Nanomolar Sensitivity Research Articles

A novel reaction-based fluorescent probe with a nanomolar sensitivity for detection of Hg(II) and its multiple application

An easily synthesized reaction-based probe for the fluorescence detection of Hg2+ ions using was reported. The designed probe exhibited “turn-on” fluorescence response towards Hg2+ ions via deprotection of the thiocarbonate-protecting group to alcohol in the HEPES/EtOH=8:2 (v/v, 5 mM, pH 7.4). The investigation results of probe Bzp-1 properties for Hg2+ detection indicated that probe Bzp-1 has satisfactory high selectivity and sensitivity. The detection limit of Bzp-1 for Hg2+ was found to be 4.2 nM. The recognition mechanism of Bzp-1 for Hg2+ was confirmed by ESI-MS. Moreover, the probe Bzp-1 has been successfully used to rapidly detect trace amounts of hazardous Hg2+ ions in real samples such as tap water, seafood and soil with good recoveries and less the relative standard deviations. Moreover, the Bzp-1 can also be used for fluorescence imagining of Hg2+ in living cells.

Thermally versatile maleimide-based polymeric thin film for detection and separation of picric acid from polluted areas

2,4,6-Trinitrophenol, commonly known as picric acid, is a powerful explosive that has been used historically in military ammunition. Picric acid is extremely toxic and sensitive to friction and heat; thus, it is prone to accidental detonation at high temperatures. It is critical to detect and separate picric acid from the impacted areas. In the present study, we designed and developed a fluorescent copolymer (P1) for temperature-dependent picric acid detection. The solvatochromic and aggregation-induced emission (AIE) properties of P1 were utilized to detect picric acid at a high temperature of 40 °C (high alert level) by turn-off emission. Additionally, a polymeric thin film (F1) was fabricated by immobilizing P1 onto the surface of quartz slide. Notably, F1 retained its emissive properties across a range of temperatures (both relatively low and high alert levels), facilitated by its AIE characteristics. The portable film, F1, displayed a nanomolar sensitivity toward picric acid, making it suitable for on-site detection in aqueous media (neutral pH). Furthermore, F1 demonstrated efficient separation of picric acid from nitroaromatic explosive mixtures with a separation efficiency of 70 % over four cycles. Importantly, F1 exhibited practical utility in detecting and separating picric acid in real samples, highlighting its potential for environmental monitoring applications.

Wearable, Biocompatible, and Dual-Emission Ocular Multisensor Patch for Continuous Profiling of Fluoroquinolone Antibiotics in Tears.

The abuse or misuse of antibiotics in clinical and agricultural settings severely endangers human health and ecosystems, which has raised profound concerns for public health worldwide. Trace detection and reliable discrimination of commonly used fluoroquinolone (FQ) antibiotics and their analogues have consequently become urgent to guide the rational use of antibiotic medicines and deliver efficient treatments for associated diseases. Herein, we report a wearable eye patch integrated with a quadruplex nanosensor chip for noninvasive detection and discrimination of primary FQ antibiotics in tears during routine eyedrop treatment. A set of dual-mode fluorescent nanoprobes of red- or green-emitting CdTe quantum dots integrated with lanthanide ions and a sensitizer, adenosine monophosphate, were constructed to provide an enhanced fluorescence up to 45-fold and nanomolar sensitivity toward major FQs owing to the aggregation-regulated antenna effect. The aggregation-driven, CdTe-Ln(III)-based microfluidic sensor chip is highly specific to FQ antibiotics against other non-FQ counterparts or biomolecular interfering species and is able to accurately discriminate nine types of FQ or non-FQ eyedrop suspensions using linear discriminant analysis. The prototyped wearable sensing detector has proven to be biocompatible and nontoxic to human tissues, which integrates the entire optical imaging modules into a miniaturized, smartphone-based platform for field use and reduces the overall assay time to ∼5 min. The practicability of the wearable eye patch was demonstrated through accurate quantification of antibiotics in a bactericidal event and the continuous profiling of FQ residues in tears after using a typical prescription antibiotic eyedrop. This technology provides a useful supplement to the toolbox for on-site and real-time examination and regulation of inappropriate daily drug use that might potentially lead to long-term antibiotic abuse and has great implications in advancing personal healthcare techniques for the regulation of daily medication therapy.

Aptamer-Based Potentiometric Sensor Enables Highly Selective and Neurocompatible Neurochemical Sensing in Rat Brain.

Selective and nondisruptive in vivo neurochemical monitoring within the central nervous system has long been a challenging endeavor. We introduce a new sensing approach that integrates neurocompatible galvanic redox potentiometry (GRP) with customizable phosphorothioate aptamers to specifically probe dopamine (DA) dynamics in live rat brains. The aptamer-functionalized GRP (aptGRP) sensor demonstrates nanomolar sensitivity and over a 10-fold selectivity for DA, even amidst physiological levels of major interfering species. Notably, conventional sensors without the aptamer modification exhibit negligible reactivity to DA concentrations exceeding 20 μM. Critically, the aptGRP sensor operates without altering neuronal activity, thereby permitting real-time, concurrent recordings of both DA flux and electrical signaling in vivo. This breakthrough establishes aptGRP as a viable and promising framework for the development of high-fidelity sensors, offering novel insights into neurotransmission dynamics in a live setting.

A naked-eye colorimetric assay for highly sensitive detection of CL-20 explosive

2,4,6,8,10,12-hexanitro-2,4,6,8,10,12-hexaazaisowurtzitane (CL-20) stands as the most energetically condensed explosive ever found, and poses a great security threat to humans. Here we report a naked-eye colorimetric assay for highly sensitive detection of CL-20. Through effective cascading of the hydrolysis process of CL-20 with nitrite-involved Griess reaction to yield a pink-colored azo product, the visual quantification of CL-20 is realized by taking advantage of corresponding colorimetric signals. Under optimal experimental conditions, the present colorimetric assay is successfully used for highly selective determination of CL-20 in deionized water and environmental water samples with a nanomolar sensitivity. The resulting limit of detection is much lower than conventional methods. Moreover, benefiting from the distinct color change of the detection solution, direct naked-eye semi-quantitative analysis of CL-20 is achieved with a threshold concentration of 1 μM. Such simple, cost-effective, selective, and highly sensitive visual colorimetric assay will have valuable applications in public safety, environmental monitoring, and other related fields.

Abstract A036: A subset of lung adenocarcinomas defined by high-level ERBB2 amplification is vulnerable to HER2-targeted therapy

Abstract Background: Most lung adenocarcinomas (LUAD) are driven by activation of receptor tyrosine kinases (RTK). HER2 (encoded by ERBB2) is an RTK capable of ligand-independent dimerization and signaling when overexpressed. ERBB2 amplification (ERBB2amp) is a common cause of HER2 overexpression. While activating ERBB2 mutations (e.g., ERBB2 exon 20 YVMA insertions) are targetable drivers of LUAD, the role of ERBB2amp in lung cancer (LC) is unclear and there is no FDA-approved therapy for this indication in LC. Here, we report the clinicopathological and molecular characteristics of LUAD with high-level ERBB2amp and investigate the efficacy of HER2-targeted agents in LC models with ERBB2amp. We also demonstrate the clinical benefit of trastuzumab-deruxtecan (T-DXd) in a patient with ERBB2amp LUAD. Design: An institutional database of hybrid capture DNA NGS panel targeting 290-447 genes (v1-v3.1) was interrogated for LUAD with high-level ERBB2amp (defined as at least 6 calculated gene copies by NGS). HER2 immunohistochemistry (IHC) was performed on 29 available cases and scored according to ASCO/CAP scoring criteria. HER2 IHC scores of 2+ and 3+ were considered overexpression. Two LC cell lines, H2170 and CALU3 were identified as ERBB2amp and their responses to HER2 inhibitors were characterized with Western blotting, viability, and apoptosis assays. CALU3 and H2170 xenograft models were also established and treated with HER2 inhibitors or T-DXd. Results: ERBB2amp was identified in 31/3,684 (0.8%) LUAD patients. HER2 overexpression was present in 24 out of 29 evaluable cases (79.3%). Eighteen cases (58%) showed high-level ERBB2amp with no concurrent drivers and were more likely to be male (P = 0.001) and have a history of tobacco use (P = 0.001), compared to patients with high-level ERBB2amp with a concurrent driver mutation (activating ERBB2 SNV/indels, activating EGFR and MET mutations, RET fusion). H2170 and CALU3 cells displayed overexpression of HER2 and phospho-HER2, and growth of these cells showed sub-nanomolar sensitivity to trastuzumab and T-DXd, and low nanomolar sensitivity to afatinib. T-DXd induced apoptosis in both cell lines (assessed by activation of caspase 3/7 and elevation of BIM and cleaved PARP levels). While trastuzumab and afatinib significantly slowed down growth of H2170 and CALU3 xenografts, only T-DXd achieved tumor shrinkage equivalent to partial (H2170) or complete response (CALU3) in all animals. We describe the case of a LUAD patient with a RET rearrangement who acquired high-level ERBB2amp after progression on selpercatinib and was treated with T-DXd, with evidence of disease stability and clinical benefit. Conclusion: High-level ERBB2 amplification detected by NGS reliably predicts HER2 overexpression in lung adenocarcinoma. LUAD with ERBB2 amplification and no concurrent oncogenic drivers is a rare event that occurs predominantly in male smokers. This data can inform future trials investigating the utility of HER2-targeting agents in this population. Citation Format: Igor Odintsov, Maisam Makarem, Sara E. Bachert, Tom Zhang, Romel Somwar, Pasi A. Jänne, Lynette M. Sholl. A subset of lung adenocarcinomas defined by high-level ERBB2 amplification is vulnerable to HER2-targeted therapy [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A036.

Bispecific Antibody Detection Using Antigen-Conjugated Synthetic Nucleic Acid Strands.

We report here the development of two different sensing strategies based on the use of antigen-conjugated nucleic acid strands for the detection of a bispecific antibody against the tumor-related proteins Mucin1 and epidermal growth factor receptor. Both approaches work well in serum samples (nanomolar sensitivity), show high specificity against the two monospecific antibodies, and are rapid. The results presented here demonstrate the versatility of DNA-based platforms for the detection of bispecific antibodies and could represent a versatile alternative to other more reagent-intensive and time-consuming analytical approaches.

Decorated DNA-Based Scaffolds as Lateral Flow Biosensors.

Here we develop Lateral Flow Assays (LFAs) that employ as functional elements DNA-based structures decorated with reporter tags and recognition elements. We have rationally re-engineered tile-based DNA tubular structures that can act as scaffolds and can be decorated with recognition elements of different nature (i.e. antigens, aptamers or proteins) and with orthogonal fluorescent dyes. As a proof-of-principle we have developed sandwich and competitive multiplex lateral flow platforms for the detection of several targets, ranging from small molecules (digoxigenin, Dig and dinitrophenol, DNP), to antibodies (Anti-Dig, Anti-DNP and Anti-MUC1/EGFR bispecific antibodies) and proteins (thrombin). Coupling the advantages of functional DNA-based scaffolds together with the simplicity of LFAs, our approach offers the opportunity to detect a wide range of targets with nanomolar sensitivity and high specificity.

Decorated DNA‐Based Scaffolds as Lateral Flow Biosensors

AbstractHere we develop Lateral Flow Assays (LFAs) that employ as functional elements DNA‐based structures decorated with reporter tags and recognition elements. We have rationally re‐engineered tile‐based DNA tubular structures that can act as scaffolds and can be decorated with recognition elements of different nature (i.e. antigens, aptamers or proteins) and with orthogonal fluorescent dyes. As a proof‐of‐principle we have developed sandwich and competitive multiplex lateral flow platforms for the detection of several targets, ranging from small molecules (digoxigenin, Dig and dinitrophenol, DNP), to antibodies (Anti‐Dig, Anti‐DNP and Anti‐MUC1/EGFR bispecific antibodies) and proteins (thrombin). Coupling the advantages of functional DNA‐based scaffolds together with the simplicity of LFAs, our approach offers the opportunity to detect a wide range of targets with nanomolar sensitivity and high specificity.

(Invited) Laser-Induced Graphene on Polymers: From Process Science to Applications

Nanocarbons like graphene and carbon nanotubes (CNTs) are promising for various applications including advanced electronic devices, novel energy systems, and next-generation healthcare diagnostics. This is owing to the excellent physical, chemical and electrochemical properties arising from the ordered atomic structure, the hierarchical nanoscale morphology, and tunable chemistry of nanocarbons. In particular, low-dimensional carbon electrodes for biosensors and neural interfaces have consistently been shown to have superior performance when compared to state-of-the-art metal electrodes. Nevertheless, major manufacturing challenges still hinder our ability to scalably produce graphene-based electrodes with tailored morphology and surface chemistry, especially on flexible substrates. While chemical vapor deposition (CVD) processes enable the synthesis of high quality graphene and CNTs, the extreme environments of high temperatures and hydrocarbon-rich gaseous atmosphere in such reactors limit the choice of substrates to silicon, quartz, metals or other rigid temperature-resistant materials. On the other hand, many emerging flexible devices require the fabrication of such nanocarbon electrodes on the surface of polymer substrates. Unlike different transfer technique of CVD-grown nanocarbons, this talk will focus on a unique bottom-up approach for directly growing different types of graphenic nanocarbons on polymer films by laser irradiation. The speaker will show how this direct-write process, often referred to as laser-induced graphene (LIG), can be controlled to produce spatially-varying morphologies and chemical compositions of LIG electrodes, by leveraging gradients of laser fluence. Moreover, a method will be introduced to control the heteroatom doping of these LIG electrodes based on controlling the molecular structure of the polymer being lased. Finally, a demonstration of these functional LIG electrodes as electrochemical biosensors will be presented for the detection of the neurotransmitter dopamine with nanomolar sensitivity.

Broad-spectrum activity of bulevirtide against clinical isolates of HDV and recombinant pan-genotypic combinations of HBV/HDV

Bulevirtide (BLV) is a small lipopeptide agent that specifically binds to the sodium taurocholate cotransporting polypeptide (NTCP) bile salt transporter and HBV/HDV receptor on the surface of human hepatocytes and inhibits HDV and HBV entry. As a satellite virus of HBV, HDV virions are formed after assembly of HDV RNA with the HBV envelope proteins (HBsAg). Because both viruses exist as eight different genotypes, this creates a potential for high diversity in the HBV/HDV combinations. To investigate the sensitivity of various combinations of HBV/HDV genotypes to BLV, clinical and laboratory strains were assessed. For the laboratory strains, the different envelopes from HBV genotypes A through H were combined with HDV genotypes 1-8 in cotransfection assays. Clinical plasma isolates were obtained from clinical studies and academic collaborations to maximise the diversity of HBV/HDV genotypes tested. The mean BLV EC50 against HDV laboratory strains ranged from 0.44 to 0.64nM. Regardless of HBV and HDV genotypes, the clinical isolates showed similar sensitivities to BLV with mean values that ranged from 0.2 to 0.73nM. These data support the use of BLV in patients infected with any HBV/HDV genotypes. This study describes the potent activity of BLV against multiple laboratory strains spanning all HBV/HDV A-H/1-8 genotype combinations and the most diverse collection of HDV clinical samples tested to date, including HBV/HDV genotype combinations less frequently observed in the clinic. Overall, all isolates and laboratory strains displayed similar invitro nanomolar sensitivity to BLV. This broad-spectrum antiviral activity of BLV has direct implications on potential simplified treatment for any patient infected with HDV, regardless of genotype, and supports the new 2023 EASL Clinical Practice Guidelines on HDV that recommend antiviral treatment for all patients with CHD.

Rapid genetic screening with high quality factor metasurfaces

Genetic analysis methods are foundational to advancing personalized medicine, accelerating disease diagnostics, and monitoring the health of organisms and ecosystems. Current nucleic acid technologies such as polymerase chain reaction (PCR) and next-generation sequencing (NGS) rely on sample amplification and can suffer from inhibition. Here, we introduce a label-free genetic screening platform based on high quality (high-Q) factor silicon nanoantennas functionalized with nucleic acid fragments. Each high-Q nanoantenna exhibits average resonant quality factors of 2,200 in physiological buffer. We quantitatively detect two gene fragments, SARS-CoV-2 envelope (E) and open reading frame 1b (ORF1b), with high-specificity via DNA hybridization. We also demonstrate femtomolar sensitivity in buffer and nanomolar sensitivity in spiked nasopharyngeal eluates within 5 minutes. Nanoantennas are patterned at densities of 160,000 devices per cm2, enabling future work on highly-multiplexed detection. Combined with advances in complex sample processing, our work provides a foundation for rapid, compact, and amplification-free molecular assays.

Experimental and simulation studies of bioinspired Au-enhanced copper single atom catalysts towards real-time expeditious dopamine sensing on human neuronal cell

Single atom catalysts (SACs) becoming important participants in biosensing owing to their characteristics such as dispersed metal active sites, signal amplification, and acceptable sensitivity and selectivity toward distinct biomolecules. A recent advancement is the extension of SACs to form dual-metal single atom catalysts with high metal loading and flexible active sites, thus enhancing the electrochemical activity and biosensing ability. This study reports carbon-supported dual-metal single atoms, bioinspired from barnacle shell-extracted chitosan, for electrochemically detecting dopamine in real-time cellular environments and biofluids. Single copper and gold atoms anchored on bioinspired chitosan-extracted carbon (CuAu SACs/BC) based electrode was fabricated and used for the selective and sensitive detection of nanomolar dopamine in biological and cellular environments. Density functional theory calculations revealed that dopamine is predominantly sensed by single Cu atoms, together with the signal enhancement by single Au atoms. Collectively, the CuAu SACs/BC-based detection platform enabled real-time dopamine detection in neuronal cells upon triggering by a precursor with good biocompatibility and nanomolar sensitivity in male, female geriatric plasma. Thus, this study demonstrated the effective utilization of bioinspired carbon with dual-metal single atom catalysts to develop promising real-time electrochemical biosensors.

Discovery of Peptidic Ligands against the SARS-CoV-2 Spike Protein and Their Use in the Development of a Highly Sensitive Personal Use Colorimetric COVID-19 Biosensor.

In addition to efficacious vaccines and antiviral therapeutics, reliable and flexible in-home personal use diagnostics for the detection of viral antigens are needed for effective control of the COVID-19 pandemic. Despite the approval of several PCR-based and affinity-based in-home COVID-19 testing kits, many of them suffer from problems such as a high false-negative rate, long waiting time, and short storage period. Using the enabling one-bead-one-compound (OBOC) combinatorial technology, several peptidic ligands with a nanomolar binding affinity toward the SARS-CoV-2 spike protein (S-protein) were successfully discovered. Taking advantage of the high surface area of porous nanofibers, immobilization of these ligands on nanofibrous membranes allows the development of personal use sensors that can achieve low nanomolar sensitivity in the detection of the S-protein in saliva. This simple biosensor employing naked-eye reading exhibits detection sensitivity comparable to some of the current FDA-approved home detection kits. Furthermore, the ligand used in the biosensor was found to detect the S-protein derived from both the original strain and the Delta variant. The workflow reported here may enable us to rapidly respond to the development of home-based biosensors against future viral outbreaks.

Directed Evolution of Herbicide Biosensors in a Fluorescence-Activated Cell-Sorting-Compatible Yeast Two-Hybrid Platform.

Developing sensory modules for specific molecules of interest represents a fundamental challenge in synthetic biology and its applications. A somewhat generalizable approach for this challenge is demonstrated here by evolving a naturally occurring chemically induced heterodimer into a genetically encoded sensor for herbicides. The interaction between PYRABACTIN-RESISTANT-like receptors and type-2C protein phosphatases is induced by abscisic acid—a small-molecule hormone in plants. We considered abscisic acid receptors as a potential scaffold for the development of biosensors because of past successes in their engineering, a structurally defined ligand cavity and the availability of large-scale assays for their activation. A panel of 475 receptor variants, mutated at ligand-proximal residues, was screened for activation by 37 herbicides from several classes. Twelve compounds activated at least one member of the mutant panel. To facilitate the subsequent improvement of herbicide receptors through directed evolution, we engineered a yeast two-hybrid platform optimized for sequential positive and negative selection using fluorescence-activated cell sorting. By utilizing this system, we were able to isolate receptors with low nanomolar sensitivity and a broad dynamic range in sensing a ubiquitous group of chloroacetamide herbicides. Aside from its possible applicative value, this work lays down conceptual groundwork and provides infrastructure for the future development of biosensors through directed evolution.

Rapid biosensor development using plant hormone receptors as reprogrammable scaffolds

A general method to generate biosensors for user-defined molecules could provide detection tools for a wide range of biological applications. Here, we describe an approach for the rapid engineering of biosensors using PYR1 (Pyrabactin Resistance 1), a plant abscisic acid (ABA) receptor with a malleable ligand-binding pocket and a requirement for ligand-induced heterodimerization, which facilitates the construction of sense–response functions. We applied this platform to evolve 21 sensors with nanomolar to micromolar sensitivities for a range of small molecules, including structurally diverse natural and synthetic cannabinoids and several organophosphates. X-ray crystallography analysis revealed the mechanistic basis for new ligand recognition by an evolved cannabinoid receptor. We demonstrate that PYR1-derived receptors are readily ported to various ligand-responsive outputs, including enzyme-linked immunosorbent assay (ELISA)-like assays, luminescence by protein-fragment complementation and transcriptional circuits, all with picomolar to nanomolar sensitivity. PYR1 provides a scaffold for rapidly evolving new biosensors for diverse sense–response applications.

Label‐Free Spontaneous Raman Sensing in Photonic Crystal Fibers with Nanomolar Sensitivity

AbstractAn approach to significantly enhance spontaneous Raman sensitivity through the formation of a thin film via thermophoresis along with evaporation at the facet of a hollow‐core photonic crystal fiber is reported for the first time. Sensitivity of detection is increased by more than 6 orders of magnitude for both organic and inorganic nanoparticles, facilitating the search for trace analytes in solution. Detection of two nanoparticles, alumina and polystyrene, is demonstrated down to 392 nM without the use of surface‐enhanced Raman spectroscopy or other chemical‐based procedures. This new thin‐film deposition approach simplifies the simultaneous detection and analysis of small trace compounds, a previously arduous task using conventional spontaneous Raman.

A simple 4-amino-1,8-naphthalimide hydrazine based “turn-on” fluorescent chemosensor for selective and reversible detection of Zn(II) ion

A new 4-amino-1,8-naphthalimide hydrazine fluorophore (Nap-NH2) has been synthesized in quantitative yield via a one-pot imidation reaction between 4-amino-1,8-naphthalic anhydride and hydrazine hydrate (N2H4·H2O) and successfully employed as a fluorescent “turn-on” chemosensor for metal cations. The sensor Nap-NH2 displayed a positive solvatochromism in different solvents with varying polarities. The fluorescence titration studies of Nap-NH2 with various metal cations disclosed the high selectivity and reversible binding affinity of Nap-NH2 towards Zn(II) ions with nanomolar (10 nm) sensitivity. The fluorescence emission intensity of Nap-NH2 was drastically enhanced upon binding with Zn(II) ion and the Stern-Volmer binding constant is KSV = 1.13 × 104 M−1. Interestingly, the presence of commonly coexisting different metal cations {Fe(II), Cu(II), Mg(II), Cd(II), Ni(II), Co(II), Mn(II), Ba(II), Al(III), Ce(III), Sn(II), Pb(II), Cr(III) and Ca(II)} showed negligible changes in the emission intensity and thus weak binding interactions with Nap-NH2. These studies demonstrate that Nap-NH2 is a potential fluorescence “turn-on” chemosensor for selective, sensitive detection and quantification of biologically relevant metal cations.

Abstract PO-036: LP184, a novel alkylating agent, is highly effective in pancreatic cancers with DNA damage repair defects

Abstract Biomarker-based chemotherapy with increased efficacy and prolonged disease-free survival are urgently needed in pancreatic cancer (PDAC). A 15-20% subset of PDAC tumors carry mutations in DNA repair pathway (BRCA1/BRCA2/PALB2/RAD51/ATM/FANCD2). Additionally, mutations in nucleotide excision repair (NER) genes (ERCC2/3/4/5/6) have been reported in ~5% of PDAC. LP184 is a novel synthetic small molecule acylfulvene analog. Using precise synthetic chemistry, we determined that only a negative enantiomer of LP184 is converted to an active alkylating agent in the strict dependency on oxidoreductase, prostaglandin reductase 1 (PTGR1). By computational analyses, we demonstrate a strong positive correlation of LP184 sensitivity with PTGR1 transcript levels (r=0.89, p<10−15) in a broad panel of cancer cell lines. Once activated by PTGR1, highly reactive LP-184 nucleophile creates covalent DNA adducts that are selectively repaired via Nucleotide Excision Repair (NER) mechanism coupled to transcription (TC-NER) and/or homologous recombination (HR). We reasoned that mutation or expression driven TC-NER and HR deficiency would predispose PDAC cells to increased sensitivity to LP184. To test the idea of LP184 activity in DNA repair-deficient tumors, we evaluated LP184 chemosensitivity in genetically defined PDAC models in vitro, ex vivo, and in xenografts. Testing in six different pancreatic cancer cell lines (Capan-1, CFPAC-1, Panc1, MiaPaCa2, Panc03.27 and BxPC-3) resulted in very potent inhibition with LP184 IC50 values ranging from 114 to 182 nM. In this cell line panel, LP184 sensitivity correlated negatively with transcript levels of an NER pathway gene ERCC8 (r = -0.94). In comparison to these PDAC cell lines, a normal pancreatic epithelial cell line HPNE was 3-6 times less sensitive to LP184 (IC50 670 nM). Ex vivo cultures of 4 out of 5 low-passage patient-derived xenografts with HR deficiency showed nanomolar sensitivity to LP184 with IC50s ranging from 45 to 270 nM. These tumor graft models which were at least 6 times less sensitive to olaparib in the same assay. Depletion of ERCC4 enhanced sensitivity to LP184 about 2-fold relative to the parental cell line. To define PTGR1 as a biomarker for LP184 activity, we used CRISPR/Cas9-mediated gene editing to deplete PTGR1 expression. We found PTGR1-null Capan-1 cell line-derived xenografts were poorly sensitive to LP184, whereas PTGR1-expressing xenografts showed near complete tumor regression in all LP184 treated animals with 109% tumor growth inhibition relative to the control group in this study. Furthermore, PTGR1 depleted cells were completely resistant to LP184 in vitro. Our preclinical data demonstrate that PDAC models carrying a range of DNA repair pathway mutations are highly sensitive to LP-184 in vitro and in vivo. Increased PTGR1 expression is a validated biomarker for LP184 cytotoxicity, and is the exclusive convertase of LP184 to an active alkylator drug. We anticipate LP184 will extend the therapeutic opportunities to a large subset of PDAC patients carrying these genetic alterations. Citation Format: Diana Restifo, Aditya Kulkarni, Caleb Schimke, Joseph McDermott, Umesh Kathad, Kishor Bhatia, Panna Sharma, Igor Astsaturov. LP184, a novel alkylating agent, is highly effective in pancreatic cancers with DNA damage repair defects [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-036.

Multiplexed detection of heavy metal ions by single plasmonic nanosensors

Detection of multiple analytes simultaneously in small liquid samples with high efficiency and precision is highly important to the fields like water quality monitoring. In this letter, we present a multiplexed nanosensors with position-encoded aptamer functionalized gold nanorods for heavy metal ions detection. The individual gold nanorods respond specifically to two different heavy metal ions (Pb2+ and Hg2+) with a spectral shift in the scattering spectrum. We used a home-built spectral imaging dark-field microscope to measure the response of thousands of single plasmonic nanosensors with relatively high time resolution and precision. To explore the performance and limit of detection (LOD) of our nanosensor and setup, we recorded the concentration-dependent response of our position-encoded nanosensors with a series of mixture solutions that contain different concentrations of Hg2+ and Pb2+ ions. The LOD levels of our system are around 5 nM for Pb2+ ions and 1 nM for Hg2+ ions. Our method and results demostrate the nanomolar sensitivity and the potential to detect more different heavy metal ions.