Folate Receptor Alpha Overexpression Research Articles

Targeted Ultrafine Iron Oxide Nanoparticles for Delivery of the Topoisomerase Inhibitor SN38 and Ovarian Cancer Treatment

Ovarian cancer remains a challenge to decrease mortality and improve diagnostic efficiency in gynecological cancers. To develop a delivery system capable of efficient cancer cell targeting and delivering novel efficacious therapeutics, we assembled folic acid (FA) conjugated ultrafine iron oxide nanoparticles (uIONP) with encapsulation of DNA topoisomerase inhibitor SN38, which target ovarian cancer cells with over-expression of folate receptor alpha (FRα) and deliver SN38 to induce apoptosis. The assembled FA-uIONP-SN38 exhibited higher drug loading efficiency than the larger counterparts with core diameters more than 10 nm. The targeting specificity of FA-uIONP-SN38 for SKOV-3 cancer cells was validated, with HEK293 kidney cells and Raw264.7 macrophages as non-targeted cell line control. It was found that more SKOV-3 cancer cells were killed due to apoptosis by FA-uIONP-SN38 at the same SN38 dosages compared with uIONP-SN38 and free SN38, respectively. The delivery of this inhibitor to SKOV-3 cancer cells by FRα-targeted FA-uIONP carrier was enhanced by about 10-folds with less cytotoxicity comparing to the free drug SN38. The developed FA-uIONP-SN38 holds a great potential as a theranostic approach in treating ovarian cancer.

Overexpression of folate receptor alpha is an independent prognostic factor for outcomes of pancreatic cancer patients.

Pancreatic cancer has a poor prognosis; hence, novel prognostic markers and effective therapeutic targets should be identified. We aimed to evaluate folate receptor alpha (FR-α) expression in pancreatic cancer and examine its association with clinicopathological features. We utilized tissue samples from 100 primary pancreatic cancer patients who underwent surgery. FR-α was expressed in 37 of 100 cases (37%). The FR-α-positive group (median, 18.8 months) had a significantly poorer prognosis than the FR-α-negative group [median 21.3 months; HR 1.89 (1.12-3.12); P = 0.017]. These groups were not significantly different regarding progression-free survival (P = 0.196). Furthermore, other serum tumor markers including CA19-9 (mean, 186 vs. 822U/ml; P = 0.001), Dupan-2 (286 vs. 1133U/ml; P = 0.000), and Span-1 (69.7 vs. 171.9U/ml; P = 0.006) were significantly downregulated in the FR-α-positive group. CA19-9 was another prognostic factor, in addition to FR-α, and patient prognosis showed clear stratification curves with the expression of these two molecules. Along with CA19-9, FR-α expression was an independent prognostic factor for the overall survival. FR-α and CA19-9 helped predict patient prognosis based on stratification curves.

Abstract B107: Antitumor activity of new antibody-drug conjugate with eribulin in triple-negative breast cancer PDX

Abstract Trastuzumab-emtansine (T-DM1) is an ADC that combines the HER2-targeting properties of trastuzumab with intracellular delivery of DM1 (a highly potent derivative of the anti-microtubule agent maytansine). A new biomarker of recent interest in the cancer field is folate receptor alpha (FRA), a membrane-bound protein with high affinity for binding and transporting folate into cells. Overexpression of FRA may confer a growth advantage to tumors by increasing folate uptake and affect cell proliferation via alternative cell signaling pathways. FRA levels have been found to be elevated in tumors of epithelial origin compared to normal tissue, including TNBC. Due to an absence of potential targeted therapy for this breast cancer subtype, the finding that a significant number of TNBCs express abundant FRA suggests an important population of patients may benefit from FRA-targeting therapy. Eighty-five percent of preclinical agents entering oncology clinical trials fail to demonstrate sufficient safety or efficacy to gain regulatory approval. This failure rate shows a weak understanding of the complexity of human cancer, the continued limitations of the predictive value of existing preclinical models, and the scale at which cancer models are interrogated in the preclinical setting. There is a need for new experimental models that better replicate the diversity of human tumor biology in a preclinical setting. It is now evidenced that PDXs recapitulate human tumor biology and predict patient drug response by directly comparing drug responses in patients and their corresponding xenografts. To extend such observations to a greater number of human cancers, we have generated in collaboration with Eisai an extensive collection of breast PDXs. In this study, MORAb-202, a novel folate receptor-targeting eribulin conjugate created through a Morphotek and Eisai collaboration, was tested in FRA-expressing TNBC PDXs. Compared to free eribulin, MORAb-202 showed higher and specific antitumor activity on FRA-positive PDXs. When injected once, MORAb-202 at 5 mg/kg (0.1 mg/kg eq eribulin) showed a treatment group/control group percent (T/C %) between 4 to 17%, whereas eribulin injected at 0.1 mg/kg following the same equivalent dose and schedule of administration showed a T/C % between 74 to 93%. On non-FRA-expressing PDXs, antitumor activities of MORAb-202 and eribulin were similar. Interestingly, to find similar antitumor activity, the dose of eribulin without vectorization should be enhanced to 32-fold compared to targeted eribulin. At this dose of eribulin, close to the maximum tolerated dose, a tumor relapse is observed (100%), whereas no relapse was observed for mice treated with MORAb-202. These encouraging results lead to a proof of concept of the use of MORAb-202 in TNBC. Citation Format: Marc Hillairet de Boisferon, Caroline Mignard, Coralie Durix, Toshimitsu Uenaka, Katherine Rybinski, Xin Cheng, Keiji Furuuchi, Eart Albone. Antitumor activity of new antibody-drug conjugate with eribulin in triple-negative breast cancer PDX [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B107.

Ovarian cancer treatment with a tumor-targeting and gene expression-controllable lipoplex.

Overexpression of folate receptor alpha (FRα) and high telomerase activity are considered to be the characteristics of ovarian cancers. In this study, we developed FRα-targeted lipoplexes loaded with an hTERT promoter-regulated plasmid that encodes a matrix protein (MP) of the vesicular stomatitis virus, F-LP/pMP(2.5), for application in ovarian cancer treatment. We first characterized the pharmaceutical properties of F-LP/pMP(2.5). The efficient expression of the MP-driven hTERT promoter in SKOV-3 cells was determined after an in-vitro transfection assay, which was significantly increased compared with a non-modified LP/pMP(2.5) group. F-LP/pMP(2.5) treatment significantly inhibited the growth of tumors and extended the survival of mice in a SKOV-3 tumor model compared with other groups. Such an anti-tumor effect was due to the increased expression of MP in tumor tissue, which led to the induction of tumor cell apoptosis, inhibition of tumor cell proliferation and suppression of tumor angiogenesis. Furthermore, a preliminary safety evaluation demonstrated a good safety profile of F-LP/pMP(2.5) as a gene therapy agent. Therefore, FRα-targeted lipoplexes with therapeutic gene expression regulated by an hTERT promoter might be a promising gene therapy agent and a potential translational candidate for the clinical treatment of ovarian cancer.

Folate Receptor Alpha Immunohistochemistry in Cytology Specimens of Metastatic Breast Carcinoma

Background: Folate receptor alpha (FRA) is involved in folate accumulation and utilization, and is expressed in varying proportions in breast, ovary and parotid epithelial cells, among others. FRA overexpression by immunohistochemistry (IHC) has been shown in estrogen/progesterone receptor (ER/PR)-negative carcinoma (40-74%) and in triple-negative breast carcinoma (TNBC; 50-86%) in histological specimens of primary breast cancers. We assessed the feasibility of IHC in detecting FRA expression and its patterns and clinical significance in metastatic TNBC in fine-needle aspiration (FNA) cell blocks (CBs). Materials and Methods: Metastatic breast ductal carcinoma cases were retrospectively immunostained with FRA IHC on FNA CBs. FRA staining was scored qualitatively (+/-), by intensity (0-3) and by staining area (0-100%). Of these metastatic cases, a subset of primary breast carcinoma cases was also immunostained with FRA. The results were correlated with ER, PR and human epidermal growth factor receptor 2 (Her2/Neu) performed by routine IHC. Results: A total of 40 FNA CBs with metastatic disease were studied, including hormone (ER/PR) positive (n = 5), triple positive (n = 5), Her2/Neu-only positive (n = 5) and TNBC (n = 25). FRA IHC showed immunoreactivity with moderate positivity in only 1 (4%) TNBC. All the remaining 39 cases were negative for FRA expression. Five cases of primary TNBC were stained with FRA IHC and were negative for FRA expression. Conclusions: Our data suggest that FRA expression by IHC was rarely associated with ER/PR-negative tumors relative to ER/PR-positive tumors and, more importantly, with TNBC in FNA CBs. This finding may have a clinical significance and prognostic implications in metastatic breast carcinoma. Furthermore, 5 primary TNBC cases did not overexpress FRA by IHC. Hence, antifolate receptor therapies do not appear to be clinically relevant in TNBC based on immunostaining of FNA CBs of metastatic breast cancers.

Assessment of folate receptor-β expression in human neoplastic tissues.

Over-expression of folate receptor alpha on cancer cells has been frequently exploited for delivery of folate-targeted imaging and therapeutic agents to tumors. Because limited information exists on expression of the beta isoform of the folate receptor in human cancers (FR-β), we have evaluated the immunohistochemical staining pattern of FR-β in 992 tumor sections from 20 different human cancer types using a new anti-human FR-β monoclonal antibody. FR-β expression was shown to be more pronounced in cells within the stroma, primarily macrophages and macrophage-like cells than cancer cells in every cancer type studied. Moreover, FR-β expression in both cancer and stromal cells was found to be statistically more prominent in females than males. A significant positive correlation was also observed between FR-β expression on stromal cells and both the stage of the cancer and the presence of lymph node metastases. Based on these data we conclude FR-β may constitute a good target for specific delivery of therapeutic agents to activated macrophages and that accumulation of FR-β positive macrophages in the stroma could serve as a useful indicator of a tumor's metastatic potential.

Paradoxical Impact of Two Folate Receptors, FRα and RFC, in Ovarian Cancer: Effect on Cell Proliferation, Invasion and Clinical Outcome

Despite being an essential vitamin, folate has been implicated to enhance tumor growth, as evidenced by reports on overexpression of folate receptor alpha (FRα) in carcinomas. The role of another folate transporter, reduced folate carrier (RFC), is largely unknown. This study investigated the roles of folate, FRα and RFC in ovarian cancers. We demonstrated FRα mRNA and protein overexpression and reduced RFC expression in association with FRα gene amplification and RFC promoter hypermethylation, respectively. FRα overexpression was associated with tumor progression while RFC expression incurred a favorable clinical outcome. Such reciprocal expression pattern was also observed in ovarian cancer cell lines. Folate was shown to promote cancer cell proliferation, migration and invasion in vitro, and down-regulate E-cadherin expression. This effect was blocked after either stable knockdown of FRα or ectopic overexpression of RFC. This hitherto unreported phenomenon suggests that, RFC can serve as a balancing partner of FRα and confer a protective effect in patients with high FRα-expressing ovarian carcinomas, as evidenced by their prolonged overall and disease-free survivals. In conclusion, we report on the paradoxical impact of FRα (putative oncogenic) and RFC (putative tumor suppressive) in human malignancies. FRα and RFC may potentially be explored as therapeutic target or prognostic marker respectively. We recommend caution and additional research on folate supplements in cancer patients.

Abstract 4121: Folate pathway in malignant pleural mesothelioma (MPM): Novel therapeutic opportunities due to folate receptor alpha overexpression

Abstract Background. The folate pathway is active in several cancers including malignant pleural mesothelioma (MPM). Membrane receptors folate receptor alpha (FRα), reduced folate carrier 1 (RFC1), proton coupled folate transporter (PCFT) and the enzyme thymidylate synthase (TS) regulate intra-cellular uptake of folate molecules and antifolate drugs. Due to the documented overexpression of folate markers in MPM they arise as potential targets for novel specific therapies with humanized monoclonal anti-FRα antibody and also to antifolate chemotherapy with pemetrexed. Material and Methods. We studied 79 surgically resected MPM (43 epithelioid, 28 biphasic and 13 sarcomatoid histology types). Protein expression of FRα, RFC1, PCFT and TS was examined by immunohistochemistry (IHC) in FFPE tissues placed in tissue microarray platform and results correlated to clinical-pathologic characteristics. Gene expression and DNA copy number changes of the FRα gene (FOLR1) were examined in 53 mesothelioma tumors using Affymetrix U133 Plus 2.0 microarray's and Illumina HumanOmni1-Quad v1.0 chips, respectively. The expression microarray data was analyzed in the GeneSpring GX 11 software (Agilent technologies Inc.) while DNA copy number changes were detected using the Nexus 5.0 software (BioDiscovery Inc.). Results. MPM frequently showed protein over-expression of FRα, RFC1 and PCFT. Nuclear TS expression positively correlated (P<0.043) with cytoplasmic TS and membrane RFC1. Patients with T1/T2 tumor had significantly (P=0.038) higher levels of membrane FRα, compared to patients with T3/T4 tumors. Epitheloid MPM had the highest level of cytoplasmic PCFT (P=0.005) among the three histological types, while patients with sarcomatoid histology had the lowest. In the multivariate analysis, with adjustment of gender, N and M stage, Patients who expressed any nuclear TS (score >0) had a lower hazard ratio than those who did not (HR=0.309; 95%CI 0.159, 0.601; P=0.0005). Patients with any level (score >0) of cytoplasmic PCFT had lower hazard ratio than those with absence of the marker (HR=0.539; 95%CI 0.306, 0.952; P=0.0332). Our gene expression data analysis on 53 mesothelioma tumors showed greater than 2-fold increase in the FOLR1 and 3-fold increase in the TYMS messenger RNA levels compared to paired normal tissue. Additionally, at least 7 of these tumors show increased gene copy number at the FOLR1 locus (Ch11q 13.4). Conclusions. Folate membrane transporters (FRα, RFC1, PCFT) and the enzyme TS are frequently over expressed in MPM tumor tissues, and PCFT and TS are associated with tumors’ clinico-pathological features. The high level of expression of FRα in MPM supports the rational use of potential utilization of humanized monoclonal anti-FRα antibody and also standard antifolate chemotherapy in this disease. Supported by grant US DoD W81XWH-07-1-0306, ASCO CDA. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4121. doi:10.1158/1538-7445.AM2011-4121