Search

The use of the specific binding properties of monoclonal antibody fragments such as single-chain variable fragments (ScFv) for the selective delivery of antitumor therapeutics for cancer cells is attractive due to their smaller size, low immunogenicity, and low-cost production. Although covalent strategies for the preparation of such ScFv-based therapeutic conjugates are prevalent, this approach is not straightforward, as it requires prior chemical activation and/or modification of both the ScFv and the therapeutics for the application of robust chemistries. A non-covalent alternative based on ScFv fused to maltose-binding protein (MBP) acting as a binding adapter is proposed for active targeted delivery. MBP-ScFv proves to be a valuable modular platform to synergistically bind maltose-derivatized therapeutic cargos through the MBP, while preserving the targeting competences provided by the ScFv. The methodology has been tested by using a mutated maltose-binding protein (MBP I334W) with an enhanced affinity toward maltose and an ScFv coding sequence toward the human epidermal growth factor receptor 2 (HER2). Non-covalent binding complexes of the resulting MBP-ScFv fusion protein with diverse maltosylated therapeutic cargos (a near-infrared dye, a maltosylated supramolecular β-cyclodextrin container for doxorubicin, and non-viral polyplex gene vector) were easily prepared and characterized. In vitro and in vivo assays using cell lines that express or not the HER2 epitope, and mice xenografts of HER2 expressing cells demonstrated the capability and versatility of MBP-ScFv for diagnosis, imaging, and drug and plasmid active targeted tumor delivery. Remarkably, the modularity of the MBP-ScFv platform allows the flexible interchange of both the cargos and the coding sequence for the ScFv, allowing ad hoc solutions in targeting delivery without any further optimization since the MBP acts as a pivotal element.

Chromophore-appended cyclodextrins combine the supramolecular loading capabilities of cyclodextrins (CDs) with the optical properties of the affixed chromophores. Among fluorescent materials, carbon dots (CNDs) are attractive and the feasibility of CND-appended CDs as sensors has been demonstrated by different authors. However, CNDs are intrinsically heterogeneous materials and their ulterior functionalization yields hybrid composites that are not well defined in terms of structure and composition. Inspired by the fluorescence properties of 5-oxo-1,2,3,5-tetrahydroimidazo[1,2-a]pyridine-7-carboxylic acid (IPCA), the most paradigmatic of the molecular fluorophores detected in CNDs, herein we report two highly efficient synthetic chemical strategies for the preparation of IPCA-appended CDs that behave as CND-based CD "turn off-on" biosensors suitable for the analysis of cholesterol and β-galactosidase activity. We have deconstructed the CND-CD systems to demonstrate that (i) the role of CNDs is limited to acting as a support for the molecular fluorophores produced during their synthesis and (ii) the molecular fluorophores suffice for the determination of the enzymatic activity based on the quenching by p-nitrophenol as a sacrificial quencher.

Two simple boron-dipyrromethene-type fluorophore (azaBODIPYs) dyes are synthesized and tested for the determination of CO 2 gas by an inner filter process. The indicators are noncovalently entrapped in suitable polymers according to their polarity, featuring absorption maxima at 620 nm and fluorescent emission maxima in the range 675-720 nm. Molar absorptivity and fluorescence quantum yield data were determined for these two synthesized azaBODIPYs. These indicators have high molar absorption coefficients of 7.1 × 10 4 and 2.1 × 10 4 M -1 cm -1 and quantum yields of 21 and 9%. The pK a values of the indicators are determined from absorbance and fluorescence measurements with values of 7.9 and 8.5, depending on the positioning of the substitution pattern of the electron-donating functionalities. The two azaBODIPYs present excellent photostability, making them suitable for long duration measurements. These azaBODIPY dyes act as fluorescent pH indicators in a polymeric sensing membrane along with microcrystalline powder of chromium-doped gadolinium aluminium borate as the luminophore, a transfer phase agent (tetraoctyl or tetramethyl ammonium hydroxide) and a plasticizer or surfactant to improve membrane permeability to gaseous CO 2 . The response time ranges from 42 to 60 s and recovery time from 103 to 120 s, with a detection limit of 0.04 and 0.57% CO 2 . The store time of the sensing membranes is longer than 570 days in the best case, and it does not need to be kept in any special atmosphere other than darkness.

Saponins are potential wide-spectrum antitumor drugs, and copper(I) catalyzed azide-alkyne 1,3-dipolar cycloaddition is a suitable approach to synthesizing saponin-like compounds by regioselective glycosylation of the C2/C3 hydroxyl and C28 carboxylic groups of triterpene aglycones maslinic acid (MA) and oleanolic acid (OA). Biological studies on the T-84 human colon carcinoma cell line support the role of the hydroxyl groups at C2/C3, the influence of the aglycone, and the bulky nature of the substituents in C28. OA bearing a α-d-mannose moiety at C28 (compound 18 ) focused our interest because the estimated inhibitory concentration 50 was similar to that reported for ginsenoside Rh2 against colon cancer cells and it inhibits the G 1 -S phase transition affecting the cell viability and apoptosis. Considering that triterpenoids from natural sources have been identified as inhibitors of nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) signaling, docking studies were conducted to evaluate whether NF-κB may be a potential target. Results are consistent with the biological study and predict a similar binding mode of MA and compound 18 to the p52 subunit from NF-κB but not for OA. The fact that the binding site is shared by the NF-κB inhibitor 6,6-dimethyl-2-(phenylimino)-6,7-dihydrobenzo[ d ][1,3]oxathiol-4(5 H )-one supports the result and points to NF-κB as a potential target of both MA and compound 18 ., Competing Interests: The authors declare no competing financial interest.

Polymer-based nanotheranostics are appealing tools for cancer treatment and diagnosis in the fast-growing field of nanomedicine. A straightforward preparation of novel engineered PEI-based nanotheranostics incorporating NIR fluorescence heptamethine cyanine dyes (NIRF-HC) to enable them with tumor targeted gene delivery capabilities is reported. Branched PEI-2 kDa (b2kPEI) is conjugated with IR-780 and IR-783 dyes by both covalent and noncovalent simple preparative methodologies varying their stoichiometry ratio. The as-prepared set of PEI-NIR-HC nanocarriers are assayed in vitro and in vivo to evaluate their gene transfection efficiency, cellular uptake, cytotoxicity, internalization and trafficking mechanisms, subcellular distribution, and tumor specific gene delivery. The results show the validity of the approach particularly for one of the covalent IR783-b2kPEI conjugates that exhibit an enhanced tumor uptake, probably mediated by organic anion transporting peptides, and favorable intracellular transport to the nucleus. The compound behaves as an efficient nanotheranostic transfection agent in NSG mice bearing melanoma G361 xenographs with concomitant imaging signal and gene concentration in the targeted tumor. By this way, advanced nanotheranostics with multifunctional capabilities (gene delivery, tumor-specific targeting, and NIR fluorescence imaging) are generated in which the NIRF-HC dye component accounts for simultaneous targeting and diagnostics, avoiding additional incorporation of additional tumor-specific targeting bioligands.