Robert Babak Faryabi, Ph.D.

Deciphering Mechanisms of Genome Mis-folding In Cancer

Our lab deploys data-rich experimental techniques to elucidate the role of genome mis-folding in controlling oncogenic gene expression programs. Specifically, we are interested in moving beyond the status quo to understand how oncogenic subversion of lineage-determining transcription factors set topology of cancer genome. To tackle this question, we combine genomics and super-resolution imaging and focus on investigating molecular mechanisms of genome mis-folding in breast and blood cancers. These mechanistic studies aim to identify precise epigenetic vulnerabilities of cancer cells and guide treatments disrupting cancer cells’ transcriptional addiction.

Representative Publication:

Oncogenic Notch Promotes Long-Range Regulatory Interactions Within Hyperconnected 3D Cliques. Petrovic J*, Zhou Y*, Fasolino M, Goldman N, Schwartz GW, Mumbach MR, Nguyen SC, Rome KS, Sela Y, Zapataro Z, Blacklow SC, Kruhlak MJ, Shi J, Aster JC, Joyce EF, Little SC, Vahedi G, Pear WS, Faryabi RB Molecular Cell. 2019;73(6):1174-90 e12

Determining Epigenetic Mechanisms Of Resistance To Targeted Therapies

Targeting oncogenic drivers of cancers commonly leads to drug resistance. Mechanisms of acquiring resistance to oncology drugs mostly remain unknown, partly due to the limitations of population-based assays in elucidating heterogeneity of drug-naive and complexity of drug-induced tumor evolution. Using single-cell genomics and imaging, we study how heterogeneity and plasticity of transcriptional dependencies confer resistance to targeted therapeutics such as Notch inhibitors.

Representative Publication

TooManyCells Identifies And Visualizes Relationships Of Single-cell Clades. Schwartz GW, Zhou Y, Petrovic J, Fasolino M, Xu L, Shaffer SM, Pear WS, Vahedi G, Faryabi RB Nature Methods, 2020; 17: 405-413

Innovating Computational Methods To Enable Cancer Discovery

Our lab innovates statistical and machine learning approaches to accelerate discovery of novel therapeutics and biomarkers by elucidating complexity and heterogeneity of tumors. Recently, we have developed a computational ecosystem for mapping molecular and spatial heterogeneity in tumors. As part of the Center for Personalized Diagnostics, we also mine cancer patient genotypic/phenotypic data to improve patient health. patient health.

Representative Publication:

Classes of ITD Predict Outcomes in AML Patients Treated With FLT3 Inhibitors. Schwartz GW, Manning B, Zhou Y, Velu P, Bigdeli A, Astles R, Lehman AW, Morrissette JJD, Perl AE, Li M, Carroll M, Faryabi RB Clinical Cancer Research. 2019;25(2):573-83

Research Interest

Cancer is typically considered a genetic disease. However, recent progress in our understanding of epigenetic aberrations in cancer has challenged this view. Overarching goal of our lab is to understand epigenetic mechanisms of transcriptional addiction in cancer and exploit this information to advance cancer therapeutics.

To pursue this objective, we use cutting-edge chromatin conformation capture, high-content imaging, single-cell epigenomics, functional genomics, and combine these technologies with our expertise in computational sciences to systematically explore: i) how epigenetic control of gene expression is disrupted in cancer, ii) why transcriptional addiction can develop, and iii) how heterogeneity and plasticity of transcriptional dependencies enable drug resistance.

Brian C. Capell, M.D., Ph.D.

  • Demonstrated the first role for ferroptosis in epidermal differentiation and tumor suppression: Our recent studies have uncovered a potential link between the emerging form of programmed cell death known as ferroptosis and the execution of both epidermal differentiation and tumor suppression in the skin. Beyond identifying the mechanisms through which MLL4 (KMT2D) promotes differentiation and exerts its tumor suppressive functions, these studies have revealed evidence that ferroptosis may be the essential mechanism by which keratinocytes ultimately die and form the cornified envelope of the epidermal barrier. Given that numerous skin disorders are driven by dysregulated epidermal differentiation, combined with the ability to both pharmacologically induce and inhibit ferroptosis, these studies have provided proof of principal that ferroptosis modulation may a viable therapeutic strategy for a variety of skin disorders.

a.    Egolf S, Zou J, Anderson A, Simpson CL, Aubert Y, Prouty S, Ge K, Seykora JT, Capell BC. MLL4 mediates differentiation and tumor suppression through ferroptosis. Sci Adv. 2021 Dec 10;7(50):eabj9141. doi: 10.1126/sciadv.abj9141. Epub 2021 Dec 10. PMID: 34890228; PMCID: PMC8664260.

Research Interest

The Capell Lab seeks to understand epigenetic gene regulatory mechanisms, how they interface with metabolism and the immune system, and how when disrupted, they may contribute to disease, and in particular, cancer. By combining the incredible accessibility of human skin with the most cutting-edge techniques, we aim to identify therapeutic vulnerabilities in cancer, and novel targets to treat disease.

George Burslem, Ph.D.

Targeted Protein Degradation
Approximately 75% of the proteome is considered undruggable by traditional small molecule inhibition approaches. As an LLS postdoctoral fellow at Yale, I contributed to the development and application of Proteolysis Targeting Chimera (PROTACs) – heterobifunctional small molecules which recruit an E3 ligase to a protein target, resulting in target ubiquitination and subsequent degradation via the proteasome. This enables small molecule induced degradation of disease relevant proteins in native systems including in vivo. One of my contributions was to expand this approach to transmembrane proteins (specifically receptor tyrosine kinases) which are not endogenously degraded via the proteasome but can be degraded in this manner under small molecule control. The PROTAC approach has also been applied to crucial targets (BCR-Abl and FLT-3 ITD) in hematological malignancies demonstrating the power of the PROTAC approach for drug discovery and revealing previously unknown scaffolding roles of these kinases.
– Proteolysis-Targeting Chimeras as Therapeutics and Tools for Biological Discovery, G.M. Burslem and C.M. Crews, Cell, 2020, 181, 102. PMCID: PMC7319047
– Targeting BCR-ABL1 in Chronic Myeloid Leukemia by PROTAC-mediated Targeted Protein Degradation, G.M. Burslem, A. Reister-Schultz, D.P. Bondeson, C. Eide, S. Savage, B. Druker and C.M. Crews, Cancer Research, 2019, 79. 4744. PMCID: PMC6893872
– Enhancing Antiproliferative Activity and Selectivity of a FLT-3 Inhibitor by Proteolysis Targeting Chimera Conversion, G.M. Burslem, J. Song, X. Chen, J. Hines and C.M. Crews, J. Am. Chem. Soc., 2018, 140, 16428. PMID: 30427680
– The Advantages of Targeted Protein Degradation over Inhibition: an RTK Case Study, G.M. Burslem, B.E. Smith, A. Lai, S. Jaime-Figueroa, D. McQuaid, D.P. Bondeson, M. Toure, H. Dong, Y. Qian, J. Wang, A.P. Crew, J. Hines and C. M. Crews, Cell Chemical Biology, 2018, 25, 67. PMCID: PMC5777153

HIF-1α/p300 Inhibition
Mammalian cells have developed an elaborate pathway for oxygen sensing with a key player in this pathway being hypoxia inducible factor 1α (HIF-1α). Under hypoxic conditions, HIF-1α accumulates, heterodimerizes and translocated to the nucleus where it forms a protein-protein interaction with p300. This complex is transcriptionally active resulting in the hypoxic response and resupply of oxygen to the hypoxic tissue. This pathway is crucial for growth and development but is also exploited by solid tumors to enable growth to continue after exhaustion of their oxygen supply. As a graduate student, I focused on elucidating the molecular recognition between HIF-1α and p300 using biophysical and biochemical approaches before applying that knowledge to develop the first biophysically characterized inhibitors of this protein-protein interaction. Furthermore, this project led to the identification of novel peptide and protein aptamer inhibitors of the HIF-1α/p300 interaction via phage display and a novel class of chemical probes which incorporates both natural and unnatural recognition elements to provide enhanced selectivity and potency.
– Hypoxia Inducible Factor as a Model for Studying Inhibition of Protein-Protein Interactions, G.M. Burslem, H.F. Kyle, A.S. Nelson, T.A. Edwards, A.J. Wilson, Chemical Science, 2017, 8, 4188. PMCID: PMC5576430
– Towards “Bionic” Proteins: Replacement of Continuous Sequences from HIF-1α with Proteomimetics to Create Functional p300 Binding HIF-1α Mimics, G.M. Burslem, H.F. Kyle, A. L. Breeze, T.A. Edwards, S.L. Warriner, A. S. Nelson and A.J. Wilson, Chem. Commun., 2016, 52, 5421. PMCID: PMC4843846
– Small molecule proteomimetic inhibitors of the HIF-1α/p300 protein-protein interaction, G.M. Burslem, H. Kyle, A. Breeze, T.A. Edwards, A. Nelson, S.L. Warriner and A.J. Wilson, ChemBioChem, 2014, 15, 1083. PMCID: PMC4159589
– Exploration of the HIF-1α/p300 binding interface using peptide and adhiron phage display technologies to locate binding hot-spots for inhibitor development, H. F. Kyle, K. F. Wickson, J. Stott, G. M. Burslem, A. L. Breeze, D. C. Tomlinson, S. L. Warriner, A. Nelson, A. J. Wilson and T. A. Edwards, Mol. Biosyst., 2015, 11, 2738. PMID: 26135796

Modulating Protein-Protein Interactions
There are an estimated 650,000 pairwise protein-protein interactions in the human interactome and these interactions are implicated in all biological pathways. As such, the ability to modulate protein-protein interactions with chemical probes can provide unique insights in the functional roles of proteins and their binding partners. Over my career I have developed chemical biology approaches to both inhibit and induce protein-protein interactions as tool compounds and therapeutic approaches. An attractive approach to the inhibition of protein-protein interactions is to develop molecules which mimic a portion of one of protein binding partners and thus preferentially occupy the binding site on the other. Peptides are capable of doing this but must pay a significant entropic penalty to adopt the required conformation due to their inherent flexibility. One approach to combat this is to pre-organize the peptide into the desired conformation by chemically “stapling” it. Another approach is to develop small molecule mimetics capable of recapitulating the recognition elements of a protein/peptide but with less conformational flexibility. I have applied both of these techniques to generate inhibitors of a variety of protein-protein interactions including p53/hDM2, Bcl-XL/BID and RNase S-peptide/S-protein. Furthermore, protein-protein interactions can be induced by small molecules known as molecular glues which we have employed to stabilize interactions between Cereblon and IKZF1. We have also demonstrated the ability to induce protein-protein interactions between various proteins using heterobifunctional compounds.
– Double Quick, Double “Click” Reversible Peptide “Stapling”, C.M. Grison, G.M. Burslem, J.A. Miles, L. Pilsl, D.J. Yeo, S.L. Warriner, M. E. Webb and A. J. Wilson, Chemical Science, 2017, 8, 5166. PMCID: PMC5618791
– Synthesis of Highly Functionalized Oligobenzamide Proteomimetic Foldamers by Late-Stage Introduction of Sensitive Groups, G.M. Burslem, H.F. Kyle, P. Prabhakaran, A. L. Breeze, T.A. Edwards, S.L. Warriner, A. Nelson and A.J. Wilson, Org. Biomol. Chem. 2016, 14, 3782. PMCID: PMC4839272
– Efficient Synthesis of Immunomodulatory Drug Analogues Enables Exploration of Structure Degradation Relationships. G.M. Burslem*, P. Ottis, S. Jaime-Figueroa, A. Morgan, P.M. Cromm, M. Toure and C.M. Crews*, ChemMedChem, 2018, 12, 1508 (*Co-corresponding authors). PMCID: PMC6291207
– Lessons on Selective Degradation with a Promiscuous Warhead: Informing PROTAC Design, D.P. Bondeson, B.E. Smith, G.M. Burslem, A.D. Buhimschi, J. Hines, S. Jaime-Figueroa, J. Wang, B. Hamman, A. Ishchenko, C.M. Crews, Cell Chemical Biology, 2018, 25, 78. PMCID: PMC5777153

Epigenetic Chemical Biology
Epigenetic drug discovery provides a wealth of opportunities for the discovery of new therapeutics but has been hampered by low hit rates, frequent identification of false-positives, and poor synthetic tractability. Since establishing my laboratory at the University of Pennsylvania, we have endeavored to remedy the low hit rate in drug discovery efforts against epigenetic targets, by careful chemo-informatic analysis of active compounds and screening libraries. We have used the information gathered in these analysis to inform the design and synthesis of privileged compound collections for epigenetic chemical biology and probe discovery.
– Photochemical Synthesis of an Epigenetic Focused Tetrahydroquinoline Library, A.I. Green and G.M. Burslem, RSC Medicinal Chemistry, 2021, DOI: 10.1039/D1MD00193K
– Focused Libraries for Epigenetic Drug Discovery: The Importance of Isosteres, A.I. Green and G.M. Burslem, J. Med. Chem, 2021, 64, 7231-7240. PMID: 34042449
– Advances and Opportunities in Epigenetic Chemical Biology, J. Beyer, N. Raniszewski and G.M. Burslem, ChemBioChem, 2021, 22, 17-42. PMID: 32786101

Research Interest

The Burslem lab is interested in developing chemical tools to understand and modulate lysine post-translational modifications, specifically acetylation and ubiquitination. The laboratory is particularly interested in novel pharmacological approaches to modulate post-translational modifications which regulate gene expression and protein stability.

Liling Wan, Ph.D.

Molecular link between histone acetylation and oncogenic gene activation Histone acetylation is a chromatin mark generally associated with gene activation, yet the molecular mechanisms underlying this correlative relationship remain incompletely understood. I led a collaborative study in which we identified a novel ‘reader’ for histone acetylation named ENL. We showed in leukemia cells that ENL interacts with histone acetylation via the well-conserved YEATS domain, and in so doing, helps to recruit and stabilize its associated transcriptional machinery to drive transcription of leukemogenic genes. By determining the structure of ENL in complex with an acetylated histone peptide, we and our collaborators demonstrated that disrupting the reader function reduced chromatin recruitment of ENL-associated transcriptional machinery and resulted in suppression of oncogenic programs. Furthermore, blocking the functionality of ENL sensitized leukaemia cells to inhibitors that target another distinct class of histone acetylation readers, the BET proteins, thus highlighting the crosstalk between epigenetic readers and potential benefit of combinatorial therapies. Our work established ENL as a missing molecular link between histone acetylation and gene activation critical for leukemia malignant state, and has inspired following studies investigating other YEATS domain-containing proteins as a new class of chromatin ‘readers’ in a broad range of human cancers. In addition to bringing novel insights into our basic understanding of chromatin regulation, this work also provides mechanistic guidance and structural basis for ongoing drug development to target chromatin reading activity of ENL in aggressive leukemias.

Wan L#, Wen H#, Li Y#, Lyu J, Xi Y, Hoshii T, Joseph JK, Wang X, Loh YE, Erb MA, Souza AL, Bradner JE, Shen L, Li W, Li H*, Allis CD*, Armstrong SA*, Shi X*. ENL Links Histone Acetylation to Oncogenic Gene Activation in Leukemias. Nature 2017 Mar 9;543(7644):265-269. (#Equal contribution). PMC5372383
Li Y*, Sabari BR*, Panchenko T*, Wen H, Zhao D, Guan H, Wan L, Huang H, Tang Z, Zhao Y, Roeder RG, Shi X, Allis CD, Li H. Molecular Coupling of Histone Crotonylation and Active Transcription by AF9 YEATS Domain. Mol Cell2016 Apr 21;62(2):181-93. (*Equal contribution) PMC4841940

New type of gain-of-function mutations in chromatin readers Recognition of modified histones by ‘reader’ proteins constitutes a key mechanism mediating the function of histone modifications, yet the mechanisms by which their dysregulation contributes to diseases remain poorly understood. Recurrent, hotspot mutations in the acetylation-reading domain (YEATS domain) of ENL were recently found in Wilms’ tumor, the most common type of pediatric kidney cancer. Whether and how these mutations cause the disease remained unknown. Our current work shows that these mutations confer ENL gain of function in driving abnormal gene expression implicated in cancer. Unexpectedly, these mutations promoted ENL self-association, resulting in the formation of discrete nuclear puncta that are characteristic of biomolecular condensates, a newly recognized form of protein assembly that often involves weak, multivalent molecular interactions and commonly underlies the formation of membrane-less organelles. We demonstrated that such a property drives ‘self-reinforced’ chromatin targeting of mutant ENL protein and associated transcriptional machinery, thus enforcing active transcription from target loci. Aberrant gene control driven by ENL mutations, in turn, perturbs developmental programs and derails normal cell fate to a path towards tumorigenesis. This work is a remarkable demonstration of how mistakes in chromatin reader-mediated process can act as a driving force for tumor formation. These mutations represent a new class of oncogenic mutations which impair cell fate through promoting self-association and reinforcing chromatin targeting.

Wan L#, Chong S, Fan X, Liang A, Cui X, Gates L, Carroll TS, Li Y, Feng L, Chen G, Wang S, Ortiz MV, Daley S, Wang X, Xuan H, Kentsis A, Muir TW, Roeder RG, Li H, Li W, Tjian R, Wen H#, Allis CD#. Impaired Cell Fate through Gain-of-function Mutations in a Chromatin Reader. Nature 2019 in press (#co-corresponding)

Targeting chromatin readers as cancer therapies My postdoctoral work suggested that the displacement of histone acetylation reader ENL from chromatin may be a promising epigenetic therapy, alone or in combination with BET inhibitors, for aggressive leukemia. I have contributed to the development of peptidomimetic and small molecule inhibitors targeting the YEATS domain protein family. The ultimate goal of these and other ongoing efforts is to develop chemical probes targeting the ‘reading’ activity of ENL and other family members as valuable research tools and potential therapeutic agents.

Li X*, Li XM*, Jiang Y, Liu Z, Cui Y, Fung K, van der Beelen S, Tian G, Wan L, Shi X, Allis CD, Li H, Li Y#, Li X#. Structure-guided Development of YEATS Domain Inhibitors by Targeting π-π-π Stacking. Nat Chem Biol. 2018 Dec;14(12):1140-1149. (*Equal contribution) PMC6503841

Molecular mechanisms of cancer metastasis Metastasis accounts for > 90% cancer-related deaths and yet is the most poorly understood aspect of cancer biology. I have contributed to studies in which we identified and characterized new molecular mechanisms for cancer metastasis. My graduate work focused on Metadherin (MTDH), a novel cancer gene identified by our group to be prevalently amplified in breast cancer and strongly associated with a high risk of metastasis and poor prognosis. What drives the strong selection of MTDH in primary tumors was unclear. By generating genetically engineered mouse models, we provided first evidence supporting an essential role of MTDH in the initiation and metastasis of diverse subtypes of breast cancer. We further showed that MTDH regulates the expansion and activity of cancer stem cells through working with its binding partner SND1. By determining the atomic structure of the complex via collaboration, we demonstrated that disrupting the complex impairs breast cancer development and metastasis in vivo. Our work establishes MTDH and SND1 as critical regulators of cancer development and provides mechanistic guidance for ongoing drug development efforts to target this complex as cancer therapy. More broadly, this work provides crucial experimental support for the emerging concept that metastatic potential could be conferred by early oncogenic events that possess additional metastasis-promoting function and advances our understanding of the origin of metastatic traits.

Wan L, Lu X, Yuan S, Wei Y, Guo F, Shen M, Yuan M, Chakrabarti R, Hua Y, Smith HA, Blanco MA, Chekmareva M, Wu H, Bronson RT, Haffty BG, Xing Y, Kang Y. MTDH-SND1 Interaction Is Crucial for Expansion and Activity of Tumor-Initiating Cells in Diverse Oncogene- and Carcinogen-Induced Mammary Tumors. Cancer Cell 2014 Jul 14;26(1):92-105. PMC4101059
Wan L, Pantel K, Kang Y. Tumor Metastasis: Moving New Biological Insights into the Clinic. Nature Medicine 2013 Nov;19(11):1450-64. PMID: 24202397
Guo F#, Wan L#, Zheng A, Stanevich V, Wei Y, Satyshur KA, Shen M, Lee W, Kang Y, Xing Y. Structural Insights into the Tumor-Promoting Function of the MTDH-SND1 Complex. Cell Reports 2014 Sep 25;8(6):1704-13.  (#Equal contribution). PMC4309369
Wan L, Hu G, Wei Y, Yuan M, Bronson RT, Yang Q, Siddiqui J, Pienta KJ, Kang Y. Genetic Ablation of Metadherin Inhibits Autochthonous Prostate Cancer Progression and Metastasis. Cancer Research 2014 Sep 15;74(18):5336-47. PMC4167565
Kang Y, Xing Y, Wan L, Guo F. Use of peptides that block metadherin-SND1 interaction as treatment for cancer. (U.S. Patent No. 10,357,539 B2).

Research Interest

The research interests in our laboratory lie in the intersection of cancer biology and epigenetics. We focus on chromatin – the complex of DNA and histone proteins – and its regulatory network. Cancer genome studies revealed that at least 50% of human cancers harbor mutations in genes encoding chromatin-associated factors, suggesting widespread roles of chromatin misregulation in cancer. We strive to understand chromatin function and its dysregulation in human cancer, with a focus on addressing how chromatin-based mechanisms regulate cellular fate transition and plasticity that endow cancer cells with tumor-promoting potentials. We use a host of different approaches in genetics, epigenetics, biochemistry, genome-wide sequencing, bioinformatics and functional genomics to address these questions. We are also interested in leveraging our basic mechanistic discoveries for therapeutics development.

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