The emergence and global spread of ceftriaxone-resistant Neisseria gonorrhoeae underscores the urgent need for an effective vaccine. Gonococcal outer membrane vesicles (OMVs) are promising as vaccine platform, but their efficacy is potentially compromised by the immunomodulatory properties of gonococcal surface-expressed proteins, particularly the essential outer membrane porin PorB. Our previous work identified that gonococcal OMVs induce epithelial cell mitophagy via PorB, dependent on lysine residues 117 and 171. Given the critical role of dendritic cells (DCs) in initiating adaptive immunity, this study investigated whether PorB-mediated mitophagy in DCs impacts OMV vaccine efficacy. Here, we demonstrated that gonococcal OMVs induce DC mitophagy in a PorB-dependent manner, a process abolished in OMVs expressing the mitophagy-deficient PorB mutant PorBK117Q/K171Q. OMVs expressing PorBK117Q/K171Q demonstrated significantly enhanced DC activation, as shown by increased CD86 and MHC-II expression, and promoted a Th1-skewed T cell response with elevated IFN-γ and TNF-α secretion. In immunized mice, OMVs containing PorBK117Q/K171Q elicited significantly higher total IgG and IgG2a antibody titres against PorB compared with OMVs expressing wild-type PorB, with antibodies displaying enhanced bactericidal activity, including against a strain associated with the high-level ceftriaxone-resistant FC428 clone. Importantly, the OMV PorBK117Q/K171Q vaccine provided enhanced protection in a mouse vaginal colonization model, accelerating bacterial clearance and reducing overall bacterial burden. Therefore, our results identify PorB-induced mitophagy in DCs as a potential immune evasion mechanism that may dampen adaptive immunity. Engineering OMV vaccines to circumvent this process represents a rational strategy to explore for enhancing gonococcal vaccine efficacy.

Emergence of ceftriaxone-resistant Neisseria gonorrhoeae strains expressing penA allele 60.001 poses a major threat to the efficacy of ceftriaxone-based therapies. This study in Hangzhou, China, investigated the molecular epidemiology and antimicrobial susceptibility of 479 N. gonorrhoeae isolates collected from 2019 to 2025 to track the evolution of this resistance. Antimicrobial susceptibility of N. gonorrhoeae isolates was determined by agar dilution method. Sequence types were identified by multi-locus sequence typing (MLST). Whole-genome sequencing and phylogenetic analysis were performed on high-level ceftriaxone-resistant isolates. While overall ceftriaxone resistance fluctuated, a significant and alarming resurgence of high-level resistance (MIC ≥ 0.5 mg/L) was observed in 2024 (35

Neisseria gonorrhoeae is a global public health threat, exacerbated by the rapid dissemination of ceftriaxone-resistant strains harboring penA allele 60.001. Current diagnostics are limited by long turnaround times, reliance on thermal cycling equipment, and poor suitability for resource-limited settings. We developed an isothermal recombinase polymerase amplification assay coupled with a lateral flow strip (RPA-LFS) for separate identification of N. gonorrhoeae and the ceftriaxone resistance-associated SNP C932T (A311V) in penA 60.001 and limited closely related alleles. Species detection employed an RPA-Nfo probe targeting the conserved porA pseudogene. Resistance detection employed an allele-specific amplification refractory mutation system RPA (ARMS-RPA) strategy, incorporating artificial mismatches to specifically discriminate the C932T SNP. Assay performance was evaluated using genomic DNA of a panel of gonococcal and non-gonococcal strains and validated directly on unprocessed clinical urine samples. The porA RPA-LFS assay demonstrated a detection limit of 30 fg of genomic DNA with no cross-reactivity against closely related Neisseria species or other bacterial pathogens. The mismatch penA 60.001 RPA-LFS assay achieved a detection limit of 300 fg and exhibited absolute analytical specificity, with no amplification observed from wild-type or other mosaic penA alleles. Validation using 50 clinical urine samples yielded 100

Neisseria gonorrhoeae imposes a substantial global health burden due to its high incidence and escalating multidrug resistance. This study investigated the immunogenicity and efficacy of a peptide-based vaccine and a monoclonal antibody (mAb) targeting the conserved Loop2 epitope of the outer membrane protein MtrE. Two multiple antigenic peptide (MAP) vaccines, displaying four copies of MtrE Loop2 with or without a Cathepsin S cleavage site, were formulated with CpG1826 adjuvant. Immunization of mice elicited robust Loop2-specific IgM-dominant antibody responses with complement-dependent anti-gonococcal serum bactericidal activity. In a murine vaginal tract infection model, both vaccines demonstrated significant prophylactic and single-dose therapeutic efficacy. Furthermore, a human-mouse chimeric mAb (M01), consisting of mouse variable domains and human IgG1 constant domains, was generated from a dominant B-cell clonotype obtained from MAP vaccine-immunized mice. M01 exhibited high-affinity binding to MtrE and potent complement-dependent bactericidal activity. In a murine infection model, intravaginal administration of M01 significantly enhanced gonococcal clearance. Furthermore, Fc-engineered M01 variants confirmed that this efficacy was critically dependent on complement activity. These findings identify MtrE Loop2 as a promising target for both active and passive immunization strategies against N. gonorrhoeae, and underscore the critical role of complement-mediated activity as a mechanistic correlate of protection.

AIMS:This study explored botanical extracts to guide discovery of alternative antimicrobials with activity against the multidrug-resistant bacterial pathogen Neisseria gonorrhoeae. MATERIALS AND METHODS:In vitro antimicrobial activity was analyzed by agar dilution method and time-kill assays, while in vivo activity was evaluated in a mouse infection model. The antimicrobial target was identified by pull-down assays and validated by interaction studies with wild-type and targeted mutant proteins. KEY FINDINGS:The extract of Chinese Populus spp. propolis (CPP) displayed the most potent antigonococcal activity. Subsequently, galangin was identified as the most active compound in CPP. However, galangin showed no in vivo activity in the mouse infection model. Screening for active structural analogues of galangin resulted in the identification of 2-hydroxychalcone (2-HC), which showed a minimum inhibitory concentration (MIC) of 8-16 μM, and enhanced gonococcal clearance in the infection model. Pull-down experiments identified EF-Tu and butanediol dehydrogenase (Bdh) as putative 2-HC targets. Further binding analysis of 2-HC with recombinant purified proteins showed an equilibrium dissociation constant (KD) of 1.28 μM for EF-Tu and 5.97 μM for Bdh. Finally, 2-HC binding pockets on EF-Tu and Bdh were identified by molecular docking studies showing that 2-HC interacted with Glu260 of EF-Tu and Ser273 of Bdh, which was validated by mutagenesis studies. For Bdh, we furthermore demonstrated that 2-HC impacted its enzyme activity with and IC50 of 44-64 μM, resulting in perturbed NAD+/NADH ratios. SIGNIFICANCE:2-HC displays a bimodal antigonococcal mechanism, making it an interesting candidate for further development as antigonococcal therapy.

OBJECTIVES:Neisseria gonorrhoeae strains associated with the high-level ceftriaxone-resistant FC428 clone or containing its main resistance determinant, penA allele 60.001, have shown global transmission. In Hangzhou, China, 10% of the isolates were associated with the FC428 clone in 2019. Here, we investigated ceftriaxone resistance and the prevalence of FC428-associated strains in Hangzhou in 2020-22. METHODS:A total of 209 gonococcal isolates were investigated for antimicrobial susceptibility to ceftriaxone and other antibiotics by agar dilution method. Sequence types and penA alleles were determined by PCR and sequence analysis. RESULTS:Resistance to ceftriaxone (MIC > 0.125 mg/L) was observed for 16% (33/209) of the isolates, whereas 6.7% (14/209) of the isolates displayed high-level ceftriaxone resistance (MIC = 1 mg/L). These 14 high-level ceftriaxone-resistant isolates and another isolate displaying an MIC = 0.25 mg/L contained penA allele 60.001, with eight of these isolates, all from 2020 to 2021 belonging to MLST ST1903, the sequence type commonly associated with the original FC428 clone. Importantly, the six penA allele 60.001-containing isolates from 2022 belonged to MLST ST8123, ST7365 and ST7367, which are among the most frequently encountered sequence types found in China. Therefore, these results indicate that endemic lineages in China have acquired penA allele 60.001. CONCLUSIONS:Here, we report continued transmission of gonococcal strains associated with the FC428 clone or containing penA allele 60.001 in Hangzhou. A major concern for public health is the acquisition of penA allele 60.001 by successful endemic lineages, which might enhance the transmission of this high-level ceftriaxone resistance trait.

The bacterial pathogen Neisseria gonorrhoeae is able to invade epithelial cells and survive intracellularly. During this process, it secretes outer membrane vesicles (OMVs), however, the mechanistic details for interactions between gonococcal OMVs and epithelial cells and their impact on intracellular survival are currently not established. Here, we show that gonococcal OMVs induce epithelial cell mitophagy to reduce mitochondrial secretion of reactive oxygen species (ROS) and enhance intracellular survival. We demonstrate that OMVs deliver PorB to mitochondria to dissipate the mitochondrial membrane potential, resulting in mitophagy induction through a conventional PINK1 and OPTN/NDP52 mechanism. Furthermore, PorB directly recruits the E3 ubiquitin ligase RNF213, which decorates PorB lysine residue 171 with K63-linked polyubiquitin to induce mitophagy in a p62-dependent manner. These results demonstrate a mechanism in which polyubiquitination of a bacterial virulence factor that targets mitochondria directs mitophagy processes to this organelle to prevent its secretion of deleterious ROS.

ABSTRACT Neisseria gonorrhoeae is a multidrug-resistant bacterial pathogen for which ceftriaxone mono-antimicrobial therapy and ceftriaxone plus azithromycin dual-antimicrobial therapy are the only remaining effective therapies. However, ceftriaxone efficacy is threatened by the global dissemination of the high-level ceftriaxone-resistant FC428 clone, while azithromycin has in recent years been removed from the recommended dual therapy in some countries as a result of increasing (high-level) azithromycin resistance. Inclusion of an alternative second antimicrobial could be an effective strategy to protect ceftriaxone from (further) resistance development. In this study, we investigated gonococcal susceptibility to bacitracin and determined its synergistic anti-gonococcal activity with ceftriaxone. Gonococcal susceptibility to bacitracin was investigated for 449 contemporary clinical isolates using the agar dilution method. Bacitracin displayed consistent activity against the N. gonorrhoeae strain collection, with a minimum inhibitory concentration (MIC) range of 2–32 mg/L and a MIC90 of 32 mg/L. Furthermore, bacitracin was bactericidal in time-kill assays, particularly against the tested FC428-associated isolate, which showed 105-fold inactivation after 8 hours of exposure to a 4× MIC dose. Importantly, bacitracin and ceftriaxone displayed synergistic activity, with a fractional inhibitory concentration index ≤0.5 for the majority of tested strains. Finally, spot assays showed strong synergistic activity against the FC428-associated isolates, with up to 200-fold reduced plating efficacy for bacitracin combined with ceftriaxone compared with ceftriaxone only. In conclusion, bacitracin shows synergistic anti-gonococcal activity with ceftriaxone and might therefore be an interesting candidate for inclusion in a dual-antimicrobial therapy with ceftriaxone. IMPORTANCE Ceftriaxone-based antimicrobial therapies for gonorrhea are threatened by waning ceftriaxone susceptibility levels and the global dissemination of the high-level ceftriaxone-resistant gonococcal FC428 clone. Combination therapy can be an effective strategy to restrain the development of ceftriaxone resistance, and for that purpose, it is important to find an alternative antimicrobial to replace azithromycin, which has recently been removed in some countries from the recommended ceftriaxone plus azithromycin dual-antimicrobial therapy. Ideally, the second antimicrobial should display synergistic activity with ceftriaxone. We hypothesized that bacitracin might display synergistic activity with ceftriaxone because of their distinct mechanisms targeting bacterial cell wall synthesis. In this study, we showed that bacitracin indeed displays synergistic activity with ceftriaxone against Neisseria gonorrhoeae. Importantly, strains associated with the FC428 clone appeared to be particularly susceptible to the bacitracin plus ceftriaxone combination, which might therefore be an interesting dual therapy for further in vivo testing.

Neisseria gonorrhoeae establishes tight interactions with mucosal epithelia through activity of its type IV pilus, while pilus retraction forces activate autophagic responses toward invading gonococci. Here we studied pilus-independent epithelial cell responses and showed that pilus-negative gonococci residing in early and late endosomes are detected and targeted by nucleotide-binding oligomerization domain 1 (NOD1). NOD1 subsequently forms a complex with immunity-related guanosine triphosphatase M (IRGM) and autophagy-related 16-like 1 (ATG16L1) to activate autophagy and recruit microtubule-associated protein light chain 3 (LC3) to the intracellular bacteria. IRGM furthermore directly recruits syntaxin 17 (STX17), which is able to form tethering complexes with the lysosome. Importantly, IRGM-STX17 interactions are enhanced by LC3 but were still observed at lower levels in an LC3 knockout cell line. These findings demonstrate key roles for NOD1 and IRGM in the sensing of intracellular N gonorrhoeae and subsequent directing of the bacterium to the lysosome for degradation. This study shows that NOD1 detects intracellular Neisseria gonorrhoeae in epithelial cells and recruits IRGM and ATG16L1 to direct LC3 to the bacterial endosomes. IRGM subsequently recruits STX17 for the formation of tethering complexes with lysosomes to restrict intracellular survival.

ABSTRACT Global dissemination of high-level ceftriaxone-resistant Neisseria gonorrhoeae strains associated with the FC428 clone poses a threat to the efficacy ceftriaxone-based therapies. Vaccination is the best strategy to contain multidrug-resistant infections. In this study, we investigated the efficacy of MtrE and its surface Loop2 as vaccine antigens when combined with a Th1-polarizing adjuvant, which is expected to be beneficial for gonococcal vaccine development. Using in vitro dendritic cell maturation and T cell differentiation assays, CpG1826 was identified as the optimal Th1-polarizing adjuvant for MtrE and Loop2 displayed as linear epitope (Nloop2) or structural epitope (Intraloop2) on a carrier protein. Loop2-based antigens raised strongly Th1-polarized and bactericidal antibody responses in vaccinated mice. Furthermore, the vaccine formulations provided protection against a gonococcal challenge in mouse vaginal tract infection model when provided as prophylactic vaccines. Also, the vaccine formulations accelerated gonococcal clearance when provided as a single therapeutic dose to treat an already established infection, including against a strain associated with the FC428 clone. Therefore, this study demonstrated that MtrE and Loop 2 are effective gonococcal vaccine antigens when combined with the Th1-polarizing CpG1826 adjuvant.

OBJECTIVES Ceftriaxone therapy for gonorrhoea has become under increasing pressure due to waning susceptibility levels and emergence of high-level resistant strains such as the FC428 clone. Moenomycin was recently identified to display potent anti-gonococcal activity against some reference strains. Therefore, the aim of this study was to investigate moenomycin in vitro and in vivo antimicrobial activity. METHODS Moenomycin in vitro antimicrobial activity was investigated against 575 clinical isolates, including strains associated with the FC428 clone, using the agar dilution method. Moenomycin in vivo activity was investigated in a mouse vaginal tract gonococcal infection model. RESULTS The moenomycin MIC range for the strain collection was 0.004-0.06 mg/L, with a MIC50 of 0.016 mg/L and a MIC90 of 0.03 mg/L. The correlation between moenomycin and ceftriaxone susceptibility levels was poor (R = 0.13), while the fractional inhibitory concentration index (FICI) resulted in indifference for all tested strains. Therefore, development of cross-resistance between moenomycin and ceftriaxone is unlikely for N. gonorrhoeae. Determination of the moenomycin mode of activity against N. gonorrhoeae by time-kill assays showed that moenomycin is bactericidal, with over 104-fold inactivation observed after 4 h exposure. Finally, an intramuscular moenomycin dose of 10 mg/kg given on 2 consecutive days was able to clear a gonococcal infection in a mouse vaginal tract infection model within 1-3 days after the second dose, which was significantly faster than for mice treated with the vehicle control (P < 0.0001). CONCLUSIONS Moenomycin displays potent in vitro and in vivo antimicrobial activity against N. gonorrhoeae, warranting further exploration as alternative therapy.

Alternative antimicrobial therapies are urgently required for the multidrugresistant bacterial pathogen Neisseria gonorrhoeae, for which currently ceftriaxone is the only remaining recommended first-line therapy. Repurposing of drugs that are approved for other clinical applications offers an efficient approach for development of alternative antimicrobial therapies. Auranofin, cannabidivarin, and tolfenamic acid were recently identified to display antimicrobial activity against N. gonorrhoeae. Here, we investigated their activity against a collection of 575 multidrug-resistant clinical isolates. All three compounds displayed consistent antimicrobial activity against all isolates, including against strains associated with the high-level ceftriaxone-resistant FC428 clone, with both the mode and MIC90 for auranofin of 0.5 mg/L, while both the mode and MIC90 for cannabidivarin and tolfenamic acid were 8 mg/L. Correlations between MICs of ceftriaxone and auranofin, cannabidivarin or tolfenamic acid were low, indicating that development of cross-resistance is unlikely. Furthermore, antimicrobial synergy analysis between ceftriaxone and auranofin, cannabidivarin, or tolfenamic acid by determination of the fractional inhibitory concentration index (FICI) resulted in an interpretation of indifference. Finally, time-kill analyses showed that all three compounds are bactericidal against both the N. gonorrhoeae ATCC 49226 reference strain and an FC428-associated clinical isolate, with particularly cannabidivarin displaying rapid bactericidal activity. Overall, auranofin, cannabidivarin, and tolfenamic acid displayed consistent antimicrobial activity against multidrug-resistant N. gonorrhoeae, warranting further exploration of their suitability as alternative antimicrobials for treatment of gonococcal infections. IMPORTANCE Neisseria gonorrhoeae is a major public health concern because of the high incidence of gonorrhea and the increasingly limited options for antimicrobial therapy. Strains associated with the FC428 clone are a particular concern because they have shown global dissemination and they display high-level resistance against the currently recommended ceftriaxone therapy. Therefore, development of alternative antimicrobial therapies is urgently required to ensure treatment of gonorrhea remains available in the future. Repurposing of clinically approved drugs could be a rapid approach for the development of such alternative antimicrobials. In this study, we showed that repurposing of auranofin, cannabidivarin, and tolfenamic acid for antimicrobial therapy of gonorrhea deserves further clinical explorations because these compounds displayed consistent antimicrobial activity against a large collection of contemporary multidrug-resistant gonococcal isolates that included strains associated with the FC428 clone.

Abstract Background Staphylococcus aureus is a leading cause for morbidity and mortality associated with skin and burn wound infections. Therapeutic options for methicillin-resistant S. aureus (MRSA) have dwindled and therefore alternative treatments are urgently needed. In this study, the immuno-stimulating and anti-MRSA effects of cyclic di-guanosine monophosphate (c-di-GMP), a uniquely bacterial second messenger and immuno-modulator, were investigated in HaCaT human epidermal keratinocytes and a murine skin wound infection model. Results Stimulation of HaCaT cells with 125 μM c-di-GMP for 12 h prior to MRSA challenge resulted in a 20-fold reduction in bacterial colonization compared with untreated control cells, which was not the result of a direct c-di-GMP toxic effect, since bacterial viability was not affected by this dose in the absence of HaCaT cells. C-di-GMP-stimulated or MRSA-challenged HaCaT cells displayed enhanced secretion of the antimicrobial peptides human β-defensin 1 (hBD-1), hBD-2, hBD-3 and LL-37, but for hBD1 and LL-37 the responses were additive in a c-di-GMP-dose-dependent manner. Secretion of the chemokines CXCL1 and CXCL8 was also elevated after stimulation of HaCaT cells with lower c-di-GMP doses and peaked at a dose of 5 μM. Finally, pre-treatment of mice with a 200 nmol dose of c-di-GMP 24 h before a challenge with MRSA in skin wound infection model resulted in a major reduction (up to 1,100-fold by day 2) in bacterial CFU counts recovered from challenged skin tissue sections compared PBS-treated control animals. Tissue sections displayed inflammatory cell infiltration and enhanced neutrophil influx in the c-di-GMP pre-treated animals, which might account for the reduced ability of MRSA to colonize c-di-GMP pre-treated mice. Conclusions These results demonstrate that c-di-GMP is a potent immuno-modulator that can stimulate anti-MRSA immune responses in vivo and might therefore be a suitable alternative prophylactic or therapeutic agent for MRSA skin or burn wound infections.

Klebsiella pneumoniae is a gram-negative bacterium that can cause many diseases in hospitals and communities. Intestinal K. pneumoniae infections are relatively rare. Most K. pneumoniae infections begin with the colonization of the gastrointestinal system. In this study, clinically isolated K. pneumoniae strains were used to infect intestinal epithelial Caco-2 cells to study the possible intestinal translocation mechanism of K. pneumoniae. We found that of the three K. pneumoniae strains tested, KP1821 exhibited the strongest adhesive and invasive abilities and that the adhesion to Caco-2 intestinal epithelial cells was affected by the acidic environment of the stomach. Transcriptome sequencing revealed the involvement of molecules associated with the extracellular matrix and cell adhesion, inflammatory response, calcium ion and transforming growth factor β (TGF-β) signaling pathways, and other abnormalities in biological processes and cell signaling pathways. Additionally, tolloid-like protein 1 (TLL1) was significantly upregulated. Knocking down TLL1 with shRNA significantly reduced KP1821's ability to invade and adhere to intestinal epithelial cells. TLL1 is involved in the activation of the TGF-β signaling pathway. Inhibition of this pathway using the inhibitor SB431542 induced significantly reduced adhesion and invasion capabilities of KP1821. Our findings demonstrate that TLL1 participates in K. pneumoniae adhesion and invasion of intestinal epithelial cells by activating the TGF-β signaling pathway.

Objective:To investigate the possible mechanism of high mobility group box-1 (HMGB1) in amplifing inflammatory responses in Leptospira interrogans hemolysin Sph2-treated J774A.1 macrophages. Methods:Recombinant Sph2 was incubated with J774A.1 macrophages. The damage of cell membrane was detected by lactate dehydrogenase(LDH) determination; the changes of cell structure were observed by cryo-electron microscope; ELISA was used to determine the expression of HMGB1. After the commercial recombinant HMGB1 was incubated with mouse J774A.1 macrophages, the phosphorylation of NF-κB, p38-MAPK and JNK signaling pathway wsa detected by Western blot, and the expression of IL-1β, IL-6, and KC (IL-8) was detected by ELISA.Results:Recombinant hemolysin rSph2 induced significant changes in the structures of J774A.1 cells, including nucleus disappearance, cell membrane structure damage, cell lysis and membrane swelling. The yields of LDH and HMGB1 also increased significantly. Phosphorylated-NF-κB, -p38-MAPK and -JNK were increased by HMGB1. The expression of IL-1β, IL-6 and KC in J774A.1 cells was up-regulated by HMGB1 and inhibited via inhibitors of NF-κB, p38-MAPK and JNK signal pathways.Conclusions:Hemolysin rSph2 damaged the membrane of J774A.1 cells, and induced the secretion of HMGB1. Secreted-HMGB1 might induce the expression of IL-1β, IL-6 and KC in J774A.1 cells via NF-κB, p38-MAPK and JNK signal pathways, thus amplifying the inflammatory responses caused by Sph2.

The nucleotide-binding domain, leucine-rich-repeat containing family, pyrin domain-containing 3 (NLRP3) inflammasome is essential in inflammation and inflammatory disorders. Phosphorylation at various sites on NLRP3 differentially regulates inflammasome activation. The Ser725 phosphorylation site on NLRP3 is depicted in multiple inflammasome activation scenarios, but the importance and regulation of this site has not been clarified. The present study revealed that the phosphorylation of Ser725 was an essential step for the priming of the NLRP3 inflammasome in macrophages. We also showed that Ser725 was directly phosphorylated by misshapen (Msn)/NIK-related kinase 1 (MINK1), depending on the direct interaction between MINK1 and the NLRP3 LRR domain. MINK1 deficiency reduced NLRP3 activation and suppressed inflammatory responses in mouse models of acute sepsis and peritonitis. Reactive oxygen species (ROS) upregulated the kinase activity of MINK1 and subsequently promoted inflammasome priming via NLRP3 Ser725 phosphorylation. Eliminating ROS suppressed NLRP3 activation and reduced sepsis and peritonitis symptoms in a MINK1-dependent manner. Altogether, our study reveals a direct regulation of the NLRP3 inflammasome by Msn family kinase MINK1 and suggests that modulation of MINK1 activity is a potential intervention strategy for inflammasome-related diseases.

Objective The aim of the study is to explore the prognosis value of PTPRH in patients with lung adenocarcinoma (LUAD). Methods Oncomine, UALCAN, and GEPIA databases were employed to examine the differential expression of PTPRH between LUAD and adjacent tissues. 100 pairs of LUAD and adjacent tissue samples were involved in this study. qRT-PCR and immunohistochemical staining were performed. Meanwhile, we analyzed The Cancer Genome Atlas (TCGA) data to investigate the correlation between PTPRH gene expression and clinicopathological characteristics. Kaplan-Meier analysis and univariate and multivariate Cox analyses were performed to estimate the relationship between PTPRH expression and LUAD prognosis. The evaluation performance was verified by drawing a ROC curve. In addition, through GSEA, the changes of PTPRH expression were analyzed by GSEA to screen out primarily affected signaling pathway. Results Oncomine, UALCAN, and GEPIA databases showed that the mRNA expression of PTPRH in LUAD tissues was significantly higher than that in adjacent tissues. qRT-PCR and immunohistochemical staining indicated the mRNA and protein levels of PTPRH in LUAD tissues were markedly upregulated. TCGA data showed that the expression of PTPRH was significantly correlated with T stage and disease stage. Kaplan-Meier analysis showed that the patients with high PTPRH expression had a poor prognosis. Univariate and multivariate Cox analyses exhibited that PTPRH expression could act as an independent prognostic factor for LUAD. The ROC curve showed that PTPRH combined with various clinicopathological features could effectively predict the prognosis of LUAD. Finally, GSEA indicated that changes in PTPRH expression level may affect p53, VEGF, Notch, and mTOR cancer-related signaling pathways. Conclusion Our results demonstrated that PTPRH was highly expressed in LUAD and may be closely correlated with the poor prognosis of LUAD patients.