Elucidating Raman Image-Guided Differential Recognition of Clinically Confirmed Grades of Cervical Exfoliated Cells by Dual Biomarker-Appended SERS-Tag.

Ultrasensitive detection of cancer biomarkers via single-cell analysis through Raman imaging is an impending approach that modulates the possibility of early diagnosis. Cervical cancer is one such type that can be monitored for a sufficiently long period toward invasive cancer phenotype. Herein, we report a surface-enhanced Raman scattering (SERS) nanotag (SERS-tag) for the simultaneous detection of p16/K-i67, a dual biomarker persisting in the progression of squamous cell carcinoma of human cervix. A nanoflower-shaped SERS-tag, constituted of hybrid gold nanostar with silver tips to achieve maximum fingerprint enhancement from the incorporated reporter molecule, was further functionalized with the cocktail monoclonal antibodies against p16/K-i67. The recognition by the SERS-tag was first validated in cervical squamous cell carcinoma cell line SiHa as a foot-step study and subsequently implemented to different grades of clinically confirmed exfoliated cells including normal cell (NC), high-grade intra-epithelial lesion (HC), and squamous cell carcinoma (CC) samples of the cervix. Precise Raman mapped images were constituted based on the average intensity gradient of the signature Raman peaks arising from different grades of exfoliated cells. We observed a distinct intensity hike of around 10-fold in the single dysplastic HC and CC samples in comparison to NC specimen, which clearly justify the prevalence of p16/Ki-67. The synthesized probe is able to map the abnormal cells within 20 min with high reproducibility and stability for 1 mm × 1 mm mapping area with good contrast. Amidst the challenges in Raman image-guided modality, the technique was further complemented with the gold standard immunocytochemistry (ICC) dual staining analysis. Even though both are time-consuming techniques, tedious steps can be avoided and real-time readout can be achieved using the SERS mapping unlike immunocytochemistry technique. Therefore, the newly developed Raman image-guided SERS imaging emphasizes the approach of uplifting of SERS in practical utility with further improvement for clinical applications for cervical cancer detection in future.

Targeted Theranostic Nano Vehicle Endorsed with Self-Destruction and Immunostimulatory Features to Circumvent Drug Resistance and Wipe-Out Tumor Reinitiating Cancer Stem Cells.

The downsides of conventional cancer monotherapies are profound and enormously consequential, as drug-resistant cancer cells and cancer stem cells (CSC) are typically not eliminated. Here, a targeted theranostic nano vehicle (TTNV) is designed using manganese-doped mesoporous silica nanoparticle with an ideal surface area and pore volume for co-loading an optimized ratio of antineoplastic doxorubicin and a drug efflux inhibitor tariquidar. This strategically framed TTNV is chemically conjugated with folic acid and hyaluronic acid as a dual-targeting entity to promote folate receptor (FR) mediated cancer cells and CD44 mediated CSC uptake, respectively. Interestingly, surface-enhanced Raman spectroscopy is exploited to evaluate the molecular changes associated with therapeutic progression. Tumor microenvironment selective biodegradation and immunostimulatory potential of the MSN-Mn core are safeguarded with a chitosan coating which modulates the premature cargo release and accords biocompatibility. The superior antitumor response in FR-positive syngeneic and CSC-rich human xenograft murine models is associated with a tumor-targeted biodistribution, favorable pharmacokinetics, and an appealing bioelimination pattern of the TTNV with no palpable signs of toxicity. This dual drug-loaded nano vehicle offers a feasible approach for efficient cancer therapy by on demand cargo release in order to execute complete wipe-out of tumor reinitiating cancer stem cells.

Emergence of Gold-Mesoporous Silica Hybrid Nanotheranostics: Dox-Encoded, Folate Targeted Chemotherapy with Modulation of SERS Fingerprinting for Apoptosis Toward Tumor Eradication.

Strategically fabricated theranostic nanocarrier delivery system is an unmet need in personalized medicine. Herein, this study reports a versatile folate receptor (FR) targeted nanoenvelope delivery system (TNEDS) fabricated with gold core silica shell followed by chitosan-folic acid conjugate surface functionalization by for precise loading of doxorubicin (Dox), resembled as Au@SiO2 -Dox-CS-FA. TNEDS possesses up to 90% Dox loading efficiency and internalized through endocytosis pathway leading to pH and redox-sensitive release kinetics. The superior FR-targeted cytotoxicity is evaluated by the nanocarrier in comparison with US Food and Drug Administration (FDA)-approved liposomal Dox conjugate, Lipodox. Moreover, TNEDS exhibits theranostic features through caspase-mediated apoptosis and envisages high surface plasmon resonance enabling the nanoconstruct as a promising surface enhanced Raman scattering (SERS) nanotag. Minuscule changes in the biochemical components inside cells exerted by the TNEDS along with the Dox release are evaluated explicitly in a time-dependent fashion using bimodal SERS/fluorescence nanoprobe. Finally, TNEDS displays superior antitumor response in FR-positive ascites as well as solid tumor syngraft mouse models. Therefore, this futuristic TNEDS is expected to be a potential alternative as a clinically relevant theranostic nanomedicine to effectively combat neoplasia.

Unveiling NIR Aza-Boron-Dipyrromethene (BODIPY) Dyes as Raman Probes: Surface-Enhanced Raman Scattering (SERS)-Guided Selective Detection and Imaging of Human Cancer Cells.

The development of new Raman reporters has attracted immense attention in diagnostic research based on surface enhanced Raman scattering (SERS) techniques, which is a well established method for ultrasensitive detection through molecular fingerprinting and imaging. Herein, for the first time, we report the unique and efficient Raman active features of the selected aza-BODIPY dyes 1-6. These distinctive attributes could be extended at the molecular level to allow detection through SERS upon adsorption onto nano-roughened gold surface. Among the newly revealed Raman reporters, the amino substituted derivative 4 showed high signal intensity at very low concentrations (ca. 0.4 µm for 4-Au). Interestingly, an efficient nanoprobe has been constructed by using gold nanoparticles as SERS substrate, and 4 as the Raman reporter (4-Au@PEG), which unexpectedly showed efficient recognition of three human cancer cells (lung: A549, cervical: HeLa, Fibrosarcoma: HT-1080) without any specific surface marker. We observed well reflected and resolved Raman mapping and characteristic signature peaks whereas, such recognition was not observed in normal fibroblast (3T3L1) cells. To confirm these findings, a SERS nanoprobe was conjugated with a specific tumour targeting marker, EGFR (Epidermal Growth Factor Receptor), a well known targeted agent for Human Fibrosarcoma (HT1080). This nanoprobe efficiently targeted the surface marker of HT1080 cells, threreby demonstrating its use as an ultrasensitive Raman probe for detection and targeted imaging, leaving normal cells unaffected.

A clinically feasible diagnostic spectro-histology built on SERS-nanotags for multiplex detection and grading of breast cancer biomarkers.

Simultaneous detection of multiple biomarkers is always an obstacle in immunohistochemical (IHC) analysis. Herein, a straightforward spectroscopy-driven histopathologic approach has emerged as a paradigm of Raman-label (RL) nanoparticle probes for multiplex recognition of pertinent biomarkers in heterogeneous breast cancer. The nanoprobes are constructed by sequential incorporation of signature RL and target specific antibodies on gold nanoparticles, which are coined as Raman-Label surface enhanced Raman scattering (RL-SERS)-nanotags to evaluate simultaneous recognition of clinically relevant breast cancer biomarkers i.e., estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor2 (HER2). As a foot-step assessment, breast cancer cell lines having varied expression levels of the triple biomarkers are investigated. Subsequently, the optimized detection strategy using RL-SERS-nanotags is subjected to clinically confirmed, retrospective formalin-fixed paraffin embedded (FFPE) breast cancer tissue samples to fish out the quick response of singleplex, duplex as well as triplex biomarkers in a single tissue specimen by adopting a ratiometric signature RL-SERS analysis which enabled to minimize the false negative and positive results. Significantly, sensitivity and specificity of 95% and 92% for singleplex, 88% and 85% for duplex, and 75% and 67% for triplex biomarker has been achieved by assessing specific Raman fingerprints of the respective SERS-tags. Furthermore, a semi-quantitative evaluation of HER2 grading between 4+/2+/1+ tissue samples was also achieved by the Raman intensity profiling of the SERS-tag, which is fully in agreement with the expensive fluorescent in situ hybridization analysis. Additionally, the practical diagnostic applicability of RL-SERS-tags has been achieved by large area SERS imaging of areas covering 0.5-5 mm2 within 45 min. These findings unveil an accurate, inexpensive and multiplex diagnostic modality envisaging large-scale multi-centric clinical validation.

Raman Imaging: An Impending Approach Towards Cancer Diagnosis.

In accordance with the recent studies, Raman spectroscopy is well experimented as a highly sensitive analytical and imaging technique in biomedical research, mainly for various disease diagnosis including cancer. In comparison with other imaging modalities, Raman spectroscopy facilitate numerous assistances owing to its low background signal, immense spatial resolution, high chemical specificity, multiplexing capability, excellent photo stability and non-invasive detection capability. In cancer diagnosis Raman imaging intervened as a promising investigative tool to provide molecular level information to differentiate the cancerous vs non-cancerous cells, tissues and even in body fluids. Anciently, spontaneous Raman scattering is very feeble due to its low signal intensity and long acquisition time but new advanced techniques like coherent Raman scattering (CRS) and surface enhanced Raman scattering (SERS) gradually superseded these issues. So, the present review focuses on the recent developments and applications of Raman spectroscopy-based imaging techniques for cancer diagnosis.

Exploring the margins of SERS in practical domain: An emerging diagnostic modality for modern biomedical applications.

Excellent multiplexing capability, molecular specificity, high sensitivity and the potential of resolving complex molecular level biological compositions augmented the diagnostic modality of surface-enhanced Raman scattering (SERS) in biology and medicine. While maintaining all the merits of classical Raman spectroscopy, SERS provides a more sensitive and selective detection and quantification platform. Non-invasive, chemically specific and spatially resolved analysis facilitates the exploration of SERS-based nano probes in diagnostic and theranostic applications with improved clinical outcomes compared to the currently available so called state-of-art technologies. Adequate knowledge on the mechanism and properties of SERS based nano probes are inevitable in utilizing the full potential of this modality for biomedical applications. The safety and efficiency of metal nanoparticles and Raman reporters have to be critically evaluated for the successful translation of SERS in to clinics. In this context, the present review attempts to give a comprehensive overview about the selected medical, biomedical and allied applications of SERS while highlighting recent and relevant outcomes ranging from simple detection platforms to complicated clinical applications.

New Insight of Tetraphenylethylene-based Raman Signatures for Targeted SERS Nanoprobe Construction Toward Prostate Cancer Cell Detection.

We have designed and synthesized novel tetraphenylethylene (TPE) appended organic fluorogens and unfold their unique Raman fingerprinting reflected by surface-enhanced Raman scattering (SERS) upon adsorption on nanoroughened gold surface as a new insight in addition to their prevalent aggregation-induced emission (AIE) and aggregation-caused quenching (ACQ) phenomena. A series of five TPE analogues has been synthesized consisting of different electron donors such as (1) indoline with propyl (TPE-In), (2) indoline with lipoic acid (TPE-In-L), (3) indoline with Boc-protected propyl amine (TPE-In-Boc), (4) benzothaizole (TPE-B), and (5) quinaldine (TPE-Q). Interestingly, all five TPE analogues produced multiplexing Raman signal pattern, out of which TPE-In-Boc showed a significant increase in signal intensity in the fingerprint region. An efficient SERS nanoprobe has been constructed using gold nanoparticles as SERS substrate, and the TPE-In as the Raman reporter, which conjugated with a specific peptide substrate, Cys-Ser-Lys-Leu-Gln-OH, well-known for the recognition of prostate-specific antigen (PSA). The designated nanoprobe TPE-In-PSA@Au acted as SERS "ON/OFF" probe in peace with the vicinity of PSA protease, which distinctly recognizes PSA expression with a limit of detection of 0.5 ng in SERS platform. Furthermore, TPE-In-PSA@Au nanoprobe was efficiently recognized the overexpressed PSA in human LNCaP cells, which can be visualized through SERS spectral analysis and SERS mapping.