Delivery of mRNA with Histidine-Lysine Peptides.

Transfection with mRNA has been considered superior to that with plasmids since the mRNA can be translated to a protein in the cytosol without entering the nucleus. One disadvantage of using mRNA is its susceptibility to enzymatic biodegradability, and consequently, significant research has occurred to determine nonviral carriers that will sufficiently stabilize this nucleic acid for cellular transport. Histidine-lysine peptides (HK) are one such class of mRNA carriers, which we think serves as a model for other peptides and polymeric carrier systems. When the HK peptide and mRNA are mixed and interact through ionic and nonionic bonds, mRNA polyplexes are formed, which can transfect cells. In contrast to linear HK peptides, branched HK peptides protected and efficiently transfected mRNA into cells. After describing the preparation and biophysical characterization of these polyplexes, we will provide protocols for in vitro and in vivo transfection for these mRNA polyplexes.

pH-Dependent Non-Covalent Release of Chemotherapy from Carriers.

Although Warburg discovered pH discrepancies between tumor and normal tissues nearly 100 years ago, developing therapies to take advantage of this concept was relatively slow for the first 70 years. During the last 30 years, there has been an exponential increase in the use of pH-dependent strategies for both low molecular weight drugs and nanoparticles. Two frequently discussed approaches are the chemotherapy's release from pH-sensitive covalent linkages of macromolecules or from pH-dependent disruption of charged polymeric nanoparticles. In contrast, pH-dependent non-covalent bonds between the chemotherapy agent and macromolecules have rarely been discussed, yet this underappreciated strategy has great potential. These non-covalent interactions are primarily ionic or hydrogen bonds with supporting roles from hydrophobic bonds. In addition to the facile coupling of the drug with the carrier, these non-covalent interactions may show marked pH dependence. Consistent with pH dependence, many of these drug-loaded carriers showed significant in vitro and, in some cases, striking in vivo activity. In this review, we will focus on pH-sensitive non-covalent bonds, highlighting the release of drugs from diverse carriers such as tetrahedron DNA structures, cyclodextrin, polymeric carriers, and carbon-based quantum particles.

Cytokine Signatures for Lung Cancer Diagnosis in African American Populations.

Lung cancer is the leading cause of cancer-related deaths among both men and women. African Americans (AAs) experience disproportionately higher incidence and mortality compared to other ethnic groups. Cytokines play multifaceted and crucial roles in the initiation, progression, and spread of cancer. Our aim was to identify cytokine biomarkers for the early detection of lung cancer in AAs. We examined eight key cytokines (Interleukin-1, IL-6, IL-8, IL-10, IL-12p70, monocyte chemotactic protein-1 (MCP-1), interferon-gamma (IFN-γ), and tumor necrosis factor-alpha (TNF-α)) in the plasma of 104 lung cancer patients and 48 cancer-free individuals using the FirePlex Immunoassay. These findings were subsequently validated in a separate cohort of 58 cases and 58 controls. IL-8, IFN-γ, and TNF-α exhibited elevated levels in both AA and White American (WA) lung cancer cases. Notably, IL-10 and MCP-1 displayed significant increases specifically in AA lung cancer patients, with MCP-1 levels associated with lung adenocarcinoma cases. Conversely, WA lung cancer patients showed heightened IL-6 levels, particularly linked to lung adenocarcinoma. The combined use of specific cytokines showed promise in lung cancer diagnosis, with IL-8, IL-10, and MCP-1 achieving 76% sensitivity and 79% specificity in AAs and IL-6 and IL-8 combined offering 76% sensitivity and 74% specificity in WAs. These diagnostic biomarkers were validated in the independent cohort. The ethnicity-related cytokine biomarkers hold promise for diagnosing lung cancer in AAs and WAs, potentially addressing the observed racial disparity.

Delivery of Chemotherapy Agents and Nucleic Acids with pH-Dependent Nanoparticles.

With less than one percent of systemically injected nanoparticles accumulating in tumors, several novel approaches have been spurred to direct and release the therapy in or near tumors. One such approach depends on the acidic pH of the extracellular matrix and endosomes of the tumor. With an average pH of 6.8, the extracellular tumor matrix provides a gradient for pH-responsive particles to accumulate, enabling greater specificity. Upon uptake by tumor cells, nanoparticles are further exposed to lower pHs, reaching a pH of 5 in late endosomes. Based on these two acidic environments in the tumor, various pH-dependent targeting strategies have been employed to release chemotherapy or the combination of chemotherapy and nucleic acids from macromolecules such as the keratin protein or polymeric nanoparticles. We will review these release strategies, including pH-sensitive linkages between the carrier and hydrophobic chemotherapy agent, the protonation and disruption of polymeric nanoparticles, an amalgam of these first two approaches, and the release of polymers shielding drug-loaded nanoparticles. While several pH-sensitive strategies have demonstrated marked antitumor efficacy in preclinical trials, many studies are early in their development with several obstacles that may limit their clinical use.

Circulating Bacterial DNA as Plasma Biomarkers for Lung Cancer Early Detection.

Lung cancer is a leading cause of cancer deaths and early diagnosis can significantly improve outcomes. Pathogenic bacteria have been shown to play a role in tumorigenesis and its analysis provides a new approach for cancer diagnosis. To evaluate the potential of bacteria as plasma biomarkers for early lung cancer detection, we analyzed eight lung-cancer-related bacterial genera in 58 lung cancer patients and 58 controls using ddPCR. Our results showed that five genera had higher DNA abundance in lung tumor tissues compared with normal tissues. Three of these genera (Selenomonas, Streptococcus, and Veillonella) displayed consistent changes in plasma, with higher DNA abundance in lung cancer patients compared with controls. When used as a panel, these three bacterial genera had a sensitivity of 75% and specificity of 78% for lung cancer detection, regardless of stage or histology. The performance of this biomarker panel was confirmed in an independent cohort of 93 lung cancer cases and 93 controls. Thus, circulating bacterial DNA has the potential to be used as plasma biomarkers for early lung cancer detection.

pH-Sensitive Targeting of Tumors with Chemotherapy-Laden Nanoparticles: Progress and Challenges.

Accumulating chemotherapeutic drugs such as doxorubicin within a tumor while limiting the drug dose to normal tissues is a central goal of drug delivery with nanoparticles. Liposomal products such as Doxil® represent one of the marked successes of nanoparticle-based strategies. To replicate this success for cancer treatment, many approaches with nanoparticles are being explored in order to direct and release chemotherapeutic agents to achieve higher accumulation in tumors. A promising approach has been stimulus-based therapy, such as the release of chemotherapeutic agents from the nanoparticles in the acidic environments of the tumor matrix or the tumor endosomes. Upon reaching the acidic environments of the tumor, the particles, which are made up of pH-dependent polymers, become charged and release the entrapped chemotherapy agents. This review discusses recent advances in and prospects for pH-dependent histidine-based nanoparticles that deliver chemotherapeutic agents to tumors. The strategies used by investigators include an array of histidine-containing peptides and polymers which form micelles, mixed micelles, nanovesicles, polyplexes, and coat particles. To date, several promising histidine-based nanoparticles have been demonstrated to produce marked inhibition of tumor growth, but challenges remain for successful outcomes in clinical trials. The lessons learned from these histidine-containing particles will provide insight in the development of improved pH-dependent polymeric delivery systems for chemotherapy.

Novel Method for Rapid Detection of COVID-19 Omicron Variant and Future Emerging Variants.

Over six million people have died worldwide as a result of SARS-CoV-19 (Covid-19). Several major and minor variant waves of Covid-19 including the Alpha (B.1.1.7), Delta (B.1.617.2), and the Omicron (B.1.1.529) have infected patients in the last two years. These individual variants have been associated with different infectivity and death rates, and it is anticipated that other variants of significance will occur. As a result, the emergence and frequency of the different Covid-19 variants at various locations in the world are critically important to monitor. To date, sequencing of these variants has been the primary method to track the prevalence of the different variants, but this approach is relatively slow and expensive. In contrast, we have developed two qPCR methods that are sensitive, specific, and economical which can distinguish the Omicron variant from the other variants. The first qPCR method detects a specific base mutation in Omicron enabling improved amplification. The second method is based on designing a primer specific for the region in which a deletion and an insertion have occurred in the Omicron variant. While the first approach can be readily adapted to identify other variants with point mutations, the second method can identify emerging deletion/insertion mutations such as BA.2, a subvariant of the Omicron.

Targeting KDM1B-dependent miR-215-AR-AGR2-axis promotes sensitivity to enzalutamide-resistant prostate cancer.

Post-translational modifications of histones by histone demethylases plays an important role in the regulation of gene transcription and are implicated in cancers. Castrate resistant prostate cancer (CRPC) is often driven by constitutively active androgen receptor and commonly becomes resistant to established hormonal therapy strategies such as enzalutamide as a result. However, the role of KDM1B involved in next generation anti-enzalutamide resistance and the mechanisms of KDM1B regulation are poorly defined. Here, we show that KDM1B is upregulated and correlated with prostate cancer progression and poor prognosis. Downregulation of miR-215 is correlated with overexpression of KDM1B in enzalutamide-resistant prostate cancer cells, which promotes AR-dependent AGR2 transcription and regulates the sensitivity to next generation AR-targeted therapy. Inhibition of KDM1B significantly inhibits prostate tumor growth and improves enzalutamide treatments through AGR2 suppression. Our studies demonstrate inhibition of KDM1B can offer a viable therapeutic option to overcome enzalutamide resistance in tumors with deregulated miR-215-KDM1B-AR-AGR2 signaling axis.

Microbiota Biomarkers for Lung Cancer.

Non-small cell lung cancer (NSCLC) is the number one cancer killer and its early detection can reduce mortality. Accumulating evidences suggest an etiopathogenic role of microorganisms in lung tumorigenesis. Certain bacteria are found to be associated with NSCLC. Herein we evaluated the potential use of microbiome as biomarkers for the early detection of NSCLC. We used droplet digital PCR to analyze 25 NSCLC-associated bacterial genera in 31 lung tumor and the paired noncancerous lung tissues and sputum of 17 NSCLC patients and ten cancer-free smokers. Of the bacterial genera, four had altered abundances in lung tumor tissues, while five were aberrantly abundant in sputum of NSCLC patients compared with their normal counterparts (all p < 0.05). Acidovorax and Veillonella were further developed as a panel of sputum biomarkers that could diagnose lung squamous cell carcinoma (SCC) with 80% sensitivity and 89% specificity. The use of Capnocytophaga as a sputum biomarker identified lung adenocarcinoma (AC) with 72% sensitivity and 85% specificity. The use of Acidovorax as a sputum biomarker had 63% sensitivity and 96% specificity for distinguishing between SCC and AC, the two major types of NSCLC. The sputum biomarkers were further validated for the diagnostic values in a different cohort of 69 NSCLC cases and 79 cancer-free controls. Sputum microbiome might provide noninvasive biomarkers for the early detection and classification of NSCLC.

Sensitive Detection of KRAS Mutations by Clustered Regularly Interspaced Short Palindromic Repeats.

Kirsten rat sarcoma viral oncogene (KRAS) is the isoform most frequently mutated in human tumors. Testing for activating KRAS mutations has important implications for diagnosis and the personalized medicine of cancers. The current techniques for detecting KRAS mutations have moderate sensitivity. The emerging clustered regularly interspaced short palindromic repeats (CRISPR) system shows great promise in the detection of nucleic acids and is revolutionizing medical diagnostics. This study aimed to develop CRISPR-Cas12a as a sensitive test to detect KRAS mutations. Serially diluted DNA samples containing KRAS mutations are subjected to CRISPR-Cas12a and polymerase chain reaction (PCR). CRISPR-Cas12a and PCR can specifically detect 0.01% and 0.1% mutant KRAS DNA in the presence of wild-type KRAS DNA, respectively. Twenty pairs of lung tumor and noncancerous lung tissues are tested by CRISPR-Cas12a, PCR, and direct sequencing. CRISPR-Cas12a could identify the G12C mutation in five of 20 tumor tissues, while both PCR and direct sequencing discovered the KRAS mutation in three of the five tumor tissues. Furthermore, the results of CRISPR-Cas12a for testing the mutation could be directly and immediately visualized by a UV light illuminator. Altogether, CRISPR-Cas12a has a higher sensitivity for the detection of KRAS mutations compared with PCR and sequencing analysis, and thus has diagnostic and therapeutic implications. Nevertheless, the technique needs to be validated for its clinical significance in a large and prospective study.

Identification of a novel differentially methylated region adjacent to ATG16L2 in lung cancer cells using methyl-CpG binding domain protein-enriched genome sequencing.

Lung cancer is the most common cancer worldwide. Epigenetic modifications like DNA methylation play fundamental roles in the dynamic process of lung cancer. The objective of this study was to use methyl-CpG binding domain protein-enriched genome sequencing (MBD-Seq) to identify novel and high-confidence DNA methylation in lung tumor. We first compared the whole-genome DNA methylation of three lung cancer cell lines, including A549, H1299, and SK-MES-1, against BEAS-2B, a lung/bronchial normal epithelial cell line. We then used pyrosequencing and OneStep qMethyl kit methods to verify the results in the cell line specimens. MBD-Seq identified 279, 8046, and 22 887 differentially methylated regions (DMRs), respectively, with 120 common DMRs among three comparison groups. Three DMRs were consistent with the MBD-Seq results by both pyrosequencing and OneStep qMethyl validations. Furthermore, OneStep qMethyl kit was also performed for functional validation of these three potential DMRs in sputum DNA from clinical participants. We successfully identified one new DMR adjacent to ATG16L2. The novel DMR might have an important function in lung carcinogenesis. Further validation of the finding in clinical specimens of lung cancer patients and functional analysis of this novel DMR in the development of lung cancer through transcriptional silencing of ATG16L2 are warranted.

Location of a single histidine within peptide carriers increases mRNA delivery.

BACKGROUND: Previously, we determined that four-branched histidine-lysine (HK) peptides were effective carriers of plasmids and small interfering RNA. In the present study, we compared several branched HK carriers and, in particular, two closely-related H3K4b and H3K(+H)4b peptides for their ability as carriers of mRNA. The H3K(+H)4b peptide differed from its parent analogue, H3K4b, by only a single histidine in each branch. METHODS: A series of four-branched HK peptides with varied sequences was synthesized on a solid-phase peptide synthesizer. The ability of these peptides to carry mRNA expressing luciferase to MDA-MB-231 cells was investigated. With gel retardation and heparin displacement assays, the stability of HK polyplexes was examined. We determined the intracellular uptake of HK polyplexes by flow cytometry and fluorescence microscopy. The size and polydispersity index of the polyplexes in several media were measured by dynamic light scattering. RESULTS: MDA-MB-231 cells transfected by H3K(+H)4b-mRNA polyplexes expressed 10-fold greater levels of luciferase than H3K4b polyplexes. With gel retardation and heparin displacement assays, the H3K(+H)4b polyplexes showed greater stability than H3K4b. Intracellular uptake and co-localization of H3K(+H)4b polyplexes within acidic endosomes were also significantly increased compared to H3K4b. Similar to H3K(+H)4b, several HK analogues with an additional histidine in the second domain of their branches were effective carriers of mRNA. When combined with DOTAP liposomes, H3K(+H)4b was synergistic in delivery of mRNA. CONCLUSIONS: H3K(+H)4b was a more effective carrier of mRNA than H3K4b. Mechanistic studies suggest that H3K(+H)4b polyplexes were more stable than H3K4b polyplexes. Lipopolyplexes formed with H3K(+H)4b markedly increased mRNA transfection.

Targeting the KDM4B-AR-c-Myc axis promotes sensitivity to androgen receptor-targeted therapy in advanced prostate cancer.

The histone demethylase KDM4B functions as a key co-activator for the androgen receptor (AR) and plays a vital in multiple cancers through controlling gene expression by epigenetic regulation of H3K9 methylation marks. Constitutively active androgen receptor confers anti-androgen resistance in advanced prostate cancer. However, the role of KDM4B in resistance to next-generation anti-androgens and the mechanisms of KDM4B regulation are poorly defined. Here we found that KDM4B is overexpressed in enzalutamide-resistant prostate cancer cells. Overexpression of KDM4B promoted recruitment of AR to the c-Myc (MYC) gene enhancer and induced H3K9 demethylation, increasing AR-dependent transcription of c-Myc mRNA, which regulates the sensitivity to next-generation AR-targeted therapy. Inhibition of KDM4B significantly inhibited prostate tumor cell growth in xenografts, and improved enzalutamide treatments through suppression of c-Myc. Clinically, KDM4B expression was found upregulated and to correlate with prostate cancer progression and poor prognosis. Our results revealed a novel mechanism of anti-androgen resistance via histone demethylase alteration which could be targeted through inhibition of KDM4B to reduce AR-dependent c-Myc expression and overcome resistance to AR-targeted therapies. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

A CRISPR Test for Rapidly and Sensitively Detecting Circulating EGFR Mutations.

The detection of EGFR mutations in circulating cell-free DNA can enable personalized therapy for cancer. The current techniques for detecting circulating EGFR mutations are expensive and time-consuming with moderate sensitivity. Emerging CRISPR is revolutionizing medical diagnostics and showing a great promise for nucleic acid detection. This study aims to develop CRISPR-Cas12a as a simple test to sensitively detect circulating EGFR mutations in plasma. Serially diluted samples of DNA containing heterozygous EGFR mutations (L858R and T790M) in wild-type genomic DNA are concurrently tested for the mutations by a CRISPR-Cas12a system and droplet digital PCR (ddPCR). The CRISPR-Cas12a system can detect both L858R and T790M with a limit of detection of 0.005% in less than three hours. ddPCR detects the mutations with a limit of detection of 0.05% for more than five hours. Plasma samples of 28 lung cancer patients and 20 cancer-free individuals are tested for the EGFR mutations by CRISPR-Cas12a system and ddPCR. The CRISPR-Cas12a system could detect L858R in plasma of two lung cancer patients whose tissue biopsies are positive for L858R, and one plasma sample of three lung cancer patients whose tissue biopsies are positive for T790M. ddPCR detects L858R in the same two plasm samples, however, does not detect T790M in any of the plasma samples. This proof of principle study demonstrates that the CRISPR-Cas12a system could rapidly and sensitively detect circulating EGFR mutations, and thus, has potential prognostic or therapeutic implications.

MicroRNA-based biomarkers for diagnosis of non-small cell lung cancer (NSCLC).

BACKGROUND: The development of biomarkers for the early detection of non-small cell lung cancer (NSCLC) is clinically important. We have developed miRNA biomarkers in sputum and plasma, respectively, for NSCLC. Herein, we evaluate whether integrated analysis of the miRNAs across the different types of specimens could improve the early detection of NSCLC. METHODS: Using reverse transcription PCR, we determined expressions of two miRNAs (miRs-31-5p and 210-3p) in sputum and three miRNAs (miRs-21-5p, 210-3p, and 486-5p) in plasma of a training cohort of 76 NSCLC patients and 72 cancer-free smokers. The results were validated in a testing cohort of 56 NSCLC patients and 55 cancer-free smokers. RESULTS: The panels of two sputum miRNAs and three plasma miRNAs had 65.8-75.0% sensitivities and 83.3-87.5% specificities for diagnosis of NSCLC in the training cohort. The individual sputum or plasma miRNA panel had a higher sensitivity for squamous cell carcinoma or adenocarcinoma of the lung, respectively. From the miRNAs, we optimized an integrated panel of biomarkers consisting of two sputum miRNAs (miRs-31-5p and 210-3p) and one plasma miRNA (miR-21-5p) that had higher sensitivity (85.5%) and specificity (91.7%) for diagnosis of NSCLC compared with the individual panels alone. Furthermore, the performance of the integrated panel of biomarkers was independent of histology and stage of NSCLC, and patients' age, sex, and ethnicity. The performance of the integrated panel of biomarkers was confirmed in the testing cohort. CONCLUSIONS: Integrating biomarkers across different body fluids would synergistically improve the early detection of NSCLC. KEY POINTS: Lung cancer is a heterogeneous disease and develops from complex aberrations. Integrating sputum and plasma miRNAs has higher accuracy than when they are used alone.

A CRISPR Test for Detection of Circulating Nuclei Acids.

Emerging CRISPR-based nucleic acid detection shows great promise in molecular diagnosis of diseases. CRISPR-Cas12a can sensitively and specifically detect human papillomavirus (HPV) DNA in anal swabs. However, the current CRISPR-Cas12a system needs auxiliary and expensive equipment, which limit its application as a point-of-care (POC) diagnostic tool. This study aimed to develop CRISPR-Cas12a as a POC test to directly target plasma for circulating HPV DNA detection by immediately reading results with naked eyes. Cell-cultured supernatants of either HPV16- or 18-positive cancer cells were treated with lysis buffer followed by isothermal amplification without DNA isolation. Cas12a, crRNA, and fluorescent-biotin reporters were incubated with the lysates. Our data showed that integrating CRISPR-Cas12a with lateral-flow strips could directly and specifically detect HPV16 and 18 in the liquid samples with the same limit of detection (0.24 fM) as did polymerase chain reaction but requiring less time. Furthermore, the CRISPR-Cas12a system could rapidly detect presence of HPV16 and HPV18 in plasma samples of 13 of 14 and 3 of 10 the patients with histopathological diagnosis of cervical cancer, respectively. Therefore, a CRISPR-Cas12a-based POC system was developed for conveniently detecting circulating nuclei acid targets in body fluids without requiring technical expertise and ancillary machineries.

Targeting Cancer with Peptide RNAi Nanoplexes.

With the recent explosion of genomic information on the root causes of disease, there is an increased interest in nucleic acid therapeutics, including siRNA and gene therapy, all of which require delivery of highly charged nucleic acids from siRNA with a molecular weight of about 1.4 × 104 to plasmids with an approximate molecular weight of 2.0-3.0 × 106. This chapter describes the delivery of shRNA via plasmid or siRNA with a peptide-based carrier. We focus on the histidine-lysine peptide which serves as an example for other peptides and polymeric carrier systems. When the HK peptide and nucleic acids are mixed together and interact with one another through ionic and nonionic interactions, nanoplexes are formed. These nanoplexes, carrying either shRNA or siRNA that target oncogenes, provide promising options for the treatment of cancer. We describe methods of preparation and characterization of these nanoplexes using dynamic light scattering, zeta potential, and gel retardation assays. We also provide protocols for transfection in vitro and in vivo for these nanoplexes.

Enhanced tumor uptake and activity of nanoplex-loaded doxorubicin.

Doxorubicin (Dox) has widespread use as a cancer chemotherapeutic agent, but Dox is limited by several side effects including irreversible cardiomyopathy. Although liposomal Dox formulations, such as Doxil, mitigate side effects, they do not prolong survival in many patients. As a result, efforts have continued to discover improved formulations of Dox. We previously found that a peptide-based nanoplex delivered plasmid DNA efficiently to tumors in murine models. Unlike the majority of nanoparticles that depend solely on enhanced permeability and retention (EPR) for their transport into the tumor, our peptide-based nanoplex has a potential advantage in that its uptake primarily depends on neuropilin-1 receptor targeting. Because Dox binds to DNA, we tested whether this delivery platform could effectively deliver Dox to tumors and reduce their size. The nanoplexes increased the levels of Dox in tumors by about 5.5-fold compared to aqueous (free) Dox controls. Consistent with enhanced levels in the tumor, the nanoplex-Dox treatment had significantly greater anti-tumor activity. Whereas low dose free Dox did not reduce the size of tumors compared to untreated controls, the low dose nanoplex-Dox reduced the size of tumors by nearly 55% (p < 0.001). The high dose nanoplex-Dox also inhibited the size of tumor significantly more than the comparable high-dose free Dox (p < 0.001). Furthermore, apoptosis and proliferation markers (Ki67) of tumors observed in the different treatment groups correlated with their ability to inhibit tumor size. This study shows the efficacy of an NRP-1 targeted nanoplexes to deliver Dox to tumors in vivo and lays the groundwork for more complex and effective formulations.

A Plasma Long Noncoding RNA Signature for Early Detection of Lung Cancer.

The early detection of lung cancer is a major clinical challenge. Long noncoding RNAs (lncRNAs) have important functions in tumorigenesis. Plasma lncRNAs directly released from primary tumors or the circulating cancer cells might provide cell-free cancer biomarkers. The objective of this study was to investigate whether the lncRNAs could be used as plasma biomarkers for early-stage lung cancer. By using droplet digital polymerase chain reaction, we determined the diagnostic performance of 26 lung cancer-associated lncRNAs in plasma of a development cohort of 63 lung cancer patients and 33 cancer-free individuals, and a validation cohort of 39 lung cancer patients and 28 controls. In the development cohort, 7 of the 26 lncRNAs were reliably measured in plasma. Two (SNHG1 and RMRP) displayed a considerably high plasma level in lung cancer patients vs. cancer-free controls (all P < .001). Combined use of the plasma lncRNAs as a biomarker signature produced 84.13% sensitivity and 87.88% specificity for diagnosis of lung cancer, independent of stage and histological type of lung tumor, and patients' age and sex (all P > .05). The diagnostic value of the plasma lncRNA signature for lung cancer early detection was confirmed in the validation cohort. The plasma lncRNA signature may provide a potential blood-based assay for diagnosing lung cancer at the early stage. Nevertheless, a prospective study is warranted to validate its clinical value.