High expression of integrin αvβ3 enables uptake of targeted fluorescent probes into ovarian cancer cells and tumors

Shaw, Scott K.; Schreiber, Cynthia L.; Roland, Felicia M.; Battles, Paul M.; Brennan, Seamus P.; Padanilam, Simon J.; Smith, Bradley D.

doi:10.1016/j.bmc.2018.03.007

Cited by 23 publications

(20 citation statements)

References 45 publications

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“…[32] We have recently demonstrated that the αvβ3 integrin receptor is expressed at high levels ([9.8 ± 2.5] × 10 4 /cell) on the surface of OVCAR-4 cells, and that the cells are amenable to molecular targeting and imaging using covalently-conjugated fluorophores bearing the cRGDfK antagonist. [33] In contrast A549 adenocarcinoma cells do not express αvβ3, but they do express the integrin αvβ5 receptor, which also has good affinity for the cRGDfK motif. [34] Expression of αvβ5 in A549 cells was corroborated using immunocytochemistry staining with an anti-β5 FITC-conjugated antibody (Figure S6).…”

Section: Resultsmentioning

confidence: 99%

“…[38] The average tumor-to-muscle ratio of 3.6 for mice that were treated with targeted 2(R) ⊃ H3H is similar to the tumor selectivity ratios reported for analogous but completely covalent RGD-targeted molecular probes in similar mouse tumor models. [33,39–41] Moreover, a tumor selectivity ratio of 3 or more is a desired benchmark for effective fluorescence guided surgery. [42,43]…”

Section: Resultsmentioning

confidence: 99%

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Non‐Covalently Pre‐Assembled High‐Performance Near‐Infrared Fluorescent Molecular Probes for Cancer Imaging

Shaw

Liu

Durán

et al. 2018

Chemistry A European J

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New fluorescent molecular probes, which can selectively target specific cell surface receptors, are needed for microscopy, in vivo imaging, and image guided surgery. The preparation of multivalent probes using standard synthetic chemistry can be a laborious process due to low reaction yields caused by steric effects. In this study, fluorescent molecular probes were prepared by a programmed non-covalent pre-assembly process that used a near-infrared fluorescent squaraine dye to thread a macrocycle bearing a cyclic arginine-glycine-aspartate peptide antagonist (cRGDfK) as a cancer targeting unit. Cell microscopy studies using OVCAR-4 (ovarian cancer) and A549 (lung cancer) cells that express high levels of the integrin αvβ3 or αvβ5 receptors, respectively, revealed a multivalent cell targeting effect. That is, there was comparatively more cell uptake of a pre-assembled probe equipped with two copies of the cRGDfK antagonist than a pre-assembled probe with only one appended cRGDfK antagonist. The remarkably high photostability and low phototoxicity of these near-infrared probes allowed for acquisition of long-term fluorescence movies showing endosome trafficking in living cells. In vivo near-infrared fluorescence imaging experiments compared the biodistribution of a targeted and untargeted probe in a xenograft mouse tumor model. The average tumor-to-muscle ratio for the pre-assembled targeted probe was 3.6 which matches the tumor targeting performance reported for analogous cRGDfK-based probes that were prepared entirely by covalent synthesis. The capability to excite these pre-assembled near-infrared fluorescent probes with blue or deep-red excitation light makes it possible to determine if a target site is located superficially or buried in tissue, a probe performance feature that is likely to be very helpful for eventual applications such as fluorescence guided surgery.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Non‐Covalently Pre‐Assembled High‐Performance Near‐Infrared Fluorescent Molecular Probes for Cancer Imaging

Shaw

Liu

Durán

et al. 2018

Chemistry A European J

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“…The linear tripeptide RGD and its cyclic-RGD analogs are well-known peptides for cell targeting. These peptide ligands bind α v β 3 integrins, which are overexpressed in several cancers, including melanomas, 93 ovarian cancer, 94 and breast cancer, as well as in metastasis. 95 Grafting such ligands to therapeutic oligonucleotides provides a means for targeting and efficient delivery.…”

Section: Bioconjugated Oligonucleotide Deliverymentioning

confidence: 99%

Bioconjugated Oligonucleotides: Recent Developments and Therapeutic Applications

et al. 2019

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Oligonucleotide-based agents have the potential to treat or cure almost any disease, and are one of the key therapeutic drug classes of the future. Bioconjugated oligonucleotides, a subset of this class, are emerging from basic research and being successfully translated to the clinic. In this review, we first briefly describe two approaches for inhibiting specific genes using oligonucleotides -antisense DNA (ASO) and RNA interference (RNAi)followed by a discussion on delivery to cells. We then summarize and analyze recent developments in bioconjugated oligonucleotides including those possessing GalNAc, cell penetrating peptides, αtocopherol, aptamers, antibodies, cholesterol, squalene, fatty acids, or nucleolipids. These novel conjugates provide a means to enhance tissue targeting, cell internalization, endosomal escape, target binding specificity, resistance to nucleases, and more. We next describe those bioconjugated oligonucleotides approved for patient use or in clinical trials. Finally, we summarize the state of the field, describe current limitations, and discuss future prospects. Bioconjugation chemistry is at the centerpiece of this therapeutic oligonucleotide revolution, and significant opportunities exist for development of new modification chemistries, for mechanistic studies at the chemical-biology interface, and for translating such agents to the clinic.

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“…In view of the uptake results, we have selected the U87MG cell line as the most suitable model for further experiments, starting with the confirmation that their nanoparticle uptake was in fact mediated by a specific, integrin mediated pathway. To this end, cells were pre-incubated for 30 min with several uptake inhibitors or competitors, including excess soluble RGD peptides 55,56 or antibodies against specific heterodimers 57 , followed by a 1 h co-incubation with rhodamine-labelled particles (please refer to Supplementary Information 4SI and Supplementary Fig. 6SI.A for cytotoxicity studies of these inhibitors/competitors).…”

Section: Evaluation Of Nanoparticle Uptake and Intracellular Sortingmentioning

confidence: 99%

Microfluidic-assisted preparation of RGD-decorated nanoparticles: exploring integrin-facilitated uptake in cancer cell lines

rosa

Spadea

Donno

et al. 2020

Sci Rep

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This study is about fine tuning the targeting capacity of peptide-decorated nanoparticles to discriminate between cells that express different integrin make-ups. Using microfluidic-assisted nanoprecipitation, we have prepared poly(lactic acid-co-glycolic acid) (PLGA) nanoparticles with a PEGylated surface decorated with two different arginine-glycine-aspartic acid (RGD) peptides: one is cyclic (RGDFC) and has specific affinity towards α v β 3 integrin heterodimers; the other is linear (RGDSP) and is reported to bind equally α v β 3 and α 5 β 1. We have then evaluated the nanoparticle internalization in two cell lines with a markedly different integrin fingerprint: ovarian carcinoma A2780 (almost no α v β 3 , moderate in α 5 β 1) and glioma U87MG (very high in α v β 3 , moderate/high in α 5 β 1). As expected, particles with cyclic RGD were heavily internalized by U87MG (proportional to the peptide content and abrogated by anti-α v β 3) but not by A2780 (same as PEGylated particles). The linear peptide, on the other hand, did not differentiate between the cell lines, and the uptake increase vs. control particles was never higher than 50%, indicating a possible low and unselective affinity for various integrins. The strong preference of U87MG for cyclic (vs. linear) peptide-decorated nanoparticles was shown in 2D culture and further demonstrated in spheroids. Our results demonstrate that targeting specific integrin make-ups is possible and may open the way to more precise treatment, but more efforts need to be devoted to a better understanding of the relation between RGD structure and their integrinbinding capacity. Integrins are targetable receptors. They are transmembrane glycoproteins, which connect cell bodies to pericellular structures and therefore are critical in cell-cell and cell-environment interactions 1. Structurally, they are heterodimeric proteins composed of non-covalently linked α and β subunits 2. To date 18α and 8β subunits, and 24 different heterodimers have been reported in humans, with each heterodimer displaying unique binding properties, tissue distribution, and biological functions 1 ,3. Although integrins are virtually ubiquitous, their heterodimeric identity and level of expression are a physio-pathological signature, which has a specific relevance in cancer 4 , e.g. in the formation of metastatic niches or in angiogenetic processes 5. Most studies have focused either on integrins in peritumoral environments, e.g. on endothelial cells (ECs), critical to (neo)angiogenesis, or on those directly present on tumor cells 6. It is now well known that α v-containing integrins are highly expressed in both scenarios, but poorly expressed on healthy cells 7. In particular, α v β 3 (e.g. in pro-tumoral ECs) is the most abundant integrin capable of regulating angiogenesis 8. Among other heterodimers, also α 5 β 1 has been

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High expression of integrin αvβ3 enables uptake of targeted fluorescent probes into ovarian cancer cells and tumors

Cited by 23 publications

References 45 publications

Non‐Covalently Pre‐Assembled High‐Performance Near‐Infrared Fluorescent Molecular Probes for Cancer Imaging

Non‐Covalently Pre‐Assembled High‐Performance Near‐Infrared Fluorescent Molecular Probes for Cancer Imaging

Bioconjugated Oligonucleotides: Recent Developments and Therapeutic Applications

Microfluidic-assisted preparation of RGD-decorated nanoparticles: exploring integrin-facilitated uptake in cancer cell lines

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