Symbiotic exclusivity between CLOCK and TFPI2 drives stemness and immunosuppression in glioblastoma models
Fei Zhou, Lizhi Pang, Yang Liu, Fatima Khan, Peiwen Chen
Fei Zhou, Lizhi Pang, Yang Liu, Fatima Khan, Peiwen Chen
View: Text | PDF
Research Article Immunology Oncology

Symbiotic exclusivity between CLOCK and TFPI2 drives stemness and immunosuppression in glioblastoma models

Abstract

Glioblastoma (GBM) is a highly aggressive brain tumor characterized by extensive crosstalk between glioblastoma stem cells (GSCs) and immunosuppressive microglia, with our previous work identifying CLOCK and TFPI2 as key regulators of this interaction. Here, we uncover a ‘symbiotic exclusivity’ pattern between CLOCK and TFPI2, showing that, despite mutually exclusive amplifications, they sustain symbiotic regulatory interactions in GBM. The CLOCK-BMAL1 complex transcriptionally upregulates TFPI2, while TFPI2-driven hypoxia inducible factor 1 α (HIF-1α) signaling activates nuclear factor k B (NF-kB) P65 to upregulate the CLOCK-BMAL1 complex, creating a positive feedback loop to promote stemness, immunosuppression, and tumor progression. Disrupting the CLOCK-TFPI2 interplay through dual inhibition of their downstream effectors reduces GSC stemness and immunosuppressive microglia, activates antitumor immunity, and synergizes with anti-PD1 therapy to achieve complete tumor regression in 50%–62.5% of tumor-bearing mice. This study uncovers a promising therapeutic strategy for a broader subset of patients with GBM with high expression of either CLOCK or TFPI2, and provides a framework for identifying ‘symbiotic exclusivity’ genes in cancer.

Authors

Fei Zhou, Lizhi Pang, Yang Liu, Fatima Khan, Peiwen Chen

×

Figure 2

CLOCK-induced GSC self renewal is regulated by TFPI2.

Options: View larger image (or click on image) Download as PowerPoint
CLOCK-induced GSC self renewal is regulated by TFPI2. (A–D) Immunoblots ...
(A–D) Immunoblots (A and B) and RT-qPCR (C and D, n = 3) for CD133 (PROM1) and SOX2 in GSC272 harboring shRNA control (shC), CLOCK shRNA (shCLOCK, A), or shBMAL1 (B) in the presence or absence of TFPI2 overexpression (OE). n = 3. (E and F) In vitro limiting dilution assays in GSC272 harboring shC, shCLOCK (E), or shBMAL1 (F) in the presence or absence of TFPI2 OE. (G–I) Representative images (G) and quantification of relative tumorsphere number (H) and size (I) of GSC272 expressing shC, shCLOCK, or shBMAL1 in the presence or absence of TFPI2 OE. Scale bar: 200 μm. n = 8. (J–M) Representative images and quantification of proliferation in GSC272 harboring shC, shCLOCK (J and K), and shBMAL1 (L and M) with or without TFPI2 OE. n = 3. (N–Q) Immunoblots (N and O) and RT-qPCR (P and Q, n = 3) for CD133 (PROM1) and SOX2 in GSC17 and GSC23 harboring Control or CLOCK OE with or without shTFPI2. (R and S) In vitro limiting dilution assays in GSC17 (R) and GSC23 (S) expressing Control or CLOCK OE with or without shTFPI2. (T and U) Representative images (T) and quantification of relative tumorsphere number (U) of GSC17 or GSC23 harboring CLOCK OE in the presence or absence of shTFPI2. Scale bar: 200 μm. n = 8. (V–Y) Representative images and quantification of proliferation in GSC17 (V and W) and GSC23 (X and Y) harboring CLOCK OE in the presence or absence of shTFPI2. n = 3. Data from multiple replicates are presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, 1-way ANOVA test (C, D, H, I, K, M, P, Q, U, W, and Y), 2-way ANOVA test (E, F, R, and S).