Bio-Conjugated Magnetic-Fluorescence Nanoarchitectures for the Capture and Identification of Lung-Tumor-Derived Programmed Cell Death Lighand 1-Positive Exosomes

Abstract

As per the American Cancer Society, lung cancer is the leading cause of cancer-related death worldwide. Since the accumulation of exosomal programmed cell death ligand 1 (PD-L1) is associated with therapeutic resistance in programmed cell death 1 (PD-1) and PD-L1 immunotherapy, tracking PD-L1-positive (PD-L1 (+)) exosomes is very important for predicting anti-PD-1 and anti-PD-L1 therapy for lung cancer. Herein, we report the design of an anti-PD-L1 monoclonal antibody-conjugated magnetic-nanoparticle-attached yellow fluorescent carbon dot (YFCD) based magnetic-fluorescence nanoarchitecture for the selective separation and accurate identification of PD-L1-expressing exosomes. In this work, photostable YFCDs with a good photoluminescence quantum yield (23%) were synthesized by hydrothermal treatment. In addition, nanoarchitectures with superparamagnetic (28.6 emu/g), biocompatible, and selective bioimaging capabilities were developed by chemically conjugating the anti-PD-L1 antibody and YFCDs with iron oxide nanoparticles. Importantly, using human non-small-cell lung cancer H460 cells lines, which express a high amount of PD-L1 (+) exosomes, A549 lung cancer cells lines, which express a low amount of PD-L1 (+) exosomes, and the normal skin HaCaT cell line, which does not express any PD-L1 (+) exosomes, we demonstrate that nanoarchitectures are capable of effectively separating and tracking PD-L1-positive exosomes simultaneously. Furthermore, as a proof-of-concept of clinical setting applications, a whole blood sample infected with PD-L1 (+) exosomes was analyzed, and our finding shows that this nanoarchitecture holds great promise for clinical applications.

Human ACE2 Peptide-Attached Plasmonic-Magnetic Heterostructure for Magnetic Separation, Surface Enhanced Raman Spectroscopy Identification, and Inhibition of Different Variants of SARS-CoV-2 Infections

“…As reported in Figure F, Raman intensity from 4-aminothiophenol was enhanced sharply in the presence of virus, after magnetic separation. The observed strong Raman bands at ∼1590 cm –1 can be assigned as a 1 vibrational modes, which is due to the ν­(C–C + NH 2 bend) vibrations. − Similarly, we also observed strong Raman bands at ∼1078 cm –1 , which is mainly due to the ν­(C–C) + δ­(C–H)] mode. − In our SERS spectra, we also observed b 2 mode Raman peaks at ∼1435 cm –1 , which is due to the CC stretch in the Ph ring + NH 2 rock. We also observed a strong peak at ∼464 cm –1 due to ν­(C–N) + ν­(C–S) + γ­(CCC) vibrational modes. − …”

“…We wanted to understand whether ACE2 peptide-attached plasmonic-magnetic heterostructures have the capability to prevent COVID infection via blocking virus entry into the host cells. Initially, to determine the biosafety, we determined the possible toxicity data of the plasmonic-magnetic heterostructures using different cell lines such as human embryonic kidney-239T (HEK-293T) cells, normal skin HaCaT cells, lung cancer H1264 cells, breast cancer SK-BR-3 cells, and prostate cancer LnCaP cells. ,− Experimental details are reported in the Supporting Information. In brief, for this purpose, plasmonic-magnetic heterostructures were treated with normal and cancerous cells for 48 h, and then a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to determine the cell viability. ,− Cell viability data reported in Figure F indicate that cell viability for normal and cancerous cells hardly changed even after 48 h of incubation with plasmonic-magnetic heterostructures.…”

“…To understand whether ACE2 peptide-attached plasmonic-magnetic heterostructures can be used for effective magnetic separation, SERS-based identification and complete inhibition of pseudotyped SARS-CoV-2 bearing the Delta variant spike from infected artificial nasal mucus fluid samples, we performed experiments using pseudotyped SARS-CoV-2 infected artificial nasal mucus fluid samples. For this purpose, initially we developed 4-aminothiolphenol (4-ATP) Raman reporter-attached bioconjugated plasmonic-magnetic heterostructures. − To design 4-ATP-attached heterostructures, we used Au–S conjugation chemistry to attach 4-ATP on the surface of GNPs on heterostructures. − Next, for the development of infected nasal mucus samples, different concentrations of pseudotyped SARS-CoV-2 bearing the Delta variant spike were added to the artificial nasal mucus (Biochemazone, Canada) sample with buffer solution. After that, we added different concentrations of 4-ATP-attached bioconjugated plasmonic-magnetic heterostructures and kept the mixture for 30 min with stirring.…”

“…To determine how “hot spot” formation helps to increase the Raman signal, we performed theoretical calculations using finite-difference time-domain (FDTD) simulation, as we have reported before. ,, Details of theoretical modeling were reported by us and others before. − In brief, for FDTD simulation, we used 10 nm spherical gold nanoparticles, as observed in the TEM image reported in Figure B. For our FDTD simulation, the GNP aggregate structure was used from TEM/SEM data, as reported in Figure D,E.…”

“…In our FDTD simulation, we used 670 nm wavelength, 0.001 nm mesh resolution, and 4000 fs time, as we have reported before. ,, As reported in Figure H, the square of the field (| E | 2 ) is enhanced 3000 times in “hot spots”. As we and others have reported before, − Raman enhancement is proportional to | E |, and as a result, from FDTD simulation data show a six orders of magnitude Raman enhancement due to the “hot spot” formation.…”