Co-targeting Bulk Tumor and CSCs in Clinically Translatable TNBC Patient-Derived Xenografts via Combination Nanotherapy

Abstract BackgroundImmune checkpoint blockade therapies, which act on T cell inhibitory receptors, including CTLA-4 and PD-1, induce durable responses across diverse cancers. However, most patients do not respond to these therapies, and initially responsive cancers may relapse. Identifying molecular mechanisms that influence therapeutic responses and resistance is critical to realize the full therapeutic potential of immune checkpoint inhibitors. The presence of immune infiltrates in the tumor microenvironment is associated with positive outcomes in breast cancer, specifically in triple-negative breast cancer (TNBC). The underlying mechanisms driving this response are unclear. We have previously identified Neuroligin 4X (NLGN4X) as a protein expressed in TNBC.MethodsBioinformatic analysis was used for pathway analysis of TCGA TNBC patient dataset. Immunohistochemistry was performed on breast cancer tissue microarray for NLGN4X protein expression. RNA-seq was performed on MDA-MB-231 breast cancer cells for differential gene expression upon gene knockdown. Cytokine array, western blot, cell adhesion array and Nanostring was performed to determine the role of NLGN4X in TNBC.ResultsIn this study, we report that NLGN4X expression is lost in breast cancer with lymph node metastasis. Its expression negatively correlates with immune markers in vitro, The Cancer Genome Atlas (TCGA) TNBC patient dataset, and metastatic breast cancer tissues. RNA-seq analysis of the MDA-MB-231 breast cancer cell line, silenced for NLGN4X by siRNA showed more than 500 differentially regulated genes. GSEA analysis of these genes revealed upregulation of interferon signaling pathway, cytokine signaling, and downregulation of cholesterol metabolism and lipid metabolism pathways. NLGN4X knockdown induced loss of cell adhesion, epithelial to mesenchymal transition (EMT), and MAVS-IRF7 signaling in breast cancer cells. Interestingly, analysis of the TCGA dataset of 104 TNBC patients also showed interferon signaling (IFN) as one of the significant pathways downregulated in TNBC patients expressing NLGN4X.ConclusionLoss of NLGN4X leads to innate immune activation in breast cancer and coincides with an aggressive phenotype of cancer. This study identifies the role of NLGN4X in regulating interferon signaling and immune microenvironment in TNBC.

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Abstract 2998: Co-targeting bulk tumor and CSCs in clinically translatable TNBC patient-derived xenografts via combination nanotherapy

10.1158/1538-7445.am2019-2998 ◽

2019 ◽

Author(s):

Andrew Sulaiman ◽

Sara El-Sahli ◽

Sarah McGarry ◽

Lisheng Wang ◽

Suresh Gadde

Keyword(s):

Tnbc Patient

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Targeting Hypoxia Sensitizes TNBC to Cisplatin and Promotes Inhibition of Both Bulk and Cancer Stem Cells

International Journal of Molecular Sciences ◽

10.3390/ijms21165788 ◽

2020 ◽

Vol 21 (16) ◽

pp. 5788

Author(s):

Andrew Sulaiman ◽

Sarah McGarry ◽

Jason Chambers ◽

Emil Al-Kadi ◽

Alexandra Phan ◽

...

Keyword(s):

Breast Cancer ◽

Therapeutic Approach ◽

Triple Negative ◽

Ex Vivo ◽

Ovarian Cancer Cells ◽

Therapeutic Approaches ◽

Organotypic Cultures ◽

Tnbc Patient ◽

Medical Need ◽

Similar Expression Pattern

Development of targeted therapies for triple-negative breast cancer (TNBC) is an unmet medical need. Cisplatin has demonstrated its promising potential for the treatment of TNBC in clinical trials; however, cisplatin treatment is associated with hypoxia that, in turn, promotes cancer stem cell (CSC) enrichment and drug resistance. Therapeutic approaches to attenuate this may lead to increased cisplatin efficacy in the clinic for the treatment of TNBC. In this report we analyzed clinical datasets of TNBC and found that TNBC patients possessed higher levels of EGFR and hypoxia gene expression. A similar expression pattern was also observed in cisplatin-resistant ovarian cancer cells. We, thus, developed a new therapeutic approach to inhibit EGFR and hypoxia by combination treatment with metformin and gefitinib that sensitized TNBC cells to cisplatin and led to the inhibition of both CD44+/CD24− and ALDH+ CSCs. We demonstrated a similar inhibition efficacy on organotypic cultures of TNBC patient samples ex vivo. Since these drugs have already been used frequently in the clinic; this study illustrates a novel, clinically translatable therapeutic approach to treat patients with TNBC.

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Targeting Hypoxia Sensitizes TNBC to Cisplatin and Promotes Inhibition of Both Bulk and Cancer Stem Cells

10.20944/preprints202007.0527.v1 ◽

2020 ◽

Author(s):

Andrew Sulaiman ◽

Sarah McGarry ◽

Jason Chambers ◽

Emil Al-Kadi ◽

Alexandra Phan ◽

...

Keyword(s):

Therapeutic Approach ◽

Triple Negative ◽

Ex Vivo ◽

Ovarian Cancer Cells ◽

Therapeutic Approaches ◽

Organotypic Cultures ◽

Tnbc Patient ◽

Clinical Dataset ◽

Medical Need ◽

Similar Expression Pattern

Development of targeted therapies for triple-negative breast cancer (TNBC) is an unmet medical need. Cisplatin has demonstrated its promising potential for the treatment of TNBC in clinical trials; however, cisplatin treatment is associated with hypoxia that in turn promotes cancer stem cell (CSC) enrichment and drug resistance. Therapeutic approaches to attenuate this may lead to increased cisplatin efficacy in the clinic for the treatment of TNBC. In this report, we analyzed clinical dataset of TNBC and found that TNBC patients possessed higher levels of EGFR and hypoxia gene expression. A similar expression pattern was also observed in cisplatin-resistant ovarian cancer cells. We thus developed a new therapeutic approach to inhibit EGFR and hypoxia by combination of metformin and gefitinib that sensitized TNBC cells to cisplatin and led to the inhibition of both CD44+/CD24- and ALDH+ CSCs. We demonstrated a similar inhibition efficacy on organotypic cultures of TNBC patient samples ex vivo. Since these drugs have already been used frequently in the clinic, this study illustrates a novel, clinically translatable therapeutic approach to treat patients with TNBC.

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Loss of Neuroligin 4X Induces An Intrinsic Innate Immune Response in TNBC

10.21203/rs.3.rs-565425/v2 ◽

2021 ◽

Author(s):

Alehegne Yirsaw ◽

Muhammad G Omar ◽

Isra A Elhussin ◽

Dequarius King ◽

Henry J Henderson ◽

...

Keyword(s):

Breast Cancer ◽

Cell Adhesion ◽

Cancer Cells ◽

Immune Checkpoint ◽

Breast Cancer Cells ◽

Innate Immune ◽

Rna Seq ◽

Interferon Signaling ◽

Tnbc Patient

Abstract Background Immune checkpoint blockade therapies, which act on T cell inhibitory receptors, including CTLA-4 and PD-1, induce durable responses across diverse cancers. However, most patients do not respond to these therapies, and initially responsive cancers may relapse. Identifying molecular mechanisms that influence therapeutic responses and resistance is critical to realize the full therapeutic potential of immune checkpoint inhibitors. The presence of immune infiltrates in the tumor microenvironment is associated with positive outcomes in breast cancer, specifically in triple-negative breast cancer (TNBC). The underlying mechanisms driving this response are unclear. We have previously identified Neuroligin 4X (NLGN4X) as a protein expressed in TNBC.Methods Bioinformatic analysis was used for pathway analysis of TCGA TNBC patient dataset. Immunohistochemistry was performed on breast cancer tissue microarray for NLGN4X protein expression. RNA-seq was performed on MDA-MB-231 breast cancer cells for differential gene expression upon gene knockdown. Cytokine array, western blot, cell adhesion array and Nanostring was performed to determine the role of NLGN4X in TNBC.Results In this study, we report that NLGN4X expression is lost in breast cancer with lymph node metastasis. Its expression negatively correlates with immune markers in vitro, The Cancer Genome Atlas (TCGA) TNBC patient dataset, and metastatic breast cancer tissues. RNA-seq analysis of the MDA-MB-231 breast cancer cell line, silenced for NLGN4X by siRNA showed more than 500 differentially regulated genes. GSEA analysis of these genes revealed upregulation of interferon signaling pathway, cytokine signaling, and downregulation of cholesterol metabolism and lipid metabolism pathways. NLGN4X knockdown induced loss of cell adhesion, epithelial to mesenchymal transition (EMT), and MAVS-IRF7 signaling in breast cancer cells. Interestingly, analysis of the TCGA dataset of 104 TNBC patients also showed interferon signaling (IFN) as one of the significant pathways downregulated in TNBC patients expressing NLGN4X.Conclusion Loss of NLGN4X leads to innate immune activation in breast cancer and coincides with an aggressive phenotype of cancer. This study identifies the role of NLGN4X in regulating interferon signaling and immune microenvironment in TNBC.

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Abstract 2998: Co-targeting bulk tumor and CSCs in clinically translatable TNBC patient-derived xenografts via combination nanotherapy

10.1158/1538-7445.sabcs18-2998 ◽

2019 ◽

Author(s):

Andrew Sulaiman ◽

Sara El-Sahli ◽

Sarah McGarry ◽

Lisheng Wang ◽

Suresh Gadde

Keyword(s):

Tnbc Patient

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Nanoparticles Loaded with Wnt and YAP/Mevalonate Inhibitors in Combination with Paclitaxel Stop the Growth of TNBC Patient‐Derived Xenografts and Diminish Tumorigenesis

Advanced Therapeutics ◽

10.1002/adtp.202000123 ◽

2020 ◽

Vol 3 (11) ◽

pp. 2000123

Author(s):

Andrew Sulaiman ◽

Sarah McGarry ◽

Sara El‐Sahli ◽

Li Li ◽

Jason Chambers ◽

...

Keyword(s):

Tnbc Patient

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Chemotherapy Shifts the Balance in Favor of CD8+ TNFR2+ TILs in Triple-Negative Breast Tumors

Cells ◽

10.3390/cells10061429 ◽

2021 ◽

Vol 10 (6) ◽

pp. 1429

Author(s):

Tamir Baram ◽

Nofar Erlichman ◽

Maya Dadiani ◽

Nora Balint-Lahat ◽

Anya Pavlovski ◽

...

Keyword(s):

Triple Negative ◽

Tumor Infiltrating Lymphocytes ◽

Breast Tumors ◽

Therapeutic Strategies ◽

Tnbc Patient ◽

Tumor Bearing ◽

Mouse Tumors ◽

Cancer Types ◽

Infiltrating Lymphocytes ◽

The Impact

Triple-negative breast cancer (TNBC) is primarily treated via chemotherapy; in parallel, efforts are made to introduce immunotherapies into TNBC treatment. CD4+ TNFR2+ lymphocytes were reported as Tregs that contribute to tumor progression. However, our published study indicated that TNFR2+ tumor-infiltrating lymphocytes (TNFR2+ TILs) were associated with improved survival in TNBC patient tumors. Based on our analyses of the contents of CD4+ and CD8+ TILs in TNBC patient tumors, in the current study, we determined the impact of chemotherapy on CD4+ and CD8+ TIL subsets in TNBC mouse tumors. We found that chemotherapy led to (1) a reduction in CD4+ TNFR2+ FOXP3+ TILs, indicating that chemotherapy decreased the content of CD4+ TNFR2+ Tregs, and (2) an elevation in CD8+ TNFR2+ and CD8+ TNFR2+ PD-1+ TILs; high levels of these two subsets were significantly associated with reduced tumor growth. In spleens of tumor-bearing mice, chemotherapy down-regulated CD4+ TNFR2+ FOXP3+ cells but the subset of CD8+ TNFR2+ PD-1+ was not present prior to chemotherapy and was not increased by the treatment. Thus, our data suggest that chemotherapy promotes the proportion of protective CD8+ TNFR2+ TILs and that, unlike other cancer types, therapeutic strategies directed against TNFR2 may be detrimental in TNBC.

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