Folate Receptor Research Articles

Proteomics analysis of extracellular vesicles for biomarkers of autism spectrum disorder

BackgroundAutism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by symptoms that include social interaction deficits, language difficulties and restricted, repetitive behavior. Early intervention through medication and behavioral therapy can eliminate some ASD-related symptoms and significantly improve the life-quality of the affected individuals. Currently, the diagnosis of ASD is highly limited.MethodsTo investigate the feasibility of early diagnosis of ASD, we tested extracellular vesicles (EVs) proteins obtained from ASD cases. First, plasma EVs were isolated from healthy controls (HCs) and ASD individuals and were analyzed using proximity extension assay (PEA) technology to quantify 1,196 protein expression level. Second, machine learning analysis and bioinformatic approaches were applied to explore how a combination of EV proteins could serve as biomarkers for ASD diagnosis.ResultsNo significant differences in the EV morphology and EV size distribution between HCs and ASD were observed, but the EV number was slightly lower in ASD plasma. We identified the top five downregulated proteins in plasma EVs isolated from ASD individuals: WW domain-containing protein 2 (WWP2), Heat shock protein 27 (HSP27), C-type lectin domain family 1 member B (CLEC1B), Cluster of differentiation 40 (CD40), and folate receptor alpha (FRalpha). Machine learning analysis and correlation analysis support the idea that these five EV proteins can be potential biomarkers for ASD.ConclusionWe identified the top five downregulated proteins in ASD EVs and examined that a combination of EV proteins could serve as biomarkers for ASD diagnosis.

Immune-based subgroups uncover diverse tumor immunogenicity and implications for prognosis and precision therapy in acute myeloid leukemia

BackgroundAlthough a considerable proportion of acute myeloid leukemia (AML) patients achieve remission through chemotherapy, relapse remains a recurring and significant event leading to treatment failure. This study aims to investigate the immune landscape in AML and its potential implications for prognosis and chemo-/immune-therapy.MethodsIntegrated analyses based on multiple sequencing datasets of AML were performed. Various algorithms estimated immune infiltration in AML samples. A subgroup prediction model was developed, and comprehensive bioinformatics and machine learning algorithms were applied to compare immune-based subgroups in relation to clinical features, mutational landscapes, immune characterizations, drug sensitivities, and cellular hierarchies at the single-cell level.ResultsTwo immune-based AML subgroups, G1 and G2, were identified. G1 demonstrated higher immune infiltration, a more monocytic phenotype, increased proportions of monocytes/macrophages, and higher FLT3, DNMT3A, and NPM1 mutation frequencies. It was associated with a poorer prognosis, lower proportions of various immune cell types and a lower T cell infiltration score (TIS). AML T-cell-based immunotherapy target antigens, including CLEC12A, Folate receptor β, IL1RAP and TIM3, showed higher expression levels in G1, while CD117, CD244, CD96, WT and TERT exhibited higher expression levels in G2. G1 samples demonstrated higher sensitivity to elesclomol and panobinostat but increased resistance to venetoclax compared to G2 samples. Moreover, we observed a positive correlation between sample immune infiltration and sample resistance to elesclomol and panobinostat, whereas a negative correlation was found with venetoclax resistance.ConclusionOur study enriches the current AML risk stratification and provides guidance for precision medicine in AML.

Deciphering Folate Receptor alphaGene Expression and mRNA Signatures in Ovarian Cancer: Implications for Precision Therapies

Deciphering Folate Receptor alphaGene Expression and mRNA Signatures in Ovarian Cancer: Implications for Precision Therapies

Folate Receptor Alpha—A Secret Weapon in Ovarian Cancer Treatment?

Epithelial ovarian cancer (EOC) is the most lethal gynecological malignancy worldwide. Due to its nonspecific symptoms and unreliable screening tools, EOC is not diagnosed at an early stage in most cases. Unfortunately, despite achieving initial remission after debulking surgery and platinum-based chemotherapy, most patients experience the recurrence of the disease. The limited therapy approaches have encouraged scientists to search for new detection and therapeutic strategies. In this review, we discuss the role of folate receptor alpha (FRα) in EOC development and its potential application as a biomarker and molecular target in designing new EOC screening and treatment methods. We summarize the mechanisms of the action of various therapeutic strategies based on FRα, including MABs (monoclonal antibodies), ADCs (antibody–drug conjugates), FDCs (folate–drug conjugates), SMDCs (small molecule–drug conjugates), vaccines, and CAR-T (chimeric antigen receptor T) cells, and present the most significant clinical trials of some FRα-based drugs. Furthermore, we discuss the pros and cons of different FR-based therapies, highlighting mirvetuximab soravtansine (MIRV) as the currently most promising EOC-targeting drug.

REFRaME-O1/ENGOT-OV79/GOG-3086: A phase II/III, open-label study evaluating the efficacy and safety of luveltamab tazevibulin versus investigator's choice of chemotherapy in women with relapsed platinum-resistant epithelial ovarian cancer expressing folate receptor alpha

REFRaME-O1/ENGOT-OV79/GOG-3086: A phase II/III, open-label study evaluating the efficacy and safety of luveltamab tazevibulin versus investigator's choice of chemotherapy in women with relapsed platinum-resistant epithelial ovarian cancer expressing folate receptor alpha

Variability of folate receptor alpha expression by race among recurrent ovarian cancer patients

Variability of folate receptor alpha expression by race among recurrent ovarian cancer patients

Folate receptor positivity in low-grade serous ovarian cancer: Exploring new therapeutic possibilities

Folate receptor positivity in low-grade serous ovarian cancer: Exploring new therapeutic possibilities

Folic acid conjugated sodium alginate based redox responsive smart functional microgels as a potential targeted anticancer drug carrier

In this study, we have fabricated a sodium alginate-based redox-responsive, and cancer cell-targeted specific microgel using a water in oil (w/o) mini emulsion process and characterized it accordingly. Initially, we synthesized folic acid (FA)-grafted sodium alginate (SA), a biocompatible polysaccharide, followed by the preparation of a semi-interpenetrating polymer network (semi-IPN) microgel by crosslinking polyacrylamide (PAAm) with a disulfide crosslinker. The presence of disulfide linkages in the microgel formulation accelerated the release of the anti-cancer drug Doxorubicin (DOX) in the reducing domain of cancer cells (in vitro) (cumulative drug release amount 11 ± 3 %). Folic acid was incorporated into the microgels to enhance its targeting towards cancer cells due to the presence of the folate receptor on cancer cells. MTT (3-(4,5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide) assay showed an IC50 of ∼30 μg/mL of the DOX-loaded microgels towards breast cancer cells (MDA-MB-231) and a nontoxic behaviour towards normal mouse fibroblast cells (L929). FACS (Fluorescence-Activated Cell Sorting) and cell uptake studies showed significant cell apoptosis upon application of the drug-loaded microgels. In our opinion, this type of microgel can be a potential drug delivery vector for cancer treatment.

Ligand-Based targeting in drug delivery system

Targeted drug delivery system has come up as a new approach that increase the effectiveness of the drug while reducing the side effects of cancer therapy. This review focuses on the application of ligand-based drug targeting, especially antibodies-drug conjugates in the case of ovarian cancer and folate-receptor targeting in the case for tumor cells. This review will provide analytical insights about the effectiveness of two approaches on improving bioavailability, targeting specific tumor cells, and enhancing drug carrying capacity. The findings show that folate receptor targeting does has significant potential in enhancing curcumin’s anticancer activity. However, there are still challenges such as rapid decomposition of curcumin. This review also highlights the need for future experiments and research to consolidate the effectiveness of the drug delivery system and explore their potential in broader clinical environment.

Platinum Resistance in Ovarian Cancer: Is This the End of the Line?

Approximately 80% of females with ovarian cancer are diagnosed with advanced disease, and around 70% of these females relapse within 3 years of first-line treatment. Five-year survival for newly diagnosed advanced ovarian cancer is less than 50%. Platinum-based chemotherapy is the cornerstone of systemic treatment; however, it is not appropriate for patients with relapsed ovarian cancer who had disease progression during previous platinum treatment, early symptomatic progression post-platinum treatment, or who are platinum intolerant. New drugs are needed to address the unmet need in patients with relapsed ovarian cancer who are not eligible for platinum-based chemotherapy. This article presents highlights from a satellite symposium conducted as part of the European Society for Medical Oncology (ESMO) Congress 2024, which took place from 13th–17th September 2024 in Barcelona, Spain. The objectives of the symposium were to improve understanding of the current treatment pathways and unmet needs for patients with ovarian cancer who are ineligible for platinum-based therapies, to raise awareness of the rationale for targeting folate receptor α (FRα) in a variety of novel therapeutics for platinum-resistant ovarian cancer (PROC), and to review the efficacy outcomes and side effects from recent clinical trials involving antibody–drug conjugates (ADC) in PROC. In this symposium, Ana Oaknin, Vall d’Hebron University Hospital, Barcelona, Spain, described the current landscape in advanced ovarian cancer and the unmet need in relapsed disease; Philipp Harter, Evangelische Kliniken Essen-Mitte, Germany, explored FRα-targeted therapeutics in PROC; and Kathleen Moore, Stephenson Cancer Center, University of Oklahoma, Norman, USA, discussed ADC development beyond FRα in PROC. The symposium concluded with a lively discussion, including questions from the audience, key examples of which are included in this article.

Evaluation of 18F FOL a PET tracer targeting folate receptor beta in a rat model of myocardial infarction

Abstract Background Expression of folate receptor-β (FR-β) on the activated macrophages is associated with inflammatory diseases. However, the role of FR-β in the immune response following myocardial infarction (MI) remains unknown. Purpose To investigate FR-β-targeted positron emission tomography (PET) tracer aluminium fluoride-18-labeled 1,4,7,-triazacylononane-1,4,7-triacetic acid conjugated folate ([18F]FOL) for imaging immune response in a rat model of MI. Methods Rats underwent PET scan from 20 to 40 minutes after injection of 50±6 MBq of [18F]FOL on 3 (n=6), 7 (n=21), 15 (n=9), and 90 (n=11) days after surgical ligation of the left coronary artery. As controls, rats were studied on 3 (n=6), 7 (n=14), 15 (n=5), and 90 (n=8) days after sham-operation. [18F]FDG PET was performed a day before [18F]FOL injection in order to localize myocardium and area of MI. After [18F]FOL PET, autoradiography, histology, and staining of CD68 were performed 3 (n=6), 7 (n=6), and 90 (n=11) days after MI in order to detect [18F]FOL uptake and activated macrophages in myocardial tissue sections. A subgroup of rats (n=15) underwent serial [18F]FOL PET 7 and 90 days after MI together with echocardiography in order to evaluate left ventricular function. Results The presence of MI was confirmed by histology in all rats after coronary ligation. PET images showed increased uptake of [18F]FOL in the area of MI. The uptake was significantly higher (p<0.001) in the area of MI than in the myocardium of sham-operated rats on day 3 (SUV 2.04±0.25 vs. 0.74±0.12), and remained increased on day 7 (SUV 1.39±0.33 vs. 0.68±0.12), day 15 (SUV 1.24±0.20 vs. 0.59±0.07), and day 90 (SUV 1.39±0.25 vs. 0.69±0.11). On day 3, pre-treatment with folate glucosamine (n=3) reduced the [18F]FOL signal by 50% in the area of MI indicating specific binding to FR-β. Autoradiography confirmed focally increased uptake of [18F]FOL in the area of MI. A positive correlation was found between uptake of [18F]FOL in PET images and areal percentage of CD68-positivity in the area of MI (r2=0.447, p<0.001). Uptake of [18F]FOL on day 7 after MI was associated with decline in left ventricular ejection fraction (R=-0.665, p<0.01) and increase in left ventricular systolic volume (R=0.561, p=0.02) between 7 and 90 days after MI. Conclusion Cardiac [18F]FOL PET detects increased macrophage-associated FR-β expression after MI in a rat model. FR-β expression peaks early and remains elevated up to 3 months in the infarcted area. Increased FR-β expression early after MI is associated with worsening of left ventricular systolic function late after MI.

Development of a Novel 99mTc-Labeled Folate Derivative Containing Phenyl Isonitrile to Target Folate Receptor with Reduced Renal Uptake.

The folate receptor has attracted much attention in the field of radiolabeled imaging agents due to the significant difference in its expression levels between tumor cells and most normal cells. However, the development of folate-based imaging agents has been limited by their high uptake in the kidney. In this study, to reduce the high renal uptake of radiolabeled folate-based tracers, a phenyl-isonitrile folate derivative (CNMBFA) was designed and labeled with technetium-99m. The complex obtained via the one-step kit labeling method had a high labeling yield (>95%) and high in vitro stability and hydrophilicity (log D7.4 = -1.72 ± 0.13). The results of the in vitro cell uptake and blocking studies and competitive binding experiments revealed that the [[99mTc]Tc-(CNMBFA)6]+ complex was specific for the folate receptor. Biodistribution and inhibition studies in KB tumor-bearing mice revealed moderate uptake and significant inhibition of the complex in tumors, whereas the renal uptake of [[99mTc]Tc-(CNMBFA)6]+ was significantly lower than that of previously reported tracers. Micro-SPECT/CT images further supported its ability to target the folate receptor for tumor imaging. Taken together, these results indicate that [[99mTc]Tc-(CNMBFA)6]+ is a potential tumor imaging agent that has good tumor-targeting properties with minimal radiation damage to the kidney.

Antibody-drug conjugates as targeted therapy for treating gynecologic cancers: update 2025.

Provide the most up-to-date information on the dynamic landscape of antibody-drug conjugates (ADCs) in gynecologic cancers. We discuss the latest research that supports the approved ADCs and outline the ongoing trials and preliminary results that may lead to ADC approvals in the future. Current gaps in knowledge and areas for future research are discussed. ADCs are rapidly changing the landscape of gynecologic cancer care. Three ADCs are currently FDA approved and used routinely in clinical practice, with many more currently in clinical development. The most common ADC target is folate receptor alpha of which there are 8 different folate receptor targeting ADCs in development. Other targets under investigation include trophoblast cell surface antigen-2 (Trop-2), claudin-6 (CLDN6), cadherin-6 (CDH6), nectin-4, HER-2 and B7-H4. ADCs can cause new and unique adverse effects, including ocular toxicities and interstitial lung disease. ADCs offer the opportunity for a more effective and personalized treatment approach for gynecologic cancer patients. Side effects must be closely monitored, and preventive measures must be followed to maximize benefit and minimize toxicity. A better understanding of the role of target proteins as biomarkers to predict response to ADCs will be critical for successful clinical implementation of ADCs and further research in this area is necessary.

Glycosyl Mobile Radical Structures of Folic Acid Receptors Impact the Internalization of Functionalized Folate Amphiphilic Alternating Copolymer in Cancer Cells

Folate receptor alpha (FRα) is a glycosylphosphatidylinositol (GPI) membrane-anchored protein containing three N-glycosylated residues at the N47, N139, and N179 termini. These glycosylation sites have been reported to be crucial for the receptor’s structural integrity and its ability to bind and internalize FA. Here, we investigated the role of FRα glycosylation in the binding and internalization efficacy of FA–DABA–SMA in pancreatic PANC-1 cancer cells. There is a strong association of the FA copolymer with FRα with a Pearson coefficient R-value of 0.7179. PANC-1 cancer cells were pretreated with maackia amurensis lectin II (MAL-2), sambucus Nigra lectin (SNA-1), peanut agglutinin (PNA), and wheat germ agglutinin lectin (WGA) at different doses followed by 20 kDa and 350 kDa FA–DABA–SMA loaded with coumarin 153 (C153). Increasing the dosage of MAL2, SNA-1, PNA, and WGA concomitantly and significantly increased the internalization of C153-loaded FA–DABA–SMA in the cells. The half maximal effective lectin concentrations (EC50) to induce cellular internalization into the cytoplasm of the lectins for MAL-2 were 35.88 µg/mL, 3.051 µg/mL for SNA-1, 7.883 µg/mL for PNA, and 0.898 µg/mL for WGA. Live cell imaging of the internalization of 20 kDa and 350 kDa FA copolymers indicated an aggregation of 350 kDa copolymer with FRα in the cytoplasm. In contrast, the 20 kDa FA copolymer remained in the membrane. The data indicate for the first time that the mobile positions of the glycosyl radical groups and the receptor tilt in generating steric hindrance impacted the individual FRα receptors in the binding and internalization of 350 kDa FA–DABA–SMA in cancer cells.

Targeting Synovial Macrophages with Astaxanthin-Loaded Liposomes for Antioxidant Treatment of Osteoarthritis.

Osteoarthritis (OA) is a chronic joint disease highly associated with an imbalance in the network of inflammatory factors and typically characterized by oxidative stress and cartilage damage. Moreover, the specificity of the joint structure makes it difficult for drugs to achieve good penetration and effective enrichment in the joint cavity. Therefore, therapeutic strategies that increase the specific targeting of drugs to inflammatory joint and incorporate antioxidative stress effects are important to improve the efficacy of OA. Here, we developed a folic acid-modified liposomal nanoparticle (AST@Lip-FA) loaded with the antioxidant astaxanthin (AST) to enhance the water solubility and stability of AST and to target the delivery of AST to the site of OA, leading to a significant improvement in therapeutic efficacy. In vitro experiments demonstrated that, due to the recognition by FA of the receptor folate receptor β on the surface of activated macrophages, the cellular uptake efficiency of AST@Lip-FA was increased. Meanwhile, intracellularly overexpressed inflammatory mediators such as reactive oxygen species and nitric oxide were efficiently removed by AST@Lip-FA. In addition, in the ACLT-induced OA mouse model, AST@Lip-FA was precisely enriched in the inflamed joints and achieved long-term retention, fully utilizing the anti-inflammatory, antioxidant, and cartilage-protecting effects of AST to effectively alleviate the progression of OA. In summary, AST@Lip-FA has an important prospect as a potential and effective therapeutic strategy for OA.

Combination therapy of Sanjie Zhentong capsules for endometriosis: Impact on hormone regulation and serum expression of FOLR1 and MSLN.

Examining the impact of Sanjie Zhentong capsules on hormone levels and serum expression of folate receptor 1 (FOLR1) and Mesothelin (MSLN) in endometriosis patients. A cohort of 150 endometriosis patients admitted between June 2019 and June 2023 was divided into 2 groups based on Sanjie Zhentong intake, comprising 75 cases each. Comparative analyses encompassed uterine artery hemodynamic parameters, ultrasound (B-mode) findings, Visual Analog Scale scores, traditional Chinese medicine symptom scores, Endometriosis Health Profile-30 scores, and serum levels of FOLR1, MSLN, and sex hormones before and after treatment. Post-treatment, both groups exhibited increased peak systolic velocity compared to pretreatment, with the Sanjie Zhentong group showing a significantly higher increase (P < .05). Moreover, resistance index decreased in both groups post-treatment, with the Sanjie Zhentong group displaying a greater reduction (P < .05). Adnexal mass diameters, endometrial thickness, and uterine volume decreased post-treatment in both groups, with the Sanjie Zhentong group experiencing a more pronounced decrease (P < .05). Serum FOLR1, MSLN, and sex hormone levels also decreased post-treatment in both groups, with the Sanjie Zhentong group exhibiting a more significant decrease (P < .05). Additionally, pain scores, traditional Chinese medicine symptoms, and Endometriosis Health Profile-30 scores decreased post-treatment in both groups, with the Sanjie Zhentong group demonstrating a greater reduction (P < .05). Sanjie Zhentong capsules demonstrate efficacy in regulating sex hormones, reducing serum FOLR1 and MSLN expression, alleviating pain, shrinking adnexal masses, decreasing uterine size, and improving uterine artery blood flow in endometriosis patients.

Development of folate receptor targeting chimeras for cancer selective degradation of extracellular proteins.

Targeted protein degradation has emerged as a novel therapeutic modality to treat human diseases by utilizing the cell's own disposal systems to remove protein target. Significant clinical benefits have been observed for degrading many intracellular proteins. Recently, the degradation of extracellular proteins in the lysosome has been developed. However, there have been limited successes in selectively degrading protein targets in disease-relevant cells or tissues, which would greatly enhance the development of precision medicine. Additionally, most degraders are not readily available due to their complexity. We report a class of easily accessible Folate Receptor TArgeting Chimeras (FRTACs) to recruit the folate receptor, primarily expressed on malignant cells, to degrade extracellular soluble and membrane cancer-related proteins in vitro and in vivo. Our results indicate that FRTAC is a general platform for developing more precise and effective chemical probes and therapeutics for the study and treatment of cancers.

Assessment of mirvetuximab soravtansine immunogenicity in patients with folate receptor alpha-positive ovarian cancer.

Aim: The aim of this research was to evaluate the immunogenicity of mirvetuximab soravtansine (MIRV), an antibody-drug conjugate in patients with folate receptor alpha-positive ovarian cancer across four clinical studies.Materials & methods: An assay was developed and validated for the detection of antidrug antibodies (ADAs) against MIRV. A cell-based method was also developed and validated for the detection of neutralizing anti-MIRV antibodies (NAbs). Both ADAs and NAbs were assessed across four clinical studies in 734 patients.Results: Across studies, MIRV demonstrated low immunogenicity with 7.8% of patients with treatment-emergent ADAs, 7.2% with treatment-unaffected ADAs, and 0.5% with treatment-enhanced ADAs. MIRV trough concentrations were comparable in ADA-negative and ADA-positive individuals. Limited data suggest that MIRV ADAs may be associated with decreased efficacy. Due to the very limited number of NAb-positive individuals, no conclusions could be drawn on the effect of NAb on efficacy.Conclusion: Both the validation tests and the data from the MIRV clinical studies demonstrated that these assays were suitable and reliable for the detection of MIRV ADAs and NAbs. These validated assays will continue to be used to monitor MIRV immunogenicity in future clinical trials.

Characterizing the genomic landscape through the lens of FOLR1 status in low and high grade serous ovarian carcinoma

ObjectiveTargeted therapy in folate receptor alpha (FOLR1)-positive high grade serous ovarian carcinoma (HGSOC) is now a mainstay for platinum-resistant disease. However, the rate of FOLR1-positivity in low grade serous ovarian carcinoma (LGSOC) is not well documented. Less common than HGSOC, LGSOC tends to respond poorly to traditional platinum-based chemotherapeutic regimens, particularly in recurrence. Thus, there is an urgent need to identify molecular targets that may assist in identifying more efficacious treatments for LGSOC. In this work, we assessed the genomic and transcriptomic landscapes in FOLR1-positive/negative LGSOC compared to its high-grade counterpart. MethodsUsing a large precision oncology database, next-generation sequencing and immunohistochemistry was performed on a cohort of 281 LGSOC and 5086 HGSOC. Associated MAPK activation was calculated based on NGS results and patient survival analysis was completed stratified by molecular alteration. ResultsCompared with LGSOC (24.6 %), HGSOC tumors have significantly higher prevalence of FOLR1+ status (43.5 %) and significantly higher PD-L1+ status. Conversely, LGSOC had higher prevalence of KRAS and NRAS mutations, with a near exclusivity for BRAF mutation compared to HGSOC. FOLR1- LGSOC and HGSOC had similar prevalences of T cell-inflamed tumors, though FOLR1+ LGSOC had a significantly lower prevalence of T-Cell inflamed tumors than FOLR1+ HGSOC. MAPK activation, quantified via MAPK activation score (MPAS), was significantly higher in low-grade tumors compared to HGSOC, yet no difference between FOLR1+ vs FOLR1- LGSOC was observed. ConclusionsThough less than in high-grade disease, a notable portion of low-grade tumors were FOLR1+, suggesting FOLR1 expression in LGSOC could be a viable target for this rare histology, particularly in the recurrent setting.

Construction of self-driving anti-αFR CAR-engineered NK cells based on IFN-γ and TNF-α synergistically induced high expression of CXCL10

IntroductionOvarian cancer is the most malignant gynecological tumor. Previous studies have demonstrated that chimeric antigen receptor (CAR)-engineered NK-92 cells targeting folate receptor α (αFR) (NK-92-αFR-CAR) can specifically kill αFR-positive ovarian cancer cells. However, the migration barrier restricts antitumor effects of CAR-engineered cells. ObjectivesTo elucidate the mechanism by which NK-92-αFR-CAR cells induce the secretion of chemokine CXCL10 during killing ovarian cancer cells. It is speculated that NK-92-αFR-CAR-CXCR3A can target αFR and have chemotaxis of CXCL10, and they may have stronger killing effect of ovarian cancer. MethodsStudy the mechanism of CXCL10 expression strongly induced by TNF-α and IFN-γ combined stimulation in ovarian cancer cells. Construct the fourth generation of NK-92-αFR-CAR-CXCR3A cells, which were co-expressed CXCR3A and αFR-CAR. Evaluate the killing and migration effects of NK-92-αFR-CAR-CXCR3A in vitro and in vivo. ResultsRNA sequencing (RNA-seq) first revealed that the expression level of the chemokine CXCL10 was most significantly increased in ovarian cancer cells co-cultured with NK-92-αFR-CAR. Secondly, cytokine stimulation experiments confirmed that IFN-γ and TNF-α secreted by NK-92-αFR-CAR synergistically induced high CXCL10 expression in ovarian cancer cells. Further signaling pathway experiments showed that IFN-γ and TNF-α enhanced the activation level of the IFN-γ-IFNGR-JAK1/2-STAT1-CXCL10 signaling axis. Cytotoxicity experiments showed that NK-92-αFR-CAR-CXCR3A cells could not only efficiently kill αFR-positive ovarian cancer cells in vitro but also secrete IFN-γ and TNF-α. Higher migration than that of NK-92-αFR-CAR was detected in NK-92-αFR-CAR-CXCR3A using transwell assay. NK-92-αFR-CAR-CXCR3A effectively killed tumor cells in different mouse xenograft models of ovarian cancer and increased infiltration into tumor tissue. ConclusionThis study confirmed that IFN-γ and TNF-α secreted by αFR-CAR-engineered NK cells can synergistically induce high expression of CXCL10 in ovarian cancer cells and constructed self-driving αFR-CAR-engineered NK cells that can break through migration barriers based on CXCL10, which may provide a new therapeutic weapon for ovarian cancer.