br Discussion br In recent years
In recent years, there has been renewed interest in the clinical de-velopment of HSP90 inhibitors. Although N-terminal HSP90 inhibitors elicit strong anticancer potency in preclinical models, clinical trial outcomes have been unsuccessful due to water solubility issues, low chemical stability, toxicity, and induction of the heat shock response [11,13,22,42,43]. New strategies are needed for targeting HSP90 that provide an appropriate therapeutic balance.
We previously synthesized L80, a deguelin-derivative and a novel C-terminal HSP90 inhibitor. Evidence suggests that L80 elicits anti-pro-liferative, anti-angiogenic and anti-tumor activity in paclitaxel-resistant non-small cell lung cancer (NSCLC), while exhibiting minimal toxicity toward normal cell lines . It is notable that deguelin treatment resulted in a large number of apoptotic normal cells, while its derivative L80 was more strongly cytotoxic in TNBC Cisplatin and less toxic towards normal cells.
The majority of TNBC cases are classified histologically as high-grade invasive ductal carcinomas with aggressive behavior [44,45]. Various chemotherapeutic approaches and combination therapies focus on the suppression of TNBC, and several potential compounds are in clinical trials, including inhibitors of PI3K/AKT, MEK/ERK, and the JAK/STAT3 signaling pathway and an irreversible inhibitor of EGFR [1,4,46,47]. In this context, several potential therapeutic targets for TNBC are also HSP90 client proteins. Our findings show that L80 considerably downregulates AKT, JAK2/STAT3 and MEK/ERK expres-sion.
The role of the STAT3 signaling pathway has been well established in diverse biological functions as well as tumor development, influen-cing cell survival, motility, invasion, angiogenesis and epithelial to mesenchymal transition [48–50]. STAT3 transcriptional activity is primarily regulated by phosphorylation of the Tyr705 residue which is mediated by JAKs associated with cytokine stimulated receptors and MAPK/ERK [51,52]. Our findings indicate that the STAT3 signaling pathway is interrupted by L80 challenge, as demonstrated by a sig-nificant downregulation in phosphorylation of STAT3 and subsequent reduced expression of its downstream target genes cyclin D1 and sur-vivin.
L80 administration also induced a significant reduction in BCSC-like
Fig. 6. L80 suppresses distant metastases via JAK2/STAT3 dysregulation in vivo. (A–B) Impact of L80 on nuclear expression of phospho-JAK2 and phospho-STAT3 in allografts derived from 4T1 mammospheres. Normal rabbit IgG was used as a negative control. Quantitative graphs of the number of phospho-JAK2-positive cells (A, ***p < 0.001) and STAT3-positive cells (B, **p < 0.01) are shown in the right panel, respectively. (C) Representative bioluminescence imaging (BLI) of metastases is shown for control- and L80-treated groups. D-luciferin sodium salt at 150 mg/kg was administered in mice and subjected to the (BLI) system after removal of the primary tumors. L80-treated syngeneic mice exhibited a dramatic decrease in bioluminescence signal intensity, indicating distant metastases. (D) H&E staining images of lung and liver sections from control and L80-treated mice. The black dotted areas indicate metastatic lesions in the lungs and liver. The number of tumor nodules in lungs (**p < 0.01) and liver (**p < 0.01) was quantified. (E–F) Effect of L80 on MMP-2 and MMP-9 serum concentrations in vivo. Serum was extracted from the CTL- and L80-treated mice and MMP-2 (E) and MMP-9 (F) expression was determined by ELISA assay. Serum from normal mice was used as a negative control. Data were analyzed by Student’ t-test (***p < 0.001, normal mouse vs control allografts; ##p < 0.01, control allografts vs L80-treated allografts).
properties, together with the inhibition of ALDH1 activity, suppression of the CD44high/CD24low subpopulation, and the impairment of mam-
mosphere formation in vitro. Accumulating evidence indicates that STAT3 is indispensable for the maintenance and self-renewal of CSCs, as it regulates the expression of stem- and differentiation-related genes, while its inhibition resulted in a loss of CSC-like traits, leading to suppression of tumor development and metastasis [28,33,53–56]. Our previous findings also demonstrate that phosphorylated-STAT3 is pre-dominantly elevated in BCSC-like populations such as ALDH + cells and mammospheres derived from TNBC cells [28,32]. Furthermore, BCSC-enriched allografts exhibit enhanced and constitutive STAT3
activation, upregulated CD49f and ALDH1 expression, and more ag-gressively metastasize to other organs . Consistent with the in vitro findings, L80 administration inhibited BCSC-enriched tumor burden, angiogenesis and distal metastatic growth in the lungs and liver via downregulation of BCSC markers. This was interpreted to indicate that L80-induced inhibitory effects on tumor growth may be attributable to the suppression of STAT3 signaling.