubation with 50% fetal bovine serum, which was significantly higher than that at 37°C (P<0.05). In addition, the expressions of Plk1 mRNA and protein were significantly suppressed in cells treated with BM-DOX-shPlk1 following hyperthermia treatment under the influence of an AMF compared to other groups (P<0.05). Furthermore, evaluation of the effect of in vitro antitumor revealed that BM-DOX-shPlk1 following hyperthermia treatment under the influence of an AMF was significantly more effective than others in tumor inhibition. In conclusion, the new heat-sensitive co-delivery system represents a promising approach for the treatment of cancer.
Department of Orthopaedics, The Second Affiliated Hospital of Anhui Medical University, Anhui Medical University, Hefei, Anhui, People's Republic of China.
Li
L
Cheng
Department of Orthopaedics, The Second Affiliated Hospital of Anhui Medical University, Anhui Medical University, Hefei, Anhui, People's Republic of China.
Youqun
Y
Ke
Department of Orthopaedics, The Second Affiliated Hospital of Anhui Medical University, Anhui Medical University, Hefei, Anhui, People's Republic of China.
Shuisheng
S
Yu
Department of Orthopaedics, The Second Affiliated Hospital of Anhui Medical University, Anhui Medical University, Hefei, Anhui, People's Republic of China.
Juehua
J
Jing
D017209:Apoptosis; D001419:Bacteria / Q000378:metabolism; D001859:Bone Neoplasms / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000008:administration & dosage / Q000378:metabolism; D045744:Cell Line, Tumor / Q000187:drug effects; D049109:Cell Proliferation; D004317:Doxorubicin / Q000008:administration & dosage*; D005434:Flow Cytometry; D020868:Gene Silencing / Q000187:drug effects; D015316:Genetic Therapy / Q000379:methods*; D050884:HSP72 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D006979:Hyperthermia, Induced / Q000379:methods; D055944:Magnetosomes / Q000737:chemistry*; D012516:Osteosarcoma / Q000378:metabolism*; D010100:Oxygen / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000008:administration & dosage / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000008:administration & dosage / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature; D000097869:Polo-Like Kinase 1
D018797:Cell Cycle Proteins; D004317:Doxorubicin; D050884:HSP72 Heat-Shock Proteins; D010100:Oxygen; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D000097869:Polo-Like Kinase 1
alternating magnetic field; chemotherapy; gene therapy; heat-inducible gene expression; magnetotactic bacteria; thermotherapy