AS1411 aptamer/RGD dual functionalized theranostic chitosan-PLGA nanoparticles for brain cancer treatment and imaging.

Conventional chemotherapy and poor targeted delivery in brain cancer resulting to poor treatment and develop resistance to anticancer drugs. Meanwhile, it is quite challenging to diagnose/detection of brain tumor at early stage of cancer which resulting in severity of the disease. Despite extensive research, effective treatment with real-time imaging still remains completely unavailable, yet. In this study, two brain cancer cell specific moieties i.e., AS1411 aptamer and RGD are decorated on the surface of chitosan-PLGA nanoparticles to improve targeted co-delivery of docetaxel (DTX) and upconversion nanoparticles (UCNP) for effective brain tumor therapy and real-time imaging. The nanoparticles were developed by a slightly modified emulsion/solvent evaporation method. This investigation also translates the successful synthesis of TPGS-chitosan, TPGS-RGD and TPGS-AS1411 aptamer conjugates for making PLGA nanoparticle as a potential tool of the targeted co-delivery of DTX and UCNP to the brain cancer cells. The developed nanoparticles have shown an average particle size <200 nm, spherical in shape, high encapsulation of DTX and UCNP in the core of nanoparticles, and sustained release of DTX up to 72 h in phosphate buffer saline (pH 7.4). AS1411 aptamer and RGD functionalized theranostic chitosan-PLGA nanoparticles containing DTX and UCNP (DUCPN-RGD-AS1411) have achieved greater cellular uptake, 89-fold improved cytotoxicity, enhanced cancer cell arrest even at lower drug conc., improved bioavailability with higher mean residence time of DTX in systemic circulation and brain tissues. Moreover, DUCPN-RGD-AS1411 have greatly facilitated cellular internalization and higher accumulation of UCNP in brain tissues. Additionally, DUCPN-RGD-AS1411 demonstrated a significant suppression in tumor growth in brain-tumor bearing xenograft BALB/c nude mice with no impressive sign of toxicities. DUCPN-RGD-AS1411 has great potential to be utilized as an effective and safe theranostic tool for brain cancer and other life-threatening cancer therapies.

Nanoaggregate-forming lipid-conjugated AS1411 aptamer as a promising tumor-targeted delivery system of anticancer agents in vitro.

Nanoparticles offer targeted delivery of drugs with minimal toxicity to surrounding healthy tissue and have great potential in the management of human papillomavirus (HPV)-related diseases. We synthesized lipid-modified AS1411 aptamers capable of forming nanoaggregates in solution containing Mg2+. The nanoaggregates presented suitable properties for pharmaceutical applications such as small size (100 nm), negative charge, and drug release. The nanoaggregates were loaded with acridine orange derivative C8 for its specific delivery into cervical cancer cell lines and HPV-positive tissue biopsies. This improved inhibition of HeLa proliferation and cell uptake without significantly affecting healthy cells. Finally, the nanoaggregates were incorporated in a gel formulation with promising tissue retention properties aiming at developing a local delivery strategy of the nanoaggregates in the female genital tract. Collectively, these findings suggest that the nanoformulation protocol has great potential for the delivery of both anticancer and antiviral agents, becoming a novel modality for cervical cancer management.

Nucleic acid therapeutics: a focus on the development of aptamers.

INTRODUCTION: Aptamers provide exciting opportunities for the development of specific and targeted therapeutic approaches. AREAS COVERED: In this review, the authors discuss different therapeutic options available with nucleic acids, including aptamers, focussing on similarities and differences between them. The authors concentrate on case studies with specific aptamers, which exemplify their distinct advantages. The reasons for failure, wherever available, are deliberated upon. Attempts to accelerate the in vitro selection process have been discussed. Challenges with aptamers in terms of their specificity and targeted delivery and strategies to overcome these are described. Examples of precise regulation of systemic half-life of aptamers using antidotes are discussed. EXPERT OPINION: Despite their nontoxic nature, a variety of reasons limit the therapeutic potential of aptamers in the clinic. The analysis of adverse effects observed with the pegnivacogin/anivamersen pair has highlighted the need to screen for preexisting PEG antibodies in any clinical trial involving pegylated molecules. Surprisingly, and promisingly, the ability of nucleic acid therapeutics to breach the blood brain barrier seems achievable. The recognition of specific motifs, e.g. G-quadruplex in thrombin-binding aptamers, or a 'nucleation' zone while designing aptamer-antidote pairs, is likely to accelerate the discovery of therapeutically efficacious molecules.

Anti-epithelial cell adhesion molecule RNA aptamer-conjugated liposomal doxorubicin as an efficient targeted therapy in mice bearing colon carcinoma tumor model.

To overcome the lack of selectivity and nonspecific biodistribution of drugs in the body, targeted delivery of chemotherapeutic agents with aptamers is a very effective method. In this strategy, aptamers could be specifically identified and attach to targeted molecules on the cancerous cells and deliver the chemotherapeutic agents to desired tissue with minimal or no damage to the normal cells. In this study, we designed anti-epithelial cell adhesion molecule (EpCAM) RNA aptamer conjugated PEGylated liposomal doxorubicin (ER-lip) to investigate its in vitro and in vivo anticancer abilities. Data showed that EpCAM aptamer was able to enhance cell uptake and cytotoxic effects of Dox in C26 cell line. The biodistribution study indicated that ER-lip enhanced the tumor accumulation of Dox compared to Caelyx. Also, double staining of isolated tumor cells with anti-CD44-PE-cy5 and anti-EpCAM Cy-7 antibodies indicated that tumor cells expressed a high level of EpCAM+ CD44+ cells (p ≤ .001) compared to cultured C26 cell line. in vivo results showed that ER-lip promoted survival and reduced tumor growth rate in animal model compared to Caelyx. In conclusion, these results suggested that the ER-lip could be served as promising formulation for the treatment of cancers with the high expression of EpCAM.

An aptamer-based drug delivery agent (CD133-apt-Dox) selectively and effectively kills liver cancer stem-like cells.

Liver cancer has no effective therapies, hence a poor survival. Cancer stem-like cells not only contribute to cancer initiation and progression, but also to drug resistance, cancer metastasis, and eventually treatment failure. Hence, any approaches that can effectively kill cancer stem-like cells hold a great potential for cancer treatment. CD133 is a robust marker for liver cancer stem-like cells. We developed a specific aptamer against CD133 (CD133-apt), and then loaded this aptamer with an anticancer drug doxorubicin (CD133-apt-Dox). The efficacy of CD133-apt-Dox in targeting liver cancer stem-like cells and its overall effect in treating liver cancer were investigated using multiple in vitro and in vivo studies including in patients-derived liver cancer organoids. We have observed that CD133-apt could preferably delivered doxorubicin to CD133-expressing cells with efficient drug accumulation and retention. CD133-apt-Dox impaired the self-renewal capacity of liver cancer stem-like cells and attenuated their stem-ness phenotypes in vitro or in vivo. CD133-apt-Dox significantly inhibited the growth of liver cancer cells and patients-derived organoids and reduced the growth of xenograft tumours in nude mice inhibited the growth of DEN-induced liver cancer in immunocompetent mice. Hence, aptamer-mediated targeting of CD133 is a highly promising approach for liver cancer therapy.

Co-delivery of doxorubicin and aptamer against Forkhead box M1 using chitosan-gold nanoparticles coated with nucleolin aptamer for synergistic treatment of cancer cells.

Herein, a nanotherapeutic delivery method was presented for co-delivery of doxorubicin (DOX) and aptamer against Forkhead box M1 (FOXM1 Apt) to cancer cells. Firstly, the vehicle composed of chitosan (CS)-Gold nanoparticles (AuNPs) conjugate was prepared. Nucleolin aptamer (AS1411) and FOXM1 Apt were loaded onto the CS-AuNPs and formed Aptamers (Apts)-CS-AuNPs. Subsequently, DOX was added to the Apts-CS-AuNPs to obtain the DOX-Apts-CS-AuNPs complex for synergistic treatment of tumor. The data of flow cytometry analysis and fluorescence imaging displayed that the complex was effectively internalized into target cells (A549 and 4T1 cells, nucleolin+) but not into CHO cells as nontarget cells. The results of the MTT assay showed that the complex significantly increased cell mortality in 4T1 and A549 cells compared to CHO cells treated with the complex. The in vivo studies demonstrated that the DOX-Apts-CS-AuNPs complex exhibited more tumor inhibitory effect and less distribution in other organs compared to free DOX.

Aptamer-Conjugated Multifunctional Polymeric Nanoparticles as Cancer-Targeted, MRI-Ultrasensitive Drug Delivery Systems for Treatment of Castration-Resistant Prostate Cancer.

Nanoscopic therapeutic systems that incorporate therapeutic agents, molecular targeting, and imaging capabilities have gained momentum and exhibited significant therapeutic potential. In this study, multifunctional polymeric nanoparticles with controlled drug delivery, cancer-targeted capability, and efficient magnetic resonance imaging (MRI) contrast characteristics were formulated and applied in the treatment of castration-resistant prostate cancer (CRPC). The "core-shell" targeted nanoparticles (NPs) were synthesized by the self-assembly of a prefunctionalized amphiphilic triblock copolymer composed of poly(lactic-co-glycolic-acid) (PLGA), polyethylene glycol (PEG), and the Wy5a aptamer (Apt), which have been screened for targeting the CRPC cell line PC-3 by cell-SELEX technique as described in our previous study. Docetaxel (Dtxl) and a cluster of hydrophobic superparamagnetic iron oxide (SPIO) nanoparticles were simultaneously encapsulated into the targeted nanoparticles. The targeted NPs showed a controlled drug release and an increased contrast-enhanced MRI capability. The presence of Wy5a on the nanoparticle surface resulted in the cancer-targeted delivery to PC-3 cells in vitro and in vivo. In vitro MRI and cytotoxicity studies demonstrated the ultrasensitive MRI and increased cytotoxicity of these targeted NPs. In vivo studies revealed that the targeted NPs exhibited a more efficacious antitumor capability without significant systemic toxicity. Our data suggested that these targeted NPs may be a promising drug delivery system for the efficacious treatment of CRPC.

A photo-triggerable aptamer nanoswitch for spatiotemporal controllable siRNA delivery.

A photo-triggerable aptamer nanoswitch was proposed for spatiotemporal regulation of siRNA delivery. Recognition between AS1411 and nucleolin was effectively blocked by a photo-labile complementary oligonucleotide, which could be reactivated with photo-irradiation, resulting in efficient tumor-targeted siRNA internalization and gene silencing in vitro and in vivo.

Evaluation of the specific uptake of radiolabeled Staphylococcus aureus aptamers in the infectious foci.

The specific uptake of 99mTc radiolabeled Staphylococcus aureus aptamers in the infectious foci was evaluated by scintigraphic imaging of infection-bearing mice. The radiotracer uptake was inhibited by non-radiolabeled aptamers in a competition assay. In addition, when a different number of bacterial cells was used to infect mice an increase in the target/non-target ratios of images correlated with the increase of CFU per gram of tissue was verified. These results confirmed that 99mTc-aptamers were specific to bacterial focus and the level of uptake was dependent on the number of bacterial cells.

AS1411 aptamer modified carbon dots via polyethylenimine-assisted strategy for efficient targeted cancer cell imaging.

OBJECTIVES: Carbon dots (CDs), as a fascinating class of fluorescent carbon nanomaterials, have been proven to be powerful tools in the field of bioimaging and biosensing due to their small size, suitable photostability and favourable biocompatibility. However, the cellular uptake of free CDs lacks selectivity and the same negative charges as cell membranes may cause inefficient cell internalization. In this study, an efficient detecting and targeting nanosystem was developed based on the DNA aptamer AS1411 modified CDs with polyethyleneimine (PEI) as connecting bridge. MATERIALS AND METHODS: Hydrothermally prepared CDs were assembled with positive-charged PEI, followed by conjugation with AS1411 through electrostatic interaction to form CDs-PEI-AS1411 nanocomplexes. The CDs, CDs-PEI and CDs-PEI-AS1411 were characterized by transmission electron microscopy (TEM), fourier transform infrared (FTIR) spectra, UV-vis spectra, zeta potential measurements and capillary electrophoresis characterizations. The cytotoxicity investigation of the CDs-PEI-AS1411 and CDs-PEI in both MCF-7 and L929 cells was carried out by the CCK-8 assay. The cellular uptake of the CDs-PEI-AS1411 was studied with confocal microscopy and flow cytometry. RESULTS: The as-prepared nanosystem possessed good photostability and no obvious cytotoxicity. On the basis of the confocal laser scanning microscope observation and the flow cytometry studies, the cellular uptake of CDs-PEI-AS1411 nanosystem in MCF-7 cells was significantly higher than that of L929 cells, which revealed the highly selective detection ability of nucleolin-positive cells. CONCLUSIONS: The results of this study indicated that the CDs-PEI-AS1411 nanosystem had a potential value in cancer cell targeted imaging.

Short-Acting Anti-VWF (von Willebrand Factor) Aptamer Improves the Recovery, Survival, and Hemostatic Functions of Refrigerated Platelets.

OBJECTIVE: Refrigeration-induced binding of VWF (von Willebrand factor) to platelets contributes to the rapid clearance of refrigerated platelets. In this study, we investigate whether inhibiting VWF binding by a DNA-based aptamer ameliorates the clearance of refrigerated platelets without significantly impeding hemostatic functions. Approach and Results: Platelets were refrigerated with or without aptamer ARC1779 for 48 hours. VWF binding, the effective lifetime of ARC1779, platelet post-transfusion recovery and survival, and the hemostatic function were measured. ARC1779 treatment during refrigeration inhibited the platelet-VWF interaction. ARC1779-treated refrigerated murine platelets exhibited increased post-transfusion recovery and survival than untreated ones (recovery of ARC1779-treated platelets: 76.7±5.5%; untreated: 63.7±0.8%; P<0.01. Half-life: 31.4±2.36 hours versus 28.1±0.86 hours; P<0.05). A similar increase was observed for refrigerated human platelets (recovery: 49.4±4.4% versus 36.8±2.1%, P<0.01; half-life: 9.2±1.5 hours versus 8.7±0.9 hours, ns). The effective lifetime of ARC1779 in mice was 2 hours. Additionally, ARC1779 improved the long-term (2 hours after transfusion) hemostatic function of refrigerated platelets (tail bleeding time of mice transfused with ARC1779-treated refrigerated platelets: 160±65 seconds; untreated: 373±96 seconds; P<0.01). The addition of an ARC1779 antidote before transfusion improved the immediate (15 minutes after transfusion) hemostatic function (bleeding time of treated platelets: 149±21 seconds; untreated: 320±36 seconds; P<0.01). CONCLUSIONS: ARC1779 improves the post-transfusion recovery of refrigerated platelets and preserves the long-term hemostatic function of refrigerated platelets. These results suggest that a short-acting inhibitor of the platelet-VWF interaction may be a potential therapeutic option to improve refrigeration of platelets for transfusion treatment.

Targeted and effective glioblastoma therapy via aptamer-modified tetrahedral framework nucleic acid-paclitaxel nanoconjugates that can pass the blood brain barrier.

Targeted DNA nanoparticles have been identified as one of the most promising nanocarriers in anti-glioma drug delivery. We established a multifunctional nanosystem for targeted glioma therapy. Tetrahedral framework nucleic acid (tFNA), entering U87MG cells and bEnd.3 cells, was chosen to deliver two aptamers, GMT8 and Gint4.T, and paclitaxel. GMT8 and Gint4.T, which specifically bind with U87MG cells and with PDGFRß, were linked with tFNA, to form Gint4.T-tFNA-GMT8 (GTG). GTG was efficiently internalized by U87MG and bEnd.3 cells and penetrated an in-vitro blood-brain-barrier model. GTG loaded with paclitaxel (GPC) had potentiated anti-glioma efficacy. It inhibited the proliferation, migration, and invasion of U87MG cells, and enhanced apoptosis induction in these cells. The expression of apoptosis-related proteins was significantly changed after treatment with GPC, confirming apoptosis induction. Our study demonstrated that the combination of GTG and paclitaxel has great potential for glioma treatment and tFNA shows great promise for use in drug delivery.

"In vitro" behaviour of aptamer-functionalized polymeric nanocapsules loaded with 5-fluorouracil for targeted therapy.

New type of nanocapsules based on carboxymethyl chitosan functionalized with AS1411 aptamer and poly(N-vinylpyrrolidone-alt-itaconic anhydride) loaded with 5-Fluorouracil (5-FU) were developed, with the potential to improve the treatment of cancer. Functionalization of nanocapsules with AS1411 aptamer will enhance their recognition by tumor cells, due to the interaction with nucleolin, and subsequent endocytosis. Nanocapsules were prepared by interfacial condensation method in the absence of any toxic crosslinking agents. The condensation reaction took place at the interface between the organic and aqueous phases by opening the anhydride cycles from the copolymer, under the action of the NH2 groups from mixture of chitosan/aptamer-functionalized carboxymethyl chitosan. The nanocapsules diameter varied between 100 and 267 nm as a function of the molar ratio of the polymers. SEM images have revealed that nanocapsules were spherical and presented relatively low dimensional polydispersity. Nanocapsules swelling degree was found between 1000 and 1680% in PBS solution (pH = 7.4) and they allowed the encapsulation of an important amount of 5-Fluorouracil (5-FU). The release efficiency of 5-FU was studied, the processes being controlled by the drug diffusion through the polymeric membrane, as confirmed by the theoretical analysis of the drug release. The cytotoxicity and haemolysis tests performed on the nanocapsules proved their lack of toxicity and their excellent hemocompatibility. The obtained results were encouraging, showing that these original 5-FU-loaded nanocapsules were able to induce a more pronounced cytotoxic effect on neoplastic MCF-7 cells, the occurrence of dead cells being more rapidly than in the case of free 5-FU.

Identification and Application of an Aptamer Targeting Papillary Thyroid Carcinoma Using Tissue-SELEX.

Aptamers, short DNA or RNA oligonucleotides, which evolved from systematic evolution of ligands by exponential enrichment (SELEX), can perform specific target recognition. Papillary thyroid carcinoma (PTC) is of high incidence worldwide, and the prognosis of advanced PTC is poor. Up to now, there is no specific biomarker that can identify PTC and defects still remain in existing diagnostic methods. Here we report an aptamer, termed TC-6, which is generated from tissue-SELEX by using sections of papillary thyroid carcinoma and a normal thyroid gland. TC-6 could specifically target intracellular components of papillary thyroid cells with high affinity ( Kd = 57.66 ± 5.93 nmol/L) and have performed excellent biocompatibility both in vivo and in vitro. Moreover, fluorescence imaging of PTC tumor-bearing mice revealed that TC-6 was able to accumulate in tumor sites and could distinguish thyroid carcinoma from other benign thyroid diseases efficiently. In addition, TC-6d, a truncated aptamer of TC-6, maintained its affinity toward PTC with Kd of 39.20 ± 8.20 nmol/L. Overall, these results indicate that TC-6 is a potential candidate for developing novel tools for diagnosis and targeted therapy of PTC.

Defined Substrate by Aptamer Modification with the Balanced Properties of Selective Capture and Stemness Maintenance of Mesenchymal Stem Cells.

The recruitment of endogenous mesenchymal stem cells (MSCs), as an alluring approach for in situ tissue regeneration, always accompanies with other types of cells. Therefore, it is of enormous value to bestow a substrate with the property of selective capture to MSCs. However, it was reported that when MSCs are cultured on a substrate with excessive affinity, their stemness diminished. Therefore, constructing a substrate with the balanced ability of selective capture and stemness maintenance becomes a big challenge. In this study, an Aptamer 19S (Apt19S)-modified substrate was fabricated by grafting Apt19S on a PEGylated glass substrate. The X-ray photoelectron spectroscopy results verified that the antifouling poly(ethylene glycol) (PEG) layer was created. Tracking by ellipsometry, the thicknesses of PEG layers were proved to increase with PEG concentration. The results of the quartz crystal microbalance also validated that the Apt19S densities could be modulated by the concentrations of the Apt19S solution. The results of the cell adhesion assay indicated that the modification of Apt19S caused a significant increase in the adhesion ratio and area of rBMSCs. Selective adhesion was confirmed by coculture of rBMSCs with macrophages and NIH3T3 cells, demonstrating that a higher proportion of rBMSCs adhered to the Apt19S-modified substrate. The results of specific capture were further confirmed by a flow model to simulate the body fluid flow. The comprehensive results of reverse transcription polymerase chain reaction, immunofluorescence staining, proliferation capacity, and differentiation assay showed that the stemness of rBMSCs was maintained better on a substrate with the appropriate Apt19S density. All of these results indicated that Apt19S modification is an effective strategy to endow a substrate with the specific capture ability of MSCs, and the balance between selective capture and stemness maintenance can be achieved by the precise regulation of the aptamer density.

Binding Characterization of Aptamer-Drug Layered Microformulations and In Vitro Release Assessment.

Efficient delivery of adequate active ingredients to targeted malignant cells is critical, attributing to recurrent biophysical and biochemical challenges associated with conventional pharmaceutical delivery systems. These challenges include drug leakage, low targeting capability, high systemic cytotoxicity, and poor pharmacokinetics and pharmacodynamics. Targeted delivery system is a promising development to deliver sufficient amounts of drug molecules to target cells in a controlled release pattern mode. Aptameric ligands possess unique affinity targeting capabilities which can be exploited in the design of high pay-load drug formulations to navigate active molecules to the malignant sites. This study focuses on the development of a copolymeric and multifunctional drug-loaded aptamer-conjugated poly(lactide-co-glycolic acid)-polyethylenimine (PLGA-PEI) (DPAP) delivery system, via a layer-by-layer synthesis method, using a water-in-oil-in-water double emulsion approach. The binding characteristics, targeting capability, biophysical properties, encapsulation efficiency, and drug release profile of the DPAP system were investigated under varying conditions of ionic strength, polymer composition and molecular weight (MW), and degree of PEGylation of the synthetic core. Experimental results showed increased drug release rate with increasing buffer ionic strength. DPAP particulate system obtained the highest drug release of 50% at day 9 at 1 M NaCl ionic strength. DPAP formulation, using PLGA 65:35 and PEI MW of ∼800 Da, demonstrated an encapsulation efficiency of 78.93%, and a loading capacity of 0.1605 mg bovine serum albumin per mg PLGA. DPAP (PLGA 65:35, PEI MW∼25 kDa) formulation showed a high release rate with a biphasic release profile. Experimental data depicted a lower targeting power and reduced drug release rate for the PEGylated DPAP formulations. The outcomes from the present study lay the foundation to optimize the performance of DPAP system as an effective synthetic drug carrier for targeted delivery.

PET imaging of HER2 expression with an 18F-fluoride labeled aptamer.

BACKGROUND/PURPOSE: Aptamers are oligonucleotide or peptide molecules that bind to a target molecule with high affinity and specificity. The present study aimed to evaluate the target specificity and applicability for in vivo molecular imaging of an aptamer labeled with a radioisotope. METHODS: The human epidermal growth factor receptor 2 (HER2/ErbB2) aptamer was radiolabeled with 18F-fluoride. HER2-positive tumor cell uptake of the aptamer was evaluated in comparison to negative controls by flow cytometry and confocal microscopy. Using 18F-labeled HER2-specific aptamer positron emission tomography (PET), in vivo molecular images of BT474 tumor-bearing mice were taken at 60, 90 and 120 minutes after injection. RESULTS: In flow cytometric analysis, HER2 aptamer showed strong binding to HER2-positive BT474 cells, while binding to HER2-negative MDA-MB231 cells was quite low. Likewise, in confocal microscopic images, the aptamer was bound to HER2-positive breast cancer cells, with minimal binding to HER2-negative cells. In vivo PET molecular imaging of BT474 tumor-bearing mice revealed significant higher uptake of the 18F-labeled HER2 specific aptamer into the tumor compared to the that of HER2-negative cell tumor(p = 0.033). HER2 aptamer was able to preferentially bind to HER2-positive breast cancer cells both in vitro and in vivo, by recognizing HER2 structure on the surface of these cells. CONCLUSION: The 18F-labeled aptamer enabled appropriate visualization of HER2 expression by human breast cancer cells. The results suggest that a radiolabeled HER2 aptamer could potentially be applied in the development of treatment strategies or in targeted therapy against HER2-positive breast cancer cells.

PET imaging of EGFR expression using an 18F-labeled RNA aptamer.

OBJECTIVE: Epidermal growth factor receptor (EGFR) is a theranostic biomarker for a variety of cancer types. The aim of the present study was to develop an 18F radiolabeled EGFR targeting RNA aptamer, and to investigate its ability to visualize and quantify EGFR in xenograft models. METHODS: Biolayer interferometry binding assay was used to detect the binding affinity of the alkyne-modified EGFR aptamer MinE07 (denoted as ME07) with recombinant human wild-type EGFR protein and the mutant EGFRvIII protein. Cy5-conjugated ME07 was used for flow cytometry and immunofluorescence staining, and an Alexa Fluor 488-labeled EGFR antibody (ab193244) was used as a control. 18F-Fluorobenzoyl (FB) azide was employed as a synthon to produce 18F-FB-ME07 via click chemistry, and the cellular uptake and internalization characteristics of 18F-FB-ME07 were investigated. Static PET scans, 60-min dynamic scans, and biodistribution study of 18F-FB-ME07 were performed in three types of tumor models. RESULTS: The Kd values of ME07 to wtEGFR and EGFRvIII proteins were 0.3 nM and 271 nM respectively. The A431, U87MG, and HCT-116 cells showed strong, weak, and negative binding with Cy5-ME07, which is consistent with EGFR expression level in these cells. Peak cell uptake values of 18F-FB-ME07 in A431, U87MG and HCT-116 cells were 2.86%, 2.19% and 0.88% of the added dose respectively. The mean internalization of 18F-FB-ME07 in these cells were 60.02%, 53.1%, and 52.8% of the total accumulated radioactivity. In static PET imaging, despite high uptake in the liver and kidneys, 18F-FB-ME07 showed reasonable accumulation in A431 tumors (1.02 ± 0.13 %ID/g at 30 min after injection). Of note, the uptake of 18F-FB-ME07 in A431 xenografts was significantly higher than that in U87MG and HCT-116 xenografts. In A431 xenografted mice, the tumor/blood ratio was 3.89 and the tumor/muscle ratio reached 8.65. CONCLUSIONS: We for the first time generated an aptamer-derived EGFR targeting PET tracer 18F-FB-ME07, which showed highly selective targeting ability in mouse tumor models expressing different levels of EGFR. Our results suggest that 18F-FB-ME07 is a potential EGFR targeting molecular imaging probe for future clinical translation.

Pharmacokinetics, pharmacodynamics and safety of aptamers.

Aptamers are synthetic molecules structured as single-stranded DNA or RNA oligonucleotides that can be designed to mimic the functional properties of monoclonal antibodies. They bind to the target molecules (typically soluble or cell-bound proteins) with high affinity (with picomolar to low nanomolar range) and specificity, and therefore can be an alternative to therapeutic antibodies or peptide ligands. This paper reviews published data regarding pharmacokinetics, pharmacodynamics and safety of aptamers from preclinical and clinical studies. Aptamers have been developed for the treatment of a variety of diseases, including cancer, macular degeneration,g cardiovascular disease, diabetes and anaemia of chronic diseases. There are several preclinical studies with unmodified aptamers, but the vast majority of aptamer trials in humans have been conducted with modified aptamers, because unmodified aptamers demonstrate metabolic instability, as well as rapid renal filtration and elimination. Various strategies have been developed to improve the pharmacokinetic profile of aptamers. Aside from chemical modification of nucleotides in order to stabilize them against nuclease degradation, the main modification to extend the half-life is pegylation. Therefore, the process of pegylation as well as its benefits and possible shortcomings will briefly be discussed.

Designed multifunctional polymeric nanomedicines: long-term biodistribution and tumour accumulation of aptamer-targeted nanomaterials.

We report a novel multifunctional hyperbranched polymer based on polyethylene glycol (PEG) as a nanomedicine platform that facilitates longitudinal and quantitative 89Zr-PET imaging, enhancing knowledge of nanomaterial biodistribution and pharmacokinetics/pharmacodynamics both in vivo and ex vivo. Anti-VEGF-A DNA aptamer functionalization increased tumour accumulation by >2-fold in a breast cancer model.