Oleksii Dubrovskyi - Academia.edu (original) (raw)

Papers by Oleksii Dubrovskyi

Research paper thumbnail of Novel Pyrrolopyridone Bromodomain and Extra-Terminal Motif (BET) Inhibitors Effective in Endocrine-Resistant ER+ Breast Cancer with Acquired Resistance to Fulvestrant and Palbociclib

Journal of Medicinal Chemistry, May 26, 2020

Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen re... more Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen receptor positive (ER+) breast cancer. ER is expressed in most resistance settings; thus, bromodomain and extra-terminal protein inhibitors (BETi) that target BET-amplified ER-mediated transcription have therapeutic potential. Novel pyrrolopyridone BETi leveraged novel interactions with L92/L94 confirmed by a cocrystal structure of 27 with BRD4. Optimization of BETi using growth inhibition in fulvestrant-resistant (MCF-7:CFR) cells was confirmed in endocrine-resistant, palbociclib-resistant, and ESR1 mutant cell lines. 27 was more potent in MCF-7:CFR cells than six BET inhibitors in clinical trials. Transcriptomic analysis differentiated 27 from the benchmark BETi, JQ-1, showing downregulation of oncogenes and upregulation of tumor suppressors and apoptosis. The therapeutic approach was validated by oral administration of 27 in orthotopic xenografts of endocrine-resistant breast cancer in monotherapy and in combination with fulvestrant. Importantly, at an equivalent dose in rats, thrombocytopenia was mitigated.

Research paper thumbnail of Abstract 1464: Patient-derived tumor xenograft are susceptible to formation of B-cell lymphoma after initial transplantation of human carcinoma to immunodeficient mice

Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for develop... more Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for developing novel anticancer therapies and personalized use of chemotherapeutic drugs. The PDX approach is based on the transplantation of primary or metastatic human tumors into highly immunodeficient NOD scid gamma (NSG) mice followed by continuous propagation of the established engrafment in mice. Clinicians can evaluate the effects of cancer drugs on their patients’ personalized tumor grafts enabling them to select the best treatment for the cancer patient. Here, we report that PDX tumors engrafted in NSG mice are susceptible to formation of B-cell lymphomas. We xenografted primary or metastatic human tumor fragments into NSG mice and found that tumors generated from patients’ samples of breast (1 out 3 cases), colon (2 out of 6 cases), pancreatic (1 out of 3 cases), bladder (1 out of 2 cases) and renal (1 out of 2 cases) cancer were histologically similar to lymphoid neoplasm. Moreover, we found that the first passage (F1) of some breast and pancreatic cancer PDX tumors could grow as a lymphoid neoplasm in one mouse and as an adenocarcinoma in another mouse after initial transplantation of the pieces from the same human tumor graft (F0). Whereas subcutaneous PDX tumors resembling primary human carcinoma histology were slow growing and non-metastatic, we found that subcutaneous PDX lymphoid neoplasms were fast growing and formed large metastatic lesions in lymph nodes, liver, lungs and spleen of the mouse. Immunohistochemical staining confirmed that lymphoma cells express human leucocyte common antigen CD45 and B-cell antigen CD19/20. By using Epstein-Barr Virus-encoded RNA (EBER) in situ hybridization, we found that PDX lymphoma cells were EBER-positive. Because B-cells are typically present in any solid malignant tumor, B-cell PDX lymphomas may evolve in a wide range of PDX tumor models. Our results suggest that histolopathological evaluation and lymphoid marker testing should be performed as part of the initial model characterization in order to exclude B-cell lymphomas during the development of PDX tumor model from solid human carcinoma. Citation Format: Gennadiy Bondarenko, Andrey Ugolkov, Piotr Kulesza, Stephen M. Rohan, Oleksii Dubrovskyi, Demirkan Gursel, Jeremy V. Mathews, Thomas V. O'Halloran, Jian-Jun Wei, Andrew P. Mazar. Patient-derived tumor xenograft are susceptible to formation of B-cell lymphoma after initial transplantation of human carcinoma to immunodeficient mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1464. doi:10.1158/1538-7445.AM2015-1464

Research paper thumbnail of Potent, Novel SARS-CoV-2 PLpro Inhibitors Block Viral Replication in Monkey and Human Cell Cultures

bioRxiv (Cold Spring Harbor Laboratory), Feb 15, 2021

Antiviral agents blocking SARS-CoV-2 viral replication are desperately needed to complement vacci... more Antiviral agents blocking SARS-CoV-2 viral replication are desperately needed to complement vaccination to end the COVID-19 pandemic. Viral replication and assembly are entirely dependent on two viral cysteine proteases: 3C-like protease (3CLpro) and the papain-like protease (PLpro). PLpro also has deubiquitinase (DUB) activity, removing ubiquitin (Ub) and Ub-like modifications from host proteins, disrupting the host immune response. 3CLpro is inhibited by many known cysteine protease inhibitors, whereas PLpro is a relatively unusual cysteine protease, being resistant to blockade by such inhibitors. A high-throughput screen of biased and unbiased libraries gave a low hit rate, identifying only CPI-169 and the positive control, GRL0617, as inhibitors with good potency (IC50 < 10 µM). Analogues of both inhibitors were designed to develop structure-activity relationships; however, without a co-crystal structure of the CPI-169 series, we focused on GRL0617 as a starting point for structure-based drug design, obtaining several co-crystal structures to guide optimization. A series of novel 2-phenylthiophene-based non-covalent SARS-CoV-2 PLpro inhibitors were obtained, culminating in low nanomolar potency. The high potency and slow inhibitor off-rate were rationalized by newly identified ligand interactions with a "BL2 groove" that is distal from the active site cysteine. Trapping of the conformationally flexible BL2 loop by these inhibitors blocks binding of viral and host protein substrates; however, until now it has not been demonstrated that this mechanism can induce potent and efficacious antiviral activity. In this study, we report that novel PLpro inhibitors have excellent antiviral efficacy and potency against infectious SARS-CoV-2 replication in cell cultures. Together, our data provide structural insights into the design of potent PLpro inhibitors and the first validation that non-covalent inhibitors of SARS-CoV-2 PLpro can block infection of human cells with low micromolar potency.

Research paper thumbnail of Supplemental Figures from Coactivation of Estrogen Receptor and IKKβ Induces a Dormant Metastatic Phenotype in ER-Positive Breast Cancer

This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen rec... more This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen receptor expression, phosphorylation, and activity, cell morphology and EMT. Additional analysis of metastases and stem cell phenotypes are included.

Research paper thumbnail of Supplemental Data Table 1 from Coactivation of Estrogen Receptor and IKKβ Induces a Dormant Metastatic Phenotype in ER-Positive Breast Cancer

This file contains gene expression profiling data for MCF-7-CA-IKKβ cells treated with E2, DOX, E... more This file contains gene expression profiling data for MCF-7-CA-IKKβ cells treated with E2, DOX, E2+DOX.

Research paper thumbnail of Nwani et al., Supplementary figures 1-11 from Melanoma Cells Block PEDF Production in Fibroblasts to Induce the Tumor-Promoting Phenotype of Cancer-Associated Fibroblasts

Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF... more Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF silencing in fibroblasts Supplemental Figure 3. The effect of PEDF expression in fibroblasts on tumor growth Supplemental Figure 4. The effect of endogenous PEDF on fibroblast proliferation and recruitment Supplemental Figure 5. The effect of exogenous TGFb and PDGF on PEDF mRNA in fibroblasts Supplemental Figure 6. The contribution of of TGFb and PDGF to PEDF repression assessed by shRNA silencing Supplemental Figure 7. The effect of TGFb on PDGFR mRNA levels Supplemental Figure 8. Regulation of PDGFB mRNA by TGFb Supplemental Figure 9. The role of PI3K in PEDF control by PDGF-BB Supplemental Figure 10. Inhibition of non-canonical TGFb signaling by PEDF Supplemental Figure 11. Altered expression of select genes in PEDF-null fibroblasts

Research paper thumbnail of Characterization Of Paxillin Domains Interacting With -Catenin: Implication In Endothelial Barrier Protection

Research paper thumbnail of Supplemental Figures from Coactivation of Estrogen Receptor and IKKβ Induces a Dormant Metastatic Phenotype in ER-Positive Breast Cancer

This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen rec... more This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen receptor expression, phosphorylation, and activity, cell morphology and EMT. Additional analysis of metastases and stem cell phenotypes are included.

Research paper thumbnail of Rap1 and afadin promote interactions between adherens junctions and tight junctions and regulate endothelial permeability

Research paper thumbnail of Novel Rap1‐dependent regulator of endothelial cell junctions afadin mediates protective effects by oxidized phospholipids in the models of acute lung injury

Research paper thumbnail of Extending the Calpain–Cathepsin Hypothesis to the Neurovasculature: Protection of Brain Endothelial Cells and Mice from Neurotrauma

ACS Pharmacology & Translational Science, 2021

The calpain-cathepsin hypothesis posits a key role for elevated calpain-1 and cathepsin-B activit... more The calpain-cathepsin hypothesis posits a key role for elevated calpain-1 and cathepsin-B activity in the neurodegeneration underlying neurotrauma and multiple disorders including Alzheimer's disease (AD). AD clinical trials were recently halted on alicapistat, a selective calpain-1 inhibitor, because of insufficient exposure of neurons to the drug. In contrast to neuroprotection, the ability of calpain-1 and cathepsin-B inhibitors to protect the blood-brain barrier (BBB), is understudied. Since cerebrovascular dysfunction underlies vascular dementia, is caused by ischemic stroke, and is emerging as an early feature in the progression of AD, we studied protection of brain endothelial cells (BECs) by selective and nonselective calpain-1 and cathepsin-B inhibitors. We show these inhibitors protect both neurons and murine BECs from ischemia-reperfusion injury. Cultures of primary BECs from ALDH2 -/- mice that manifest enhanced oxidative stress were sensitive to ischemia, leading to reduced cell viability and loss of tight junction proteins; this damage was rescued by calpain-1 and cathepsin-B inhibitors. In ALDH2 -/- mice 24 h after mild traumatic brain injury (mTBI), BBB damage was reflected by significantly increased fluorescein extravasation and perturbation of tight junction proteins, eNOS, MMP-9, and GFAP. Both calpain and cathepsin-B inhibitors alleviated BBB dysfunction caused by mTBI. No clear advantage was shown by selective versus nonselective calpain inhibitors in these studies. The lack of recognition of the ability of calpain inhibitors to protect the BBB may have led to the premature abandonment of this therapeutic approach in AD clinical trials and requires further mechanistic studies of cerebrovascular protection by calpain-1 inhibitors.

Research paper thumbnail of Novel Pyrrolopyridone Bromodomain and Extra-Terminal Motif (BET) Inhibitors Effective in Endocrine-Resistant ER+ Breast Cancer with Acquired Resistance to Fulvestrant and Palbociclib

Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen re... more Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen receptor positive (ER+) breast cancer. ER is expressed in most resistance settings, thus, bromodomain and extra-terminal protein inhibitors (BETi) that target BET-amplified ER-mediated transcription have therapeutic potential. Novel pyrrolopyridone BETi, leveraged novel interactions with L92/L94 confirmed by a co-crystal structure of 27 with BRD4. Optimization of BETi using growth inhibition in fulvestrant-resistant (MCF-7:CFR) cells was confirmed in endocrine-resistant, palbociclib-resistant, and ESR1 mutant cell lines. 27 was more potent in MCF-7:CFR cells than six BET inhibitors in clinical trials. Transcriptomic analysis differentiated 27 from the benchmark BETi, JQ-1, showing: downregulation of oncogenes; and upregulation of tumor suppressors and apoptosis. The therapeutic approach was validated by oral administration of 27 in orthotopic xenografts of endocrine-resistant breast cancer in monotherapy and in combination with fulvestrant. Importantly, at an equivalent dose in rats, thrombocytopenia was mitigated.

Research paper thumbnail of Methods and Compositions for Measuring Cell Permeability

Research paper thumbnail of Identification of HDAC6‐Selective Inhibitors of Low Cancer Cell Cytotoxicity

ChemMedChem, 2015

The histone deacetylases (HDACs) occur in 11 different isoforms, and these enzymes regulate the a... more The histone deacetylases (HDACs) occur in 11 different isoforms, and these enzymes regulate the activity of a large number of proteins involved in cancer initiation and progression. The discovery of isoform‐selective HDAC inhibitors (HDACIs) is desirable, as it is likely that such compounds would avoid some of the undesirable side effects found with the first‐generation inhibitors. A series of HDACIs previously reported by us were found to display some selectivity for HDAC6 and to induce cell‐cycle arrest and apoptosis in pancreatic cancer cells. In the present work, we show that structural modification of these isoxazole‐based inhibitors leads to high potency and selectivity for HDAC6 over HDAC1–3 and HDAC10, while unexpectedly abolishing their ability to block cell growth. Three inhibitors with lower HDAC6 selectivity inhibit the growth of cell lines BxPC3 and L3.6pl, and they only induce apoptosis in L3.6pl cells. We conclude that HDAC6 inhibition alone is insufficient for disrup...

Research paper thumbnail of Abstract 1464: Patient-derived tumor xenograft are susceptible to formation of B-cell lymphoma after initial transplantation of human carcinoma to immunodeficient mice

Cancer Research, 2015

Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for develop... more Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for developing novel anticancer therapies and personalized use of chemotherapeutic drugs. The PDX approach is based on the transplantation of primary or metastatic human tumors into highly immunodeficient NOD scid gamma (NSG) mice followed by continuous propagation of the established engrafment in mice. Clinicians can evaluate the effects of cancer drugs on their patients’ personalized tumor grafts enabling them to select the best treatment for the cancer patient. Here, we report that PDX tumors engrafted in NSG mice are susceptible to formation of B-cell lymphomas. We xenografted primary or metastatic human tumor fragments into NSG mice and found that tumors generated from patients’ samples of breast (1 out 3 cases), colon (2 out of 6 cases), pancreatic (1 out of 3 cases), bladder (1 out of 2 cases) and renal (1 out of 2 cases) cancer were histologically similar to lymphoid neoplasm. Moreover, we ...

Research paper thumbnail of Outside-In Signaling By VE-Cadherin Promotes Local Rac Activation And Iloprost-Induced Enhancement Of Lung Endothelial Barrier

D27. SIGNALING MOLECULES INVOLVED IN PULMONARY ENDOTHELIAL INJURY AND REPAIR, 2012

Research paper thumbnail of Characterization Of Paxillin Domains Interacting With -Catenin: Implication In Endothelial Barrier Protection

B60. REGULATION OF THE ENDOTHELIUM: BARRIER FUNCTION AND ENDOTHELIAL CELL INTEGRITY, 2010

Research paper thumbnail of Nwani et al., Supplementary figures 1-11 from Melanoma Cells Block PEDF Production in Fibroblasts to Induce the Tumor-Promoting Phenotype of Cancer-Associated Fibroblasts

Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF... more Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF silencing in fibroblasts Supplemental Figure 3. The effect of PEDF expression in fibroblasts on tumor growth Supplemental Figure 4. The effect of endogenous PEDF on fibroblast proliferation and recruitment Supplemental Figure 5. The effect of exogenous TGFb and PDGF on PEDF mRNA in fibroblasts Supplemental Figure 6. The contribution of of TGFb and PDGF to PEDF repression assessed by shRNA silencing Supplemental Figure 7. The effect of TGFb on PDGFR mRNA levels Supplemental Figure 8. Regulation of PDGFB mRNA by TGFb Supplemental Figure 9. The role of PI3K in PEDF control by PDGF-BB Supplemental Figure 10. Inhibition of non-canonical TGFb signaling by PEDF Supplemental Figure 11. Altered expression of select genes in PEDF-null fibroblasts

Research paper thumbnail of C-Terminal Part Of Paxilin Directly Binds Beta-Catenin: Implication In Endothelial Barrier Protection

C26. REGULATORS OF THE LUNG ENDOTHELIAL BARRIER, 2011

Research paper thumbnail of Nwani et al, Supplementary figure legends from Melanoma Cells Block PEDF Production in Fibroblasts to Induce the Tumor-Promoting Phenotype of Cancer-Associated Fibroblasts

Legends for supplementary figures

Research paper thumbnail of Novel Pyrrolopyridone Bromodomain and Extra-Terminal Motif (BET) Inhibitors Effective in Endocrine-Resistant ER+ Breast Cancer with Acquired Resistance to Fulvestrant and Palbociclib

Journal of Medicinal Chemistry, May 26, 2020

Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen re... more Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen receptor positive (ER+) breast cancer. ER is expressed in most resistance settings; thus, bromodomain and extra-terminal protein inhibitors (BETi) that target BET-amplified ER-mediated transcription have therapeutic potential. Novel pyrrolopyridone BETi leveraged novel interactions with L92/L94 confirmed by a cocrystal structure of 27 with BRD4. Optimization of BETi using growth inhibition in fulvestrant-resistant (MCF-7:CFR) cells was confirmed in endocrine-resistant, palbociclib-resistant, and ESR1 mutant cell lines. 27 was more potent in MCF-7:CFR cells than six BET inhibitors in clinical trials. Transcriptomic analysis differentiated 27 from the benchmark BETi, JQ-1, showing downregulation of oncogenes and upregulation of tumor suppressors and apoptosis. The therapeutic approach was validated by oral administration of 27 in orthotopic xenografts of endocrine-resistant breast cancer in monotherapy and in combination with fulvestrant. Importantly, at an equivalent dose in rats, thrombocytopenia was mitigated.

Research paper thumbnail of Abstract 1464: Patient-derived tumor xenograft are susceptible to formation of B-cell lymphoma after initial transplantation of human carcinoma to immunodeficient mice

Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for develop... more Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for developing novel anticancer therapies and personalized use of chemotherapeutic drugs. The PDX approach is based on the transplantation of primary or metastatic human tumors into highly immunodeficient NOD scid gamma (NSG) mice followed by continuous propagation of the established engrafment in mice. Clinicians can evaluate the effects of cancer drugs on their patients’ personalized tumor grafts enabling them to select the best treatment for the cancer patient. Here, we report that PDX tumors engrafted in NSG mice are susceptible to formation of B-cell lymphomas. We xenografted primary or metastatic human tumor fragments into NSG mice and found that tumors generated from patients’ samples of breast (1 out 3 cases), colon (2 out of 6 cases), pancreatic (1 out of 3 cases), bladder (1 out of 2 cases) and renal (1 out of 2 cases) cancer were histologically similar to lymphoid neoplasm. Moreover, we found that the first passage (F1) of some breast and pancreatic cancer PDX tumors could grow as a lymphoid neoplasm in one mouse and as an adenocarcinoma in another mouse after initial transplantation of the pieces from the same human tumor graft (F0). Whereas subcutaneous PDX tumors resembling primary human carcinoma histology were slow growing and non-metastatic, we found that subcutaneous PDX lymphoid neoplasms were fast growing and formed large metastatic lesions in lymph nodes, liver, lungs and spleen of the mouse. Immunohistochemical staining confirmed that lymphoma cells express human leucocyte common antigen CD45 and B-cell antigen CD19/20. By using Epstein-Barr Virus-encoded RNA (EBER) in situ hybridization, we found that PDX lymphoma cells were EBER-positive. Because B-cells are typically present in any solid malignant tumor, B-cell PDX lymphomas may evolve in a wide range of PDX tumor models. Our results suggest that histolopathological evaluation and lymphoid marker testing should be performed as part of the initial model characterization in order to exclude B-cell lymphomas during the development of PDX tumor model from solid human carcinoma. Citation Format: Gennadiy Bondarenko, Andrey Ugolkov, Piotr Kulesza, Stephen M. Rohan, Oleksii Dubrovskyi, Demirkan Gursel, Jeremy V. Mathews, Thomas V. O'Halloran, Jian-Jun Wei, Andrew P. Mazar. Patient-derived tumor xenograft are susceptible to formation of B-cell lymphoma after initial transplantation of human carcinoma to immunodeficient mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1464. doi:10.1158/1538-7445.AM2015-1464

Research paper thumbnail of Potent, Novel SARS-CoV-2 PLpro Inhibitors Block Viral Replication in Monkey and Human Cell Cultures

bioRxiv (Cold Spring Harbor Laboratory), Feb 15, 2021

Antiviral agents blocking SARS-CoV-2 viral replication are desperately needed to complement vacci... more Antiviral agents blocking SARS-CoV-2 viral replication are desperately needed to complement vaccination to end the COVID-19 pandemic. Viral replication and assembly are entirely dependent on two viral cysteine proteases: 3C-like protease (3CLpro) and the papain-like protease (PLpro). PLpro also has deubiquitinase (DUB) activity, removing ubiquitin (Ub) and Ub-like modifications from host proteins, disrupting the host immune response. 3CLpro is inhibited by many known cysteine protease inhibitors, whereas PLpro is a relatively unusual cysteine protease, being resistant to blockade by such inhibitors. A high-throughput screen of biased and unbiased libraries gave a low hit rate, identifying only CPI-169 and the positive control, GRL0617, as inhibitors with good potency (IC50 < 10 µM). Analogues of both inhibitors were designed to develop structure-activity relationships; however, without a co-crystal structure of the CPI-169 series, we focused on GRL0617 as a starting point for structure-based drug design, obtaining several co-crystal structures to guide optimization. A series of novel 2-phenylthiophene-based non-covalent SARS-CoV-2 PLpro inhibitors were obtained, culminating in low nanomolar potency. The high potency and slow inhibitor off-rate were rationalized by newly identified ligand interactions with a "BL2 groove" that is distal from the active site cysteine. Trapping of the conformationally flexible BL2 loop by these inhibitors blocks binding of viral and host protein substrates; however, until now it has not been demonstrated that this mechanism can induce potent and efficacious antiviral activity. In this study, we report that novel PLpro inhibitors have excellent antiviral efficacy and potency against infectious SARS-CoV-2 replication in cell cultures. Together, our data provide structural insights into the design of potent PLpro inhibitors and the first validation that non-covalent inhibitors of SARS-CoV-2 PLpro can block infection of human cells with low micromolar potency.

Research paper thumbnail of Supplemental Figures from Coactivation of Estrogen Receptor and IKKβ Induces a Dormant Metastatic Phenotype in ER-Positive Breast Cancer

This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen rec... more This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen receptor expression, phosphorylation, and activity, cell morphology and EMT. Additional analysis of metastases and stem cell phenotypes are included.

Research paper thumbnail of Supplemental Data Table 1 from Coactivation of Estrogen Receptor and IKKβ Induces a Dormant Metastatic Phenotype in ER-Positive Breast Cancer

This file contains gene expression profiling data for MCF-7-CA-IKKβ cells treated with E2, DOX, E... more This file contains gene expression profiling data for MCF-7-CA-IKKβ cells treated with E2, DOX, E2+DOX.

Research paper thumbnail of Nwani et al., Supplementary figures 1-11 from Melanoma Cells Block PEDF Production in Fibroblasts to Induce the Tumor-Promoting Phenotype of Cancer-Associated Fibroblasts

Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF... more Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF silencing in fibroblasts Supplemental Figure 3. The effect of PEDF expression in fibroblasts on tumor growth Supplemental Figure 4. The effect of endogenous PEDF on fibroblast proliferation and recruitment Supplemental Figure 5. The effect of exogenous TGFb and PDGF on PEDF mRNA in fibroblasts Supplemental Figure 6. The contribution of of TGFb and PDGF to PEDF repression assessed by shRNA silencing Supplemental Figure 7. The effect of TGFb on PDGFR mRNA levels Supplemental Figure 8. Regulation of PDGFB mRNA by TGFb Supplemental Figure 9. The role of PI3K in PEDF control by PDGF-BB Supplemental Figure 10. Inhibition of non-canonical TGFb signaling by PEDF Supplemental Figure 11. Altered expression of select genes in PEDF-null fibroblasts

Research paper thumbnail of Characterization Of Paxillin Domains Interacting With -Catenin: Implication In Endothelial Barrier Protection

Research paper thumbnail of Supplemental Figures from Coactivation of Estrogen Receptor and IKKβ Induces a Dormant Metastatic Phenotype in ER-Positive Breast Cancer

This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen rec... more This file contains supplemental data related to T47D-CA-IKKβ and control cell lines, estrogen receptor expression, phosphorylation, and activity, cell morphology and EMT. Additional analysis of metastases and stem cell phenotypes are included.

Research paper thumbnail of Rap1 and afadin promote interactions between adherens junctions and tight junctions and regulate endothelial permeability

Research paper thumbnail of Novel Rap1‐dependent regulator of endothelial cell junctions afadin mediates protective effects by oxidized phospholipids in the models of acute lung injury

Research paper thumbnail of Extending the Calpain–Cathepsin Hypothesis to the Neurovasculature: Protection of Brain Endothelial Cells and Mice from Neurotrauma

ACS Pharmacology & Translational Science, 2021

The calpain-cathepsin hypothesis posits a key role for elevated calpain-1 and cathepsin-B activit... more The calpain-cathepsin hypothesis posits a key role for elevated calpain-1 and cathepsin-B activity in the neurodegeneration underlying neurotrauma and multiple disorders including Alzheimer's disease (AD). AD clinical trials were recently halted on alicapistat, a selective calpain-1 inhibitor, because of insufficient exposure of neurons to the drug. In contrast to neuroprotection, the ability of calpain-1 and cathepsin-B inhibitors to protect the blood-brain barrier (BBB), is understudied. Since cerebrovascular dysfunction underlies vascular dementia, is caused by ischemic stroke, and is emerging as an early feature in the progression of AD, we studied protection of brain endothelial cells (BECs) by selective and nonselective calpain-1 and cathepsin-B inhibitors. We show these inhibitors protect both neurons and murine BECs from ischemia-reperfusion injury. Cultures of primary BECs from ALDH2 -/- mice that manifest enhanced oxidative stress were sensitive to ischemia, leading to reduced cell viability and loss of tight junction proteins; this damage was rescued by calpain-1 and cathepsin-B inhibitors. In ALDH2 -/- mice 24 h after mild traumatic brain injury (mTBI), BBB damage was reflected by significantly increased fluorescein extravasation and perturbation of tight junction proteins, eNOS, MMP-9, and GFAP. Both calpain and cathepsin-B inhibitors alleviated BBB dysfunction caused by mTBI. No clear advantage was shown by selective versus nonselective calpain inhibitors in these studies. The lack of recognition of the ability of calpain inhibitors to protect the BBB may have led to the premature abandonment of this therapeutic approach in AD clinical trials and requires further mechanistic studies of cerebrovascular protection by calpain-1 inhibitors.

Research paper thumbnail of Novel Pyrrolopyridone Bromodomain and Extra-Terminal Motif (BET) Inhibitors Effective in Endocrine-Resistant ER+ Breast Cancer with Acquired Resistance to Fulvestrant and Palbociclib

Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen re... more Acquired resistance to fulvestrant and palbociclib is a new challenge to treatment of estrogen receptor positive (ER+) breast cancer. ER is expressed in most resistance settings, thus, bromodomain and extra-terminal protein inhibitors (BETi) that target BET-amplified ER-mediated transcription have therapeutic potential. Novel pyrrolopyridone BETi, leveraged novel interactions with L92/L94 confirmed by a co-crystal structure of 27 with BRD4. Optimization of BETi using growth inhibition in fulvestrant-resistant (MCF-7:CFR) cells was confirmed in endocrine-resistant, palbociclib-resistant, and ESR1 mutant cell lines. 27 was more potent in MCF-7:CFR cells than six BET inhibitors in clinical trials. Transcriptomic analysis differentiated 27 from the benchmark BETi, JQ-1, showing: downregulation of oncogenes; and upregulation of tumor suppressors and apoptosis. The therapeutic approach was validated by oral administration of 27 in orthotopic xenografts of endocrine-resistant breast cancer in monotherapy and in combination with fulvestrant. Importantly, at an equivalent dose in rats, thrombocytopenia was mitigated.

Research paper thumbnail of Methods and Compositions for Measuring Cell Permeability

Research paper thumbnail of Identification of HDAC6‐Selective Inhibitors of Low Cancer Cell Cytotoxicity

ChemMedChem, 2015

The histone deacetylases (HDACs) occur in 11 different isoforms, and these enzymes regulate the a... more The histone deacetylases (HDACs) occur in 11 different isoforms, and these enzymes regulate the activity of a large number of proteins involved in cancer initiation and progression. The discovery of isoform‐selective HDAC inhibitors (HDACIs) is desirable, as it is likely that such compounds would avoid some of the undesirable side effects found with the first‐generation inhibitors. A series of HDACIs previously reported by us were found to display some selectivity for HDAC6 and to induce cell‐cycle arrest and apoptosis in pancreatic cancer cells. In the present work, we show that structural modification of these isoxazole‐based inhibitors leads to high potency and selectivity for HDAC6 over HDAC1–3 and HDAC10, while unexpectedly abolishing their ability to block cell growth. Three inhibitors with lower HDAC6 selectivity inhibit the growth of cell lines BxPC3 and L3.6pl, and they only induce apoptosis in L3.6pl cells. We conclude that HDAC6 inhibition alone is insufficient for disrup...

Research paper thumbnail of Abstract 1464: Patient-derived tumor xenograft are susceptible to formation of B-cell lymphoma after initial transplantation of human carcinoma to immunodeficient mice

Cancer Research, 2015

Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for develop... more Patient-derived xenograft (PDX) tumor models have emerged as new and effective models for developing novel anticancer therapies and personalized use of chemotherapeutic drugs. The PDX approach is based on the transplantation of primary or metastatic human tumors into highly immunodeficient NOD scid gamma (NSG) mice followed by continuous propagation of the established engrafment in mice. Clinicians can evaluate the effects of cancer drugs on their patients’ personalized tumor grafts enabling them to select the best treatment for the cancer patient. Here, we report that PDX tumors engrafted in NSG mice are susceptible to formation of B-cell lymphomas. We xenografted primary or metastatic human tumor fragments into NSG mice and found that tumors generated from patients’ samples of breast (1 out 3 cases), colon (2 out of 6 cases), pancreatic (1 out of 3 cases), bladder (1 out of 2 cases) and renal (1 out of 2 cases) cancer were histologically similar to lymphoid neoplasm. Moreover, we ...

Research paper thumbnail of Outside-In Signaling By VE-Cadherin Promotes Local Rac Activation And Iloprost-Induced Enhancement Of Lung Endothelial Barrier

D27. SIGNALING MOLECULES INVOLVED IN PULMONARY ENDOTHELIAL INJURY AND REPAIR, 2012

Research paper thumbnail of Characterization Of Paxillin Domains Interacting With -Catenin: Implication In Endothelial Barrier Protection

B60. REGULATION OF THE ENDOTHELIUM: BARRIER FUNCTION AND ENDOTHELIAL CELL INTEGRITY, 2010

Research paper thumbnail of Nwani et al., Supplementary figures 1-11 from Melanoma Cells Block PEDF Production in Fibroblasts to Induce the Tumor-Promoting Phenotype of Cancer-Associated Fibroblasts

Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF... more Supplemental Figure 1. Verification of PEDF knockdown Supplemental Figure 2. Verification of PEDF silencing in fibroblasts Supplemental Figure 3. The effect of PEDF expression in fibroblasts on tumor growth Supplemental Figure 4. The effect of endogenous PEDF on fibroblast proliferation and recruitment Supplemental Figure 5. The effect of exogenous TGFb and PDGF on PEDF mRNA in fibroblasts Supplemental Figure 6. The contribution of of TGFb and PDGF to PEDF repression assessed by shRNA silencing Supplemental Figure 7. The effect of TGFb on PDGFR mRNA levels Supplemental Figure 8. Regulation of PDGFB mRNA by TGFb Supplemental Figure 9. The role of PI3K in PEDF control by PDGF-BB Supplemental Figure 10. Inhibition of non-canonical TGFb signaling by PEDF Supplemental Figure 11. Altered expression of select genes in PEDF-null fibroblasts

Research paper thumbnail of C-Terminal Part Of Paxilin Directly Binds Beta-Catenin: Implication In Endothelial Barrier Protection

C26. REGULATORS OF THE LUNG ENDOTHELIAL BARRIER, 2011

Research paper thumbnail of Nwani et al, Supplementary figure legends from Melanoma Cells Block PEDF Production in Fibroblasts to Induce the Tumor-Promoting Phenotype of Cancer-Associated Fibroblasts

Legends for supplementary figures