Cervical cancer with transferrin receptor has a poor prognosis and is associated with immune infiltration, according to a comprehensive bioinformatics study

Background:The molecular mechanism underlying the involvement of the Transferrin receptor(TFRC)in cervical cancer remains poorly understood.This study aims to elucidate the role of TFRC in cervical cancer by analyzing data from The Cancer Genome Atlas(TCGA)and Genotype-Tissue Expression(GTEx)databases.Methods:TFRC protein expression was obtained from Human Protein Altas(HPA).All datas were collected from TCGA and GTEx.In this study,we analyzed the expression of TFRC in cervical cancer and its clinical significance.Through Kyoto Encyclopedia of Genes and Genomes(KEGG)and Gene set enrichment analyses(GSEA),investigated the related molecular pathways of TFRC.The relationship between TFRC and immune infiltration was then examined.The prognosis of different immune cell subsets was then analyzed after dividing cervical cancer patients into high and low expression of TFRC groups.Results:TFRC is highly expressed in various tumor tissues compared to control normal tissues,including cervical cancer.An increased expression of TFRC was associated with higher Tumor(T)and Node(N)stage,as well as a higher clinical stage.Kaplan–Meier(KM)survival analysis investigated that higher TFRC expression patients have a poor overall survival(OS),disease specific survival(DSS)and progress free interval(PFI).Both KEGG and GSEA enriched signaling pathway by high TFRC and low TFRC groups.There was a significant negative linear correlation between TFRC expression and immune infiltration.TFRC affects the prognosis of cervical cancer patients through immune pathway.Conclusions:Cervical cancer patients with TFRC expression may have a worse prognosis.

Construction of single chain Fv antibody against transferrin receptor and its protein fusion with alkaline phosphatase

AIM: To construct fusion protein of a single-chain antibody (scFv) against transferrin receptor (TfR) with alkaline phosphatase(AP). METHODS: The VH-linker-VL,namely scFv gene,was prepared by amplifying the VH and VL genes from plasmid pGEM-T-VH and pGEM-T-VL with splicing overlap extension polymerase chain reaction (SOE PCR). After the ScFv gene was modified by 5/71 and Not I,it was subcloned into the secretory expression vector pUC19/119, and then was transformed into E.coli TG1.The positive colonies were screened by colony PCR and their expressions were induced by IPTG.ScFv gene was gained by digesting ScFv expression vector pUC19/119 with 5/71 and NotI restriction enzymes, then subcloned into expression vector pDAP2, followed by transformation in E.coli TG1.The positive colonies were selected by bacterial colony PCR.The expression of fusion protein (scFv-AP) was induced by IPTG.Its activity was detected by enzyme immunoassay. The molecular weights of scFv and scFv-AP were measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). RESULTS: The product of SOE PCR formed a band of 700 bp in agarose gel electrophoresis. SDS-PAGE demonstrated the molecular weight of scFv was 27 ku.Immunofluorescent assay (IFA) demonstrated its reactivity with TfR.The molecular weight of scFv-AP was 75 ku.Enzyme immunoassay showed that scFv-AP could specifically bind to human TfR and play AP activity. CONCLUSION: We have successfully prepared the anti-human TfR scFv and constructed the fusion protein of scFv and AP.It is promising for immunological experiments.

Transferrin receptor and ferritin-H are developmentally regulated in oligodendrocyte lineage cells

Iron is an essential trophic element that is required for cell viability and differentiation, especially in oligodendrocytes, which consume relatively high rates of energy to produce myelin. Multiple iron metabolism proteins are expressed in the brain including transferrin receptor and ferritin-H. However, it is still unknown whether they are developmentally regulated in oligodendrocyte lineage cells for myelination. Here, using an in vitro cultured differentiation model of oligodendrocytes, we found that both transferrin receptor and ferritin-H are significantly upregulated during oligodendrocyte maturation, implying the essential role of iron in the development of oligodendrocytes. Additional different doses of Fe3＋ in the cultured medium did not affect oligodendrocyte precursor cell maturation or ferritin-H expression but decreased the expression of the transferrin receptor. These results indicate that upregulation of both transferrin receptor and ferritin-H contributes to maturation and myelination of oligodendrocyte precursor cells.

Study on The Method of Quantitative Analysis of Serum Ferritin and Soluble Transferrin Receptor with Protein Microarray Technology

Objective To establish and evaluate a protein serum ferritin （SF） and soluble transferrin receptor microarray method for combined measurement of （sTfR）. Methods Microarrayer was used to print both anti-SF antibodies I and anti-sTfR antibodies I on each protein microarray. Anti-SF antibodies II and anti-sTfR antibodies II were used as detection antibodies and goat antibodies coupled to Cy3 were used as antibodies Ill. The detection conditions of the quantitative analysis method for simultaneous measurement of SF and sTfR with protein microarray were optimized and evaluated. The protein microarray was compared with commercially available traditional tests with 26 serum samples. Results By comparison experiment, mouse monoclonal antibodies were chosen as the probes and contact printing was chosen as the printing method. The concentrations of SF and sTfR probes were 0.5 mg/mL and 0.5 mg/mL respectively, while those of SF and sTfR detection antibodies were 5 μg/mL and 0.36 μg/mL respectively. Intra- and inter-assay variability was between 3.26% and 18.38% for all tests. The regression coefficients comparing protein microarray with traditional test assays were better than 0.81 for SF and sTfR. Conclusion The present study has established a protein microarray method for combined measurement of SF and sTfR.

Preparation and Identification of scFv and bsFv against Transferrin Receptor

To obtain single chain variable fragment （scFv） and bivalent single chain variable fragment （bsFv） against transferrin receptor, up-stream and down-stream primers were designed according to the complementary sequences of FR1 region of variable heavy （VH） and FR4 of variable light （VL）, respectively, which contained inter-linker G4S and the restriction endonuclease SfiI, AscI and NotI. Two pieces of scFv fragments were first amplified through PCR and then inserted into plasmid pAB1, which could express scFv protein once induced by IPTG in the host bacteria. To express scFv and bsFv, E. coli TG1 was cultured in LB broth and was induced by IPTG. The restriction enzyme digestion map and DNA sequencing demonstrated that scFv and bsFv genes were successfully inserted into the expression plasmid. SDS-PAGE and Western blotting revealed the protein band at 35kD and 60kD, which were consistent with the molecular weight of scFv and bsFv respectively. Flow cytometry showed that scFv and bsFv harbored the specific binding activity with TfR expressed in various tumor cells, and the avidity of bsFv was higher than that of the parent scFv.

Selection for Anti-transferrin Receptor Bispecific T-cell Engager in Different Molecular Formats

Selecting an ideal molecular format from diverse structures is a major challenge in developing a bispecific antibody(BsAb).To choose an ideal format of anti-CD3 x anti-transferrin receptor(TfR)bispecific antibodies for clinical application,we constructed TfR bispecific T-cell engager(BiTE)in two extensively applied formats,including single-chain tandem singlechain variable fragments(scFvs)and double-chain diabodies,and evaluated their functional characterizations in vitro.Results demonstrated that TfR-BiTE in both formats directed potent killing of TfR+HepG2 cells.However,compared to two・chain diabodies,scFvs were more efficient in antigen binding and TfR target killing.Furthermore,different domain orders in scFvs would also be evaluated because single-TfR-CD3-His was preferable to single-CD3-TfR-His in immunotherapeutic strategies.Thus,the single-chain tandem TfR-CD3 format was favored for further investigation in cancer therapy.

Effect of high flux hemodialysis on renal anemia and soluble transferrin receptor in hemodialysis patients

Objective: To investigate the effect of high throughput hemodialysis on soluble transferrin receptor in hemodialysis patients and the improvement of renal anemia. Methods: 132 patients receiving maintenance hemodialysis in our hospital from July 2017 to July 2019 were selected and divided into control group and observation group according to the random number table method, with 66 cases each. The observation group was treated with high-flux hemodialysis, while the control group was treated with low-flux hemodialysis for 6 months. Compare two groups before and after treatment serum beta 2 microglobulin (beta 2 - MG), serum creatinine (Scr), blood urea nitrogen (BUN) level, anemia related index [red blood cells deposited (HCT), hemoglobin (Hb), reticulocyte percentage (Ret%)], iron metabolism index [serum ferritin (SF), transferrin saturation (TSAT)、Hepcidin(Hepc)], soluble transferrin receptor (sTfR) levels and adverse reactions. Results: the levels of 2-MG, Scr and BUN in the two groups before treatment were compared (P>0.05). After treatment, Scr and BUN levels in the two groups were significantly decreased (P<0.05), but were compared between the two groups (P>0.05). The level of 2-MG in the observation group was lower than that in the control group (P<0.05). Before treatment, sTfR, Hb, HCT level and Ret% of the two groups were compared(P>0.05). After treatment, Hb and HCT levels in the observation group were higher than those in the control group, while Ret% were lower than those in the control group, (P<0.05). Before treatment, the levels of ST、TAST、sTfR and Hepc in the two groups were compared (P>0.05). After treatment, the level of ST and TAST in the observation group was higher than that in the control group, The levels of sTfR and Hepc were lower than the control group (P<0.05). The overall incidence of adverse reactions in the observation group (8.93%) was lower than that in the control group (10.14%), with no significant difference (P>0.05). Conclusion: The high-throughput hemodialysis department significantly improved renal anemia in hemodialysis patients, reduced serum sTfR level, and had fewer adverse reactions and higher safety.

DL-3-n-butylphthalide alleviates motor disturbance by suppressing ferroptosis in a rat model of Parkinson’s disease

DL-3-n-butylphthalide(NBP)-a compound isolated from Apium graveolens seeds-is protective against brain ischemia via various mechanisms in humans and has been approved for treatment of acute ischemic stroke.NBP has shown recent potential as a treatment for Parkinson’s disease.However,the underlying mechanism of action of NBP remains poorly understood.In this study,we established a rat model of Parkinson’s disease by intraperitoneal injection of rotenone for 28 successive days,followed by intragastric injection of NBP for 14-28 days.We found that NBP greatly alleviated rotenone-induced motor disturbance in the rat model of Parkinson’s disease,inhibited loss of dopaminergic neurons and aggregation ofα-synuclein,and reduced iron deposition in the substantia nigra and iron content in serum.These changes were achieved by alterations in the expression of the iron metabolism-related proteins transferrin receptor,ferritin light chain,and transferrin 1.NBP also inhibited oxidative stress in the substantia nigra and protected mitochondria in the rat model of Parkinson’s disease.Our findings suggest that NBP alleviates motor disturbance by inhibition of iron deposition,oxidative stress,and ferroptosis in the substantia nigra.

TfR1 Extensively Regulates the Expression of Genes Associated with Ion Transport and Immunity

Transferrin receptor 1(TfR1),encoded by the TFRC gene,is the gatekeeper of cellular iron uptake for cells.A variety of molecular mechanisms are at work to tightly regulate TfR1 expression,and abnormal TfR1 expression has been associated with various diseases.In the current study,to determine the regulation pattern of TfR1,we cloned and overexpressed the human TFRC gene in HeLa cells.RNA-sequencing(RNA-seq)was used to analyze the global transcript levels in overexpressed(OE)and normal control(NC)samples.A total of 1669 differentially expressed genes(DEGs)were identified between OE and NC.Gene ontology(GO)analysis was carried out to explore the functions of the DEGs.It was found that multiple DEGs were associated with ion transport and immunity.Moreover,the regulatory network was constructed on basis of DEGs associated with ion transport and immunity,highlighting that TFRC was the node gene of the network.These results together suggested that precisely controlled TfR1 expression might be not only essential for iron homeostasis,but also globally important for cell physiology,including ion transport and immunity.

Non-HFE haemochromatosis

Non-HFE hereditary haemochromatosis （HH） refers to a genetically heterogeneous group of iron overload disorders that are unlinked to mutations in the HFE gene. The four main types of non-HFE HH are caused by mutations in the hemojuvelin, hepcidin, transferrin receptor 2 and ferroportin genes. Juvenile haemochromatosis is an autosomal recessive disorder and can be caused by mutations in either hemojuvelin or hepcidin. Ar~ adult onset form of HH similar to HFE-HH is caused by homozygosity for mutations in transferrin receptor 2. The autosomal dominant iron overload disorder ferroportin disease is caused by mutations in the iron exporter ferroportin. The clinical characteristics and molecular basis of the various types of non-HFE haemochromatosis are reviewed. The study of these disorders and the molecules involved has been invaluable in improving our understanding of the mechanisms involved in the regulation of iron metabolism.

Construction and Identification of the Adenoviral Vector with Dual Reporter Gene for Multimodality Molecular Imaging

Summary： In this study, the recombinant adenovirus （Ad） vector containing dual reporter gene [i.e. human transferrin receptor gene （TFRC） and firefly luciferase reporter gene] was constructed to provide a novel experimental tool for magnetic resonance （MR） and bioluminescence dual-modality molecular imaging. The cDNA of TFRC was amplified by polymerase chain reaction （PCR） and cloned into the multiple cloning site of pShuttle-CMV-CMV-Luciferase vector. After identification by Sfi I digestion and sequencing, pShuttle-TFRC-Luciferase vector and the adenoviral backbone vector （pAdeno） were subjected to homologous recombination. The correct recombinant plasmid was then transfected into 293 packaging cells to produce adenoviral particles and confirmed by PCR. After infection of human colo- rectal cancer LOVO cells with Ad-TFRC-Luciferase, the expressions of transferrin receptor （TfR） and luciferase protein were detected respectively by Western blotting and bioluminescence imaging in vitro. The results showed that TFRC gene was successfully inserted into the adenoviral shuttle vector carrying luciferase gene. DNA sequence analysis indicated that the TFRC gene sequence in the shuttle plasmid was exactly the same as that reported in GenBank. The recombinant plasmid was identified correct by restriction digestion. Ad-TFRC-Luciferase recombinant adenovirns was constructed successfully, and the virus titer was 1.6x10^10 pfu/mL. Forty-eight h after dual reporter gene transfection, the expressions of TfR and luciferase protein were increased significantly （P〈0.01）. It was concluded that the recombinant adenovirus vector with dual reporter gene was successfully established, which may be used for in vivo tracing target cells in multimodality imaging.

Upregulation of CDGSH iron sulfur domain 2 attenuates cerebral ischemia/reperfusion injury

CDGSH iron sulfur domain 2 can inhibit ferroptosis,which has been associated with cerebral ischemia/reperfusion,in individuals with head and neck cancer.Therefore,CDGSH iron sulfur domain 2 may be implicated in cerebral ischemia/reperfusion injury.To validate this hypothesis in the present study,we established mouse models of occlusion of the middle cerebral artery and HT22 cell models of oxygen-glucose deprivation and reoxygenation to mimic cerebral ischemia/reperfusion injury in vivo and in vitro,respectively.We found remarkably decreased CDGSH iron sulfur domain 2 expression in the mouse brain tissue and HT22 cells.When we used adeno-associated virus and plasmid to up-regulate CDGSH iron sulfur domain 2 expression in the brain tissue and HT22 cell models separately,mouse neurological dysfunction was greatly improved;the cerebral infarct volume was reduced;the survival rate of HT22 cells was increased;HT22 cell injury was alleviated;the expression of ferroptosis-related glutathione peroxidase 4,cystine-glutamate antiporter,and glutathione was increased;the levels of malondialdehyde,iron ions,and the expression of transferrin receptor 1 were decreased;and the expression of nuclear-factor E2-related factor 2/heme oxygenase 1 was increased.Inhibition of CDGSH iron sulfur domain 2 upregulation via the nuclear-factor E2-related factor 2 inhibitor ML385 in oxygen-glucose deprived and reoxygenated HT22 cells blocked the neuroprotective effects of CDGSH iron sulfur domain 2 up-regulation and the activation of the nuclear-factor E2-related factor 2/heme oxygenase 1 pathway.Our data indicate that the up-regulation of CDGSH iron sulfur domain 2 can attenuate cerebral ischemia/reperfusion injury,thus providing theoretical support from the perspectives of cytology and experimental zoology for the use of this protein as a therapeutic target in patients with cerebral ischemia/reperfusion injury.

Eph receptor A4 regulates motor neuron ferroptosis in spinal cord ischemia/reperfusion injury in rats

Previous studies have shown that the receptor tyrosine kinase Eph receptor A4(EphA4) is abundantly expressed in the nervous system. The EphA4 signaling pathway plays an important role in regulating motor neuron ferroptosis in motor neuron disease. To investigate whether EphA4 signaling is involved in ferroptosis in spinal cord ischemia/reperfusion injury, in this study we established a rat model of spinal cord ischemia/reperfusion injury by clamping the left carotid artery and the left subclavian artery. We found that spinal cord ischemia/reperfusion injury increased EphA4 expression in the neurons of anterior horn, markedly worsened ferroptosis-related indicators, substantially increased the number of mitochondria exhibiting features consistent with ferroptosis, promoted deterioration of motor nerve function, increased the permeability of the blood-spinal cord barrier, and increased the rate of motor neuron death. Inhibition of EphA4 largely rescued these effects. However, intrathecal administration of the ferroptosis inducer Erastin counteracted the beneficial effects conferred by treatment with the EphA4 inhibitor. Mass spectrometry and a PubMed search were performed to identify proteins that interact with EphA4, with the most notable being Beclin1 and Erk1/2. Our results showed that inhibition of EphA4 expression reduced binding to Beclin1, markedly reduced p-Beclin1, and reduced Beclin1-XCT complex formation. Inhibition of EphA4 also reduced binding to p-Erk1/2 and markedly decreased the expression of c-Myc, transferrin receptor 1, and p-Erk1/2. Additionally, we observed co-localization of EphA4 and p-Beclin1 and of EphA4 and p-ERK1/2 in neurons in the anterior horn. In conclusion, EphA4 participates in regulating ferroptosis of spinal motor neurons in the anterior horn in spinal cord ischemia/reperfusion injury by promoting formation of the Beclin1-XCT complex and activating the Erk1/2/c-Myc/transferrin receptor 1 axis.

Occult celiac disease prevents penetrance of hemochromatosis

AIM:To report a patient with C282Y homozygocity,depleted body iron and intestinal atrophy caused by celiac disease (CD) who experienced resolution of the enteropathy with subsequent normalization of iron metabolism upon gluten free diet. METHODS:To obtain information on the tissue distribution and quantitative expression of proteins involved in duodenal iron trafficking,we determined the expression of divalent-metal transporter 1 (DMT1),ferroportin 1 (FP1) and transferrin receptor (TfR1) by means of immunohist-ochemistry and real-time PCR in duodenal biopsies of this patient. RESULTS:Whereas in hereditary hemochromatosis patients without CD, DMT1 expression was up-regulated leading to excessive uptake of iron, we identified a significant reduction in protein ana mRNA expression of DMT1 as a compensatory mechanism in this patient with HH and CD. CONCLUSION:Occult CD may compensate for increased DMT1 expression in a specific subset of individuals with homozygous C282Y mutations in the hemochromatosis (HFE) gene,thus contributing to the low penetrance of HH.

Transferrin receptor-targeted immunostimulant for photodynamic immunotherapy against metastatic tumors through β-catenin/CREB interruption

The immunosuppressive phenotype of tumor cells extensively attenuates the immune activation effects of traditional treatments.In this work,a transferrin receptor(TfR)targeted immunostimulant(PTI)is fabricated for photodynamic immunotherapy against metastatic tumors by interrupting β-catenin signal pathway.To synthesize PTI,the photosensitizer conjugated TfR targeting peptide moiety(Palmitic-K(PpIX)-HAIYPRH)is unitized to encapsulate the transcription interrupter of ICG-001.On the one hand,the recognition of PTI and TfR can promote drug delivery into tumor cells to destruct primary tumors through photodynamic therapy and initiate an immunogenic cell death with the release of tumorassociated antigens.On the other hand,PTI will interrupt the binding between b-catenin andcAMP response element-binding protein(CREB),regulating the gene transcription to downregulate programmed death ligand 1(PD-L1)while upregulating CeC motif chemokine ligand 4(CCL4).Furthermore,the elevated CCL4 can recruit the dendritic cells to present tumor-specific antigens and promote T cells activation and infiltration,and the downregulated PD-L1 can avoid the immune evasion of tumor cells and activate systemic anti-tumor immunity to eradicate lung metastasis.This work may inspire the development of antibody antibody-free strategy to activate systemic immune response in consideration of immunosuppressive conditions.

Drug delivery via the transferrin receptor-mediated endocytosis pathway

The membrane transferrin receptor-mediated endocytosis has been exploited for developing novel targeted drug delivery systems, which could have a variety of applications in the site-specific delivery of anticancer drugs, proteins and therapeutic genes into proliferating malignant cells that overexpress the transferrin receptors. This is achieved by coupling transferrin or monoclonal antibody to transferrin receptor with therapeutic drugs or drug delivery vesicles. The transferrin conjugates can be obtained by use of either bifunctional chemical linkers or by genetic infusion of therapeutic peptides/proteins into the structure of transferrin / and monoclonal antibody to transferrin receptor. A variety of drug carriers such as liposomes, nanoparticles and DNApolymer complexes （i.e. polyplex and lipoplex） were used to efficiently deliver the Wansferrin conjugates. Use of transferrin conjugates results in improvement in drug efficacy, selectivity and drug release as well as reduction in drug toxicity. This paper reviews the basic biochemistry of transferrin and the transferrin receptor as well as the strategy for developing targeted drug delivery system.

Transferrin receptor and Fc α/μ receptor may not be the major IgA_1 receptor on human mesangial cells

IgA nephropathy (IgAN) is the most common form of primary glomerulonephritis. 1 The histopathology of IgAN is characterized by abundance of mesangial matrix and proliferation of mesangial cells. IgA_1 deposition in the mesangium plays an important role in the inflammatory process in this disease.

Ultraviolet-B induced expression of hypoxia-inducible factor 1α, transferrin receptor through EGFR/PI3K/AKT/DEC1 pathway

The aim of this research was to explore the effects and signaling pathway of ultraviolet-B(UVB)irradiation on the expression of hypoxia-inducible factor 1a(HIF-1α)and transferrin receptor(TfR).HIF-1α protein was measured by Western blot method.Expressions of epidermal growth factor receptor(EGFR),phosphor-EGF-R and TfR after UVB irradiation were determined with flow cytometry.After UVB irradiation,mRNA levels of HIF-1α and TfR were detected by real time-PCR.Results showed that compared with control groups,UVB was able to induce HIF1α and TfR protein expression in a dose-and time-dependent manner in HaCat cells(P<0.05).TfR mRNA was expressed in a dose-dependent manner and reached a peak at the 8th hour in HaCat cells(P<0.05)whereas HIF-1α mRNA expression was not affected by UVB treatment(P>0.05).The EGFR/PI3K/AKT signaling pathway was required for the induction of HIF-1α and TfR expression induced by UVB.UVB induced activation of EGFR in HaCat cells and EGFR regulated expression of TfR and HIF-1α.EGFR(−/−)MEF did not increase the HIF1 expression following UVB irradiation(P>0.05).In contrast,EGFR(+/+)MEF strongly enhanced HIF1a expression after UVB irradiation(P<0.05).PD153035,a selective inhibitor of EGFR tyrosine kinase,inhibited the TfR protein expression in UVB-treated cells in a dose-dependent manner(P<0.05).PI3K inhibitors,LY294002 and wortmannin,inhibited HIF-1a and TfR expressions induced by UVB(P<0.05).The DEC1(−/−)Ha-Cat cells did not increase their TfR and HIF-1α expressions following UVB irradiation(P>0.05).In contrast,DEC1(+/+)HaCat cells strongly enhanced TfR and HIF-1α protein expression after UVB irradiation(P<0.05).We conclude that UVB induces TfR and HIF-1α expressions via EGFR/PI3K/AKT/DEC1 signaling pathway.

Imaging Fast Cellular Uptake of Polymer Dots via Receptor‑Mediated Endocytosis

During the past decade,semiconducting polymer dots(Pdots)have been prevailing in the family of fluorescent probes due to its high photon budget,excellent photo stability,and good biocompatibility.In this study,holo-Transferrin human(Tf)was utilized to covalently couple with Pdots for a highly efficient endocytosis process through transferrin receptors(TfRs)mediated internalization.As a result,the endocytosis efficiency of Tf-conjugated Pdots in HeLa cells dramatically increased as compared to that of unconjugated Pdots in the same condition.This acute increment demonstrates that holo-Transferrin molecules are of great capability for intracellular delivery of Pdots to TfRs overexpressed cells.The transportation route of Tf-conjugated Pdots is quite different from the uptake mechanism of unconjugated Pdots via nonspecific endocytic trafficking pathway.Considering the overexpression of TfRs in various cancer cells,Tf-conjugated Pdots hold potential to function as a nanocarrier for efficient drug delivery in cancer diagnostics and therapy.

Brain endothelial cell-derived extracellular vesicles with a mitochondria-targeting photosensitizer effectively treat glioblastoma by hijacking the blood-brain barrier

Glioblastoma(GBM)is the most aggressive malignant brain tumor and has a high mortality rate.Photodynamic therapy(PDT)has emerged as a promising approach for the treatment of malignant brain tumors.However,the use of PDT for the treatment of GBM has been limited by its low blood-brain barrier(BBB)permeability and lack of cancer-targeting ability.Herein,brain endothelial cell-derived extracellular vesicles(bEVs)were used as a biocompatible nanoplatform to transport photosensitizers into brain tumors across the BBB.To enhance PDT efficacy,the photosensitizer chlorin e6(Ce6)was linked to mitochondria-targeting triphenylphosphonium(TPP)and entrapped into bEVs.TPPconjugated Ce6(TPP-Ce6)selectively accumulated in the mitochondria,which rendered brain tumor cells more susceptible to reactive oxygen species-induced apoptosis under light irradiation.Moreover,the encapsulation of TPP-Ce6 into b EVs markedly improved the aqueous stability and cellular internalization of TPP-Ce6,leading to significantly enhanced PDT efficacy in U87MG GBM cells.An in vivo biodistribution study using orthotopic GBM-xenografted mice showed that b EVs containing TPP-Ce6[b EV(TPP-Ce6)]substantially accumulated in brain tumors after BBB penetration via transferrin receptor-mediated transcytosis.As such,b EV(TPP-Ce6)-mediated PDT considerably inhibited the growth of GBM without causing adverse systemic toxicity,suggesting that mitochondria are an effective target for photodynamic GBM therapy.