Research Highlight


In this image, I am immunoblotting with p130, E2F4, and E2F1 antibodies. I incubated the primary antibodies overnight, and washed each nitrocellulose membrane with TBST the following morning. I visualized the antibody binding by using secondary antibody conjugated to Horse Radish Peroxidase and measured chemiluminescence. This experiment can tell me relative amounts of p130, E2F4, and E2F1 in different experimental conditions.

Analysis of DREAM and E2F1 Competition for Cell Cycle Promoters during G1.

The DREAM complex maintains G0 by binding to the sequence specific E2F and cell cycle gene homology region (CHR) promoter elements to repress cell cycle gene expression. The DREAM complex is composed of DP1, retinoblastoma-like p130 or p107, E2F4 or E2F5, and MuvB, which is comprised of LIN37, LIN52, LIN9, LIN54, and RBBP4. The retinoblastoma tumor suppressor protein, RB, cooperates with DREAM to repress cell cycle gene expression by binding and repressing transactivation of gene expression by activator E2F1. RB and DREAM respond similarly to growth factors and oncogenic stimuli, but they are phosphorylated differently. RB loses its binding to E2F1 after it is hypo-phosphorylated by Cyclin D-CDK4/6 in early G1 and hyperphosphorylated by Cyclin E-CDK2 at the late G1 restriction point, allowing for E2F1-driven expression of early cell cycle genes. In contrast, Cyclin D-CDK4/6 phosphorylation of p130 alone results in DREAM complex disruption. However, the mechanism of loss of E2F4 from cell cycle promoters is not known. Temporally, E2F4 is present on cell cycle promoters during early/mid G1 while E2F1-RB is present later during mid-late G1. We hypothesize that E2F4 and E2F1 compete with each other for the E2F promoter elements, with E2F1 having a higher binding affinity. Additionally, E2F1 is removed from cell cycle promoters during G1 by Cyclin A-CDK2 phosphorylation. Because these sites are conserved in E2F4, we hypothesize that Cyclin E-CDK2 phosphorylates E2F4, resulting in loss of E2F4 binding on cell cycle promoters. Cyclin E-CDK2 is active when E2F4 loses binding to cell cycle promoters, so we hypothesize that E2F4 is phosphorylated by this cyclin. The focus of my research is to determine if E2F1 competes with E2F4 for E2F promoter elements, and if E2F1 a higher affinity for these elements. We will clone TFDP1 and E2F4 into a bacterial protein expression vector called pGEX, which has an N-terminal GST tag. We will include TFDP1 (protein name DP1) because heterodimerization of E2F1-DP1 and E2F4-DP1 is required for DNA binding. We will express the proteins in E.coli BL21*(DE3) E.coli. BL21 STAR (DE3) is the best strain of E.coli for our purposes since it has higher mRNA stability and lower protease activity. We will purify the bacterially expressed, N-terminally GST-tagged proteins using immobilized glutathione beads. To test if in the presence of E2F1 DNA-bound E2F4 levels will drop or stay the same, we will perform a DNA-binding affinity assay. We will do this by conjugating biotin to dsDNA sequence with the MCM5 gene promoter that contains E2F and CHR elements. We will pull down DNA using streptavidin beads and qualitatively measure bound protein by immunoblot. To test if Cyclin E-CDK2 affects E2F4 binding affinity, we will incubate E2F4 with Cyclin E-CDK2 and then measure DNA binding affinity. These experiments will give us insight into how DREAM and RB coordinate to repress G1/S phase genes.

 

 

Trainee Research

CaNCURE provides trainees with a 6-month hands-on research experience and one-on-one mentoring by leading researchers in cancer nanomedicine.   Projects performed by current and past participants include:

Radiation enhancement in cancer cells using gold and gadolinium nanoparticles

Targeting CXCR4/SDF-1a using phytochemicals to inhibit progression and metastasis of pancreatic cancer

Targeting WASp using Wiskostatin-gold nanoparticles

Combined delivery of targeted liposomal chemotherapeutics and photodynamic therapy to treat pancreatic cancer

Use of a Triblock Copolymer Hydrogel for Controlled Release of Cisplatin and BMN-673

Co-delivery of protective substrate and chemotherapy drugs via lipid Bilayer Mesoporous Silica Nanoparticles

Software with built-in neuroanatomy atlas provides insight into cancer treatment

Discovering the Genetic Mechanism of Enhanced Metastatic Colonization in SMAD4 Mutant Tumors

Nano-plasmonic exosome (nPLEX) assays for exosome analysis and antibody validation

Surface-targeting, ligand-switching nanoparticles for mitochondrial drug delivery in prostate cancer

Iron-chelating PEG-like nanoprobes as therapeutic and 89Zr/PET imaging agents

Online monitoring and image-guided treatment of chemoresistant micrometastases

Co-delivery of antibiotics and topoisomerase inhibitors to overcome chemoresistance

Optimization of macrophage-targeted nanoparticles for positron emission tomography imaging in cancer

Development of a novel nanogel for non-invasive transdermal delivery of cancer vaccines using hyaluronic acid

Development of a Point of Care Assay for Detecting High Risk HPV in Resource Limited Settings

Delivery of Titanium Dioxide Nanoparticles via Biodegradable PLGA Dissolving Microneedles for Sustainable Release

Combined Cisplatin and Olaparib nanoparticles for ovarian cancer therapy

Longitudinal assessment of tumor heterogeneity during immunotherapy for metastatic melanoma

Assessment of Atherosclerotic Changes using Ferumoxytol as MRI Contrast Agent

Protein-encapsulated nanoparticles for oral delivery of anti-mitotic agents in prostate cancer

Implementation of novel MR-based attenuation correction in PET/MR pelvic scans

Analysis of DREAM and E2F1 Competition for Cell Cycle Promoters during G1.

Use of CT Texture Analysis for Characterization and Prognostication of Incidental Adnexal Lesions

Tracking pancreatic adenocarcinoma response to treatment using targeted, multi-modal nanoparticles

Soleil Doggett (Biology, '16) talks to her fellow peers about her research on oxygenating tumors to stimulate the anti-tumor immune response.

 

Trainee e-portfolios

Photo credit: Tom Kates Photography

While on co-op, trainees document their research in an e-portfolio.  This gives trainees the opportunity to provide regular updates on their research progress, reflect on training they are receiving, and explain how their research fits within the field of cancer nanomedicine.  These research e-portfolios can be accessed through individual trainee profiles.  The complete collection may be found here.

 

Presentation at CaNCURE Nanomedicine Day

At the completion of their co-op, trainees are provided with the opportunity to present their research to a wider audience.  For our June CaNCURE Nanomedicine Day, trainees prepare interactive, digital posters to display on electronic poster boards.  Over 100 faculty, students, and researchers attend this annual event!

Check out the news article about our first CaNCURE Day!

Trainee Publications

Our Trainees have published 22 peer-reviewed since January 2015.  A full list of Trainee publications is found below.

  1. Patrick Sheedy, Zdravka Medarova. The fundamental role of miR-10b in metastatic cancer. Am J Cancer Res 2018;8(9):1674-1688. Link
  2. Chen X, Ling X, Zhao L, Xiong F, Hollett G, Kang Y, Barrett A, Wu J. “Biomimetic Shells Endow Sub-50 nm Nanoparticles with Ultrahigh Paclitaxel Payloads for Specific and Robust Chemotherapy.”  ACS Appl Mater Interfaces. 2018 Sep 25. doi: 10.1021/acsami.8b11571. PMID: 30203956  Link
  3. Hedgire S, Krebill C, Wojtkiewicz GR, Oliveira I, Ghoshhajra BB, Hoffmann U, Harisinghani MG. “Ultrasmall superparamagnetic iron oxide nanoparticle uptake as noninvasive marker of aortic wall inflammation on MRI: proof of concept study.”   Br J Radiol. 2018 Sep 12:20180461. doi: 10.1259/bjr.20180461. PMID: 30160173  Link
  4. Application of the BLADE Sequence in Upper Abdominal MR Imaging. Krebill C.  Radiol Technol. 2018 May;89(5):495-497. PMID:29793909 Link
  5. Torrado-Carvajal A, Vera-Olmos J, Izquierdo-Garcia D1, Catalano OA, Morales MA, Margolin J, Soricelli A, Salvatore M, Malpica N, Catana C1. Dixon-VIBE Deep Learning (DIVIDE) Pseudo-CT Synthesis for Pelvis PET/MR Attenuation Correction. J Nucl Med. 2018 Aug 30. pii: jnumed.118.209288. doi: 10.2967/jnumed.118.209288. PMID: 30166357  Link
  6. Xiaoyuan Ji, Jie Wang, Lin Mei, Wei Tao, Austin Barrett, Zhiguo Su, Shaomin Wang. Guanghui Ma, Jinjun Shi, Songping Zhang. Artificial Photosynthesis: Porphyrin/SiO2 /Cp*Rh(bpy)Cl Hybrid Nanoparticles Mimicking Chloroplast with Enhanced Electronic Energy Transfer for Biocatalyzed Artificial Photosynthesis. Advanced Functional Materials. Link
  7. Yang KS, Im H, Hong S, Pergolini I, Del Castillo AF, Wang R, Clardy S, Huang CH, Craig Pille, Ferrone, Yang R, Castro CM, Lee H, Del Castillo CF, Weissleder R. Multiparametric plasma EV profiling facilitates diagnosis of pancreatic malignancy. Sci Transl Med. 2017; 9(391): eaal3226. PMC5846089
  8. Zhu X, Ji X, Kong N, Chen Y, Mahmoudi M, Xu X, Ding L, Tao W, Cai T, Li Y, Gan T, Austin Barrett, Bharwani Z, Chen H, Farokhzad OC. Intracellular Mechanistic Understanding of 2D MoS2 Nanosheets for Anti-Exocytosis-Enhanced Synergistic Cancer Therapy.  ACS Nano. 2018 Mar 27;12(3):2922-2938. PMC6097229
  9. Miller MA1, Kim E, Cuccarese MF, Alec Plotkin, Prytyskach M, Kohler RH, Pittet MJ, Weissleder R. “Near infrared imaging of Mer tyrosine kinase (MERTK) using MERi-SiR reveals tumor associated macrophage uptake in metastatic disease.” Chem Commun. 2017 Dec 19;54(1):42-45. PMC5736449
  10. Ding L, Zhu X, Wang Y, Shi B, Ling X, Chen H, Nan W, Austin Barrett, Guo Z, Tao W, Wu J, Shi X. “Intracellular Fate of Nanoparticles with Polydopamine Surface Engineering and a Novel Strategy for Exocytosis-Inhibiting, Lysosome Impairment-Based Cancer Therapy”. Nano Lett. 2017 Nov 8;17(11):6790-6801.  PMC6071871
  11. Yoo B, Ann-Marie, Billig, Medarova Z. “Guidelines for Rational Cancer Therapeutics. Frontiers in Oncology Journal”. Front Oncol. 2017 Dec 12;7:310. PMC5732930
  12. Gharagouzloo C, Timms L, Qiao J, Fang Z, Joseph Nneji, Pandya A, Kulkarni P, van de Ven AL, Ferris C, Sridhar S. “Neural circuits and brain function: New insights using quantitative vascular mapping of the rat.” Neuroimage, 2017. 16C:24-33  PMC5824692
  13. Gharagouzloo C, Timms L, Qiao J, Fang Z, Joseph Nneji, Pandya A, Kulkarni P, van de Ven AL, Ferris C, Sridhar S.   “Dataset on a 173 region awake resting state quantitative cerebral blood volume rat brain atlas and regional changes to cerebral blood volume under isoflurane anesthetization and CO2 challenge”. Data in Brief, 2018. 17:393-396.  Link
  14. Qin L, Li A, Qu J, Reinshagen K, Li X, Cheng S, Annie Bryant, Young GS. Normalization of ADC does not improve correlation with overall survival in patients with high-grade glioma (HGG). J Neurooncol. 2018 Apr;137(2):313-319.   PMC6071871
  15. Belz J, Kumar R, Baldwin P, Noelle Castilla Ojo, Leal AS, Royce DB, Di Zhang D, van de Ven AL, Liby K, Sridhar S. “Sustained-release Talazoparib implants for localized treatment of BRCA1-deficient breast cancer”. Theranostics, 7(17): 4340-4349.  PMC5695017
  16. Qin L, Li X, Amanda Stroiney, Qu J, Helgager J, Reardon DA, Young GS. “Advanced MRI assessment to predict benefit of anti-programmed cell death 1 protein immunotherapy response in patients with recurrent glioblastoma.” 2017 Feb;59(2):135-145.  PMC6097616
  17. Jodi Belz, Noelle Castilla Ojo,Srinivas Sridhar, Rajiv Kumar.  Radiosensitizing silica nanoparticles encapsulating docetaxel for treatment of prostate cancer, In Cancer Nanotechnology. Reema Zeineldin (Ed).  Series: Methods in Molecular Biology. Springer Press. Methods Mol Biol. 2017; 1530:403-409. PMC5531609
  18. Christian Berrios, Megha Padi, Mark A. Keibler, Donglim Esther Park, Vadim Molla, Gregory Stephanopoulos, John Quackenbush, James A. DeCaprio. “Merkel cell polyomavirus small T antigen promotes pro-glycolytic metabolic perturbations required for transformation”. 2016 Nov 23;12(11):e1006020.   PMC5120958
  19. Song C, Liu Y, Rachel Fontana, Makrigiorgos A, Mamon H, Kulke MH, G. Mike Makrigiorgos. “Elimination of unaltered DNA in mixed clinical samples via nuclease-assisted minor-allele enrichment”.  2016 Nov 2;44(19):e146. PMC5100565
  20. Andrew L. Hong, Yuen-Yi Tseng, Glenn S. Cowley, Oliver Jonas, Jaime H. Cheah, Bryan D. Kynnap, Mihir B. Doshi, Coyin Oh, Stephanie C. Meyer, Alanna J. Church, Shubhroz Gill, Craig M. Bielski, Paula Keskula, Alma Imamovic, Sara Howell, Gregory V. Kryukov, Paul A. Clemons, Aviad Tsherniak, Francisca Vazquez, Brian D. Crompton, Alykhan F. Shamji, Carlos Rodriguez-Galindo, Katherine A. Janeway, Charles W. M. Roberts, Kimberly Stegmaier, Paul van Hummelen,
Michael J. Cima, Robert S. Langer, Levi A. Garraway, Stuart L. Schreiber, David E. Root,
William C. Hahn, & Jesse S. Boehm. “Integrated genetic and pharmacologic interrogation of rare cancers”. Nat Commun. 2016 Jun 22;7:11987.  PMC4917959
  21. Wang P, Yoo B, Sherman S, Mukherjee P, Ross A, Pantazopoulos P, Petkova V, Farrar C, Medarova Z, Moore A. “Predictive imaging of chemotherapeutic response in a transgenic mouse model of pancreatic cancer.” Int J Cancer. 2016 Aug 1;139(3):712-8. PMCID: PMC4925171
  22. Nazila Kamaly, Gabrielle Fredman, Jhalique J. Fojas, Manikandan Subramanian, Won II Choi, Katherine Zepeda, Cristian Vilos, Mikyung Yu, Suresh Gadde, Jun Wu, Jaclyn Milton, Renata Leitao, Livia Rosa, Moaraj Hasan, Huayi Gao, Vance Nguyen, Jordan Harris, Ira Tabas, and Omid C. Farokhzad. “Interleukin-10 Targeted Nanotherapeutics Developed with a Microfluidic Chip Enhance Resolution of Inflammation in Advanced Atherosclerosis”. ACS Nano. 2016 May 24;10(5):5280-92. PMC5199136