Simultaneous targeting of Eph receptors in glioblastoma

Abstract

Eph tyrosine kinase receptors are frequently overexpressed and functional in many cancers, and they are attractive candidates for targeted therapy. Here, we analyzed the expression of Eph receptor A3, one of the most up-regulated factors in glioblastoma cells cultured under tumorsphere-forming conditions, together with EphA2 and EphB2 receptors. EphA3 was overexpressed in up to 60% of glioblastoma tumors tested, but not in normal brain. EphA3 was localized in scattered areas of the tumor, the invasive ring, and niches near tumor vessels. EphA3 co-localized with macrophage/leukocyte markers, suggesting EphA3 expression on tumor-infiltrating cells of bone marrow origin. We took advantage of the fact that ephrinA5 (eA5) is a ligand that binds EphA3, EphA2 and EphB2 receptors, and used it to construct a novel targeted anti-glioblastoma cytotoxin. The eA5-based cytotoxin potently and specifically killed glioblastoma cells with an IC50 of at least 10−11 M. This and similar cytotoxins will simultaneously target different compartments of glioblastoma tumors while mitigating tumor heterogeneity.

Maximizing Local Access to Therapeutic Deliveries in Glioblastoma. Part I: Targeted Cytotoxic Therapy

“…The gene for EphA3 was highly upregulated in G48a GBM cell tumorspheres (56) and even more so in nonpassaged GBM cells. Others found the EphA3 receptor is important for the self-renewing potential and tumorigenicity of GSCs (64).…”

“…For example, all three receptors are expressed in tumor cells of the core of GBM tumors. Importantly, IL-13RA2, EphA2, and EphA3 are present on tumorinfiltrating cells, while EphA2 is also overexpressed in tumor neovasculature (56,57). Interestingly, IL-13RA2, EphA2, and EphA3 were associated with, and play crucial roles in, the pathobiology of GSCs.…”

“…Hence, we produced a dimeric form of eA5 in a fusion with an Fc fragment of human IgG 1 , eA5-Fc ( Figure 4A) (56). We also made an eA5-Fc-PE38QQR cytotoxin chemical conjugate ( Figure 4A).…”

“…The IC 50 of eA5-Fc-PE38QQR was in the range of 10 −11 M. To confirm the specificity of the cytotoxin in targeting EphA2 and EphA3, the three GBM cell lines were pretreated with either eA1-Fc or eA5-Fc at 10 µg/mL for 1 h. As expected, the cytotoxin was less active on the three cell lines tested when pretreated with eA1-Fc, which binds only the EphA2 receptor, and completely lost its activity when cells were pretreated with eA5-Fc, which binds EphA2, EphA3, and EphB2 ( Figure 4B). Even though the readings in colorimetric cell viability assay were not reaching 100% kill, the live/dead assay (Life Technologies) demonstrated that vast majority of GBM cells were dead at 10 ng/ml of conjugate concentration and almost all were dead at 100 ng/ml of conjugate (56).…”

Molecular mechanisms of perineural spread of cutaneous carcinoma: a potential role for receptor tyrosine kinases and other biomarkers

“…(77,81) The Eph receptors/ephrins have also been implicated in tumourigenesis, progression and metastasis across a wide range of malignancies (Table 3). ( (83) Prostate cancer (84,85) Breast cancer (86,87) Melanoma (88) Glioblastoma (89,90) Gastric cancer (91) EphA3 Gastric cancer (92) Colorectal cancer (93) EphB2 Advanced cSCC (94) Glioblastoma (95)(96)(97) Cervical cancer (98) Breast cancer (99) EphB4 HNSCC (100) Non-small cell lung cancer (101) Breast cancer (99) Downregulated EphA1 NMSC (77) …”