Triple-negative breast cancer (TNBC) includes basal-like and claudin-low subtypes for which no specific treatment is currently available. Although the retinoblastoma tumor-suppressor gene (RB1) is frequently lost together with TP53 in TNBC, it is not directly targetable. There is thus great interest in identifying vulnerabilities downstream of RB1 that can be therapeutically exploited. Here, we determined that combined inactivation of murine Rb and p53 in diverse mammary epithelial cells induced claudin-low-like TNBC with Met, Birc2/3-Mmp13-Yap1, and Pvt1-Myc amplifications. Gene set enrichment analysis revealed that Rb/p53-deficient tumors showed elevated expression of the mitochondrial protein translation (MPT) gene pathway relative to tumors harboring p53 deletion alone. Accordingly, bioinformatic, functional, and biochemical analyses showed that RB1-E2F complexes bind to MPT gene promoters to regulate transcription and control MPT. Additionally, a screen of US Food and Drug Administration-approved (FDA-approved) drugs identified the MPT antagonist tigecycline (TIG) as a potent inhibitor of Rb/p53-deficient tumor cell proliferation. TIG preferentially suppressed RB1-deficient TNBC cell proliferation, targeted both the bulk and cancer stem cell fraction, and strongly attenuated xenograft growth. It also cooperated with sulfasalazine, an FDA-approved inhibitor of cystine xCT antiporter, in culture and xenograft assays. Our results suggest that RB1 deficiency promotes cancer cell proliferation in part by enhancing mitochondrial function and identify TIG as a clinically approved drug for RB1-deficient TNBC.
Triple negative breast cancer (TNBC) includes basal-like and claudin-low subtypes for which only chemotherapy and radiation therapy are currently available. The retinoblastoma (RB1) tumor suppressor is frequently lost in human TNBC. Knockdown of RB1 in luminal BC cells was shown to affect response to endocrine, radiation and several antineoplastic drugs. However, the effect of RB1 status on radiation and chemo-sensitivity in TNBC cells and whether RB1 status affects response to divergent or specific treatment are unknown. Using multiple basal-like and claudin-low cell lines, we hereby demonstrate that RB-negative TNBC cell lines are highly sensitive to gamma-irradiation, and moderately more sensitive to doxorubicin and methotrexate compared to RB-positive TNBC cell lines. In contrast, RB1 status did not affect sensitivity of TNBC cells to multiple other drugs including cisplatin (CDDP), 5-fluorouracil, idarubicin, epirubicin, PRIMA-1met, fludarabine and PD-0332991, some of which are used to treat TNBC patients. Moreover, a non-biased screen of ∼3400 compounds, including FDA-approved drugs, revealed similar sensitivity of RB-proficient and -deficient TNBC cells. Finally, ESA+/CD24−/low/CD44+ cancer stem cells from RB-negative TNBC lines were consistently more sensitive to gamma-irradiation than RB-positive lines, whereas the effect of chemotherapy on the cancer stem cell fraction varied irrespective of RB1 expression. Our results suggest that patients carrying RB-deficient TNBCs would benefit from gamma-irradiation as well as doxorubicin and methotrexate therapy, but not necessarily from many other anti-neoplastic drugs.
Triple-negative breast cancer (TNBC) represents an aggressive subtype, for which radiation and chemotherapy are the only options. Here we describe the identification of disulfiram, an FDA-approved drug used to treat alcoholism, as well as the related compound thiram, as the most potent growth inhibitors following high-throughput screens of 3185 compounds against multiple TNBC cell lines. The average IC50 for disulfiram was ~300 nM. Drug affinity responsive target stability (DARTS) analysis identified IQ motif-containing factors IQGAP1 and MYH9 as direct binding targets of disulfiram. Indeed, knockdown of these factors reduced, though did not completely abolish, cell growth. Combination treatment with 4 different drugs commonly used to treat TNBC revealed that disulfiram synergizes most effectively with doxorubicin to inhibit cell growth of TNBC cells. Disulfiram and doxorubicin cooperated to induce cell death as well as cellular senescence, and targeted the ESA+/CD24-/low/CD44+ cancer stem cell population. Our results suggest that disulfiram may be repurposed to treat TNBC in combination with doxorubicin.
Notch plays a protumorigenic role in many cancers including prostate cancer (PCa). Global notch inhibition of multiple Notch family members using γ‐secretase inhibitors has shown efficacy in suppressing PCa growth in murine models. However, global Notch inhibition is associated with marked toxicity due to the widespread function of many different Notch family members in normal cell physiology. Accordingly, in the current study, we explored if specific inhibition of Notch1 would effectively inhibit PCa growth in a murine model. The androgen‐dependent VCaP and androgen‐independent DU145 cell lines were injected subcutaneously into mice. The mice were treated with either control antibody 1B7.11, anti‐Notch1 antibody (OMP‐A2G1), docetaxel or the combination of OMP‐A2G1 and docetaxel. Tumor growth was measured using calipers. At the end of the study, tumors were assessed for proliferative response, apoptotic response, Notch target gene expression, and DNA damage response (DDR) expression. OMP‐A2G1 alone inhibited tumor growth of both PCa cell lines to a greater extent than docetaxel alone. There was no additive or synergistic effect of OMP‐A2G1 and docetaxel. The primary toxicity was weight loss that was controlled with dietary supplementation. Proliferation and apoptosis were affected differentially in the two cell lines. OMP‐A2G1 increased expression of the DDR gene GADD45α in VCaP cells but downregulated GADD45α in Du145 cells. Taken together, these data show that Notch1 inhibition decreases PCa xenograft growth but does so through different mechanisms in the androgen‐dependent VCaP cell line vs the androgen‐independent DU145 cell line. These results provide a rationale for further exploration of targeted Notch inhibition for therapy of PCa.
BackgroundProstate cancer (PCa) bone metastases have been shown to be more resistant to docetaxel than soft tissue metastases. The proinflammatory chemokine receptor CXCR4 has been shown to confer resistance to docetaxel (DOC) in PCa cells. Balixafortide (BLX) is a protein epitope mimetic inhibitor of CXCR4. Accordingly, we hypothesized that BLX would enhance DOC‐mediated antitumor activity in PCa bone metastases.MethodsPC‐3 luciferase‐labeled cells were injected into the tibia of mice to model bone metastases. Four treatment groups were created: vehicle, DOC (5 mg/kg), BLX (20 mg/kg), and combo (receiving both DOC and BLX). Mice were injected twice daily subcutaneously with either vehicle or BLX starting on Day 1 and weekly intraperitoneally with DOC starting on Day 1. Tumor burden was measured weekly via bioluminescent imaging. At end of study (29 days), radiographs were taken of the tibiae and blood was collected. Serum levels of TRAcP, IL‐2, and IFNγ levels were measured using ELISA. Harvested tibiae were decalcified and stained for Ki67, cleaved caspase‐3, and CD34 positive cells or microvessels were quantified.ResultsTumor burden was lower in the combo group compared to the DOC alone group. Treatment with the combination had no impact on the number of mice with osteolytic lesions, however the area of osteolytic lesions was lower in the combo group compared to the vehicle and BLX groups, but not the DOC group. Serum TRAcP levels were lower in the combo compared to vehicle group, but not the other groups. No significant difference in Ki67 staining was found among the groups; whereas, cleaved caspase‐3 staining was lowest in the Combo group and highest in the BLX group. The DOC and combo groups had more CD34+ microvessels than the control and BLX groups. There was no difference between the treatment groups for IL‐2, but the combo group had increased levels of IFNγ compared to the DOC group.ConclusionsOur data demonstrate that a combination of BAL and DOC has greater antitumor activity in a model of PCa bone metastases than either drug alone. These data support further evaluation of this combination in metastatic PCa.
Sustained locoregional control of breast cancer is a significant issue in patients with high risk disease. We recently reported the results of a phase I clinical trial describing acute and late toxicity of PARP1 inhibition (PARPi) with concurrent veliparib and RT. The purpose of this study was to determine predictive biomarkers of acute (during treatment) and late (1, 2, and 3 year) toxicity in patients treated with PARPi and RT from that trial. We hypothesized that protein and gene expression changes in skin epithelial cells after concurrent PARPi and RT might be used as predictive biomarkers of toxicity. Materials/Methods: Acute toxicity was any dose-limiting toxicity defined in the protocol; late toxicity was defined as any CTCAE v4 Grade 3 event, regardless of attribution. Skin punch biopsies from the irradiated field were taken from patients at baseline (pretreatment TZ0) and 6 hrs after first fraction of RT alone (TZ1). Veliparib was then started and a 3rd punch taken 6 hrs after the second fraction of RT (TZ2). RNA was isolated from skin epithelial cells and gene expression assessed using Affymetrix Human Gene ST 2.1 arrays. GSEA and MSigDB software was used for analysis. Significant difference was predetermined as a log2 fold change of+/-1.5 and FDR adjusted p-value <0.05. Protein changes (pgH2AX, PAR, Ki67) were detected using IHC and quantitated using Aperio Digital Path software. Results: Severe acute and late toxicity rates with combined therapy were 15% (5/33) and 21% (7/33), respectively. Acute toxicity was not a predictor of late toxicity. Indeed, the two were almost mutually exclusive. Gene expression analysis identified 31 genes whose expression was significantly different 6 hrs after RT and 54 genes differed after combined treatment, including genes associated with DNA damage repair (ATM, MDM2, XPC) and proliferation (Ki67, TP53TG1). 67 genes were associated with acute toxicity, including overrepresentation of miRNAs associated with gene repression. Additionally, 63 genes were associated with late toxicity and were associated with metabolism, inflammation, and DNA damage response. There was no overlap in biomarkers of acute and late toxicity. Both pgH2AX and PAR protein levels increased and Ki67 decreased significantly (more than 2 fold for all proteins) with RT and combined therapy (T1 and T2) but were not significantly associated with acute or late toxicity at any time point. Conclusion: This demonstrates the feasibility of using skin punch biopsy in the irradiated field for biomarker analysis and identification of a number of putative biomarkers of early and late radiation toxicity following combined PARPi and RT treatment. Additionally, for the first time in patient skin punch samples we report (1) gene expression changes in acute responding genes shortly after RT and (2) decreased proliferation after combined treatment. Future trials will validate the utility of these biomarkers for predicting toxicity and will seek to identify biomarkers of treatment efficacy.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.