Cytomegalovirus Immediate-Early Proteins Promote Stemness Properties in Glioblastoma

Abstract

Glioblastoma (GBM) is the most common and aggressive human brain tumor. Human cytomegalovirus (HCMV) immediate early (IE) proteins that are endogenously expressed in GBM cells are strong viral transactivators with onconcogenic properties. Here, we show how HCMV IE are preferentially expressed in glioma stem-like cells (GSC), where they co-localize with the other GBM stemness markers, CD133, Nestin, and Sox2. In patient-derived GSC that are endogenously infected with HCMV, attenuating IE expression by an RNA-i-based strategy, was sufficient to inhibit tumorsphere formation, Sox2 expression, cell cycle progression, and cell survival. Conversely, HCMV infection of HMCV-negative GSC elicited robust self-renewal and proliferation of cells that could be partially reversed by IE attenuation. In HCMV-positive GSC, IE attenuation induced a molecular program characterized by enhanced expression of mesenchymal markers and pro-inflammatory cytokines, resembling the therapeutically-resistant GBM phenotype. Mechanistically, HCMV/IE regulation of Sox2 occurred via inhibition of miRNA-145, a negative regulator of Sox2 protein expression. In a spontaneous mouse model of glioma, ectopic expression of the IE1 gene (UL123) specifically increased Sox2 and Nestin levels in the IE1-positive tumors, upregulating stemness and proliferation markers in vivo. Similarly, human GSC infected with the HCMV strain Towne but not the IE1-deficient strain CR208 showed enhanced growth as tumorspheres and intracranial tumor xenografts, compared to mock-infected human GSC. Overall, our findings offer new mechanistic insights into how HCMV/IE control stemness properties in glioblastoma cells.

The Role of RhoA, RhoB and RhoC GTPases in Cell Morphology, Proliferation and Migration in Human Cytomegalovirus (HCMV) Infected Glioblastoma Cells

“…[34]. Furthermore, HCMV IE1 can promote glioblastoma stemness, cell-cycle progression and survival demonstrating a novel role of IE1 as potent driver for glioblastoma stem-like cells [35] and revealing that HCMV infection might promote pathogenesis in gliomas [36]. In addition, there are studies demonstrating that the IE1 expression increases the proliferation rate in primary glioblastoma cells via the suppression of p53 protein and the activation of Akt signaling [37][38][39] inducing the expression of a negative p53 protein regulator [40].…”

The role of CMV in glioblastoma and implications for immunotherapeutic strategies

“…Soroceanu et al found that attenuation of endogenous IE expression in glioma stem-like cells inhibited growth of the tumor cells in spheres which is a technique that enriches for stem-like cells. 35 Simultaneously, non CMV-expressing cell lines that were infected with CMV had an increase in self-renewal and proliferation. The same group found that CMV pp71 was preferentially expressed in CD133+ glioma stem-like cells.…”

“…These cells have been shown to demonstrate stem cell properties 35,36,38,39 and therefore may represent the tumor population driving tumor growth, invasiveness and treatment resistance. Therefore, although CMV based immunotherapy may eliminate only a portion of GBM tumor cells, these changes remain clinically significant due to the downstream impact of removing this cell population (Figure 1a).…”

The association between human cytomegalovirus and glioblastomas: a review

“…self-renewal) and proliferation of glioma stemlike cells in vivo. [21] Follow-up studies demonstrated that IE1 promotes the tumor phenotype in these settings through inactivation of the p53 and Rb tumor suppressor proteins and through activation of the PI3-K/AKT signaling pathway. [22] Long-term HCMV infected glioma cultures demonstrated upregulation of key signaling mediators such as SOX2, STAT3, BMX and IL-6.…”