RESUMEN
Following three decades of systematic primary empirical screening against mice bearing two transplantable murine leukemias, the NCI took the bold step of switching to a radically different approach-initial screening of 10,000 diverse compounds/year against a panel of 60 human tumor cell lines in vitro. The establishment of the "NCI-60" screen was announced in the landmark Cancer Research article by Alley and colleagues, published in 1988, which exemplified the technological basis for the new microculture screen, operating at unprecedented scale. The underlying concept was that NCI-60 might expedite the discovery of innovative cancer drugs, especially those with predicted activity against particular solid cancers-not then possible. We discuss how NCI-60 provided a major technological advance and delivered a successful legacy for cancer research and development. While not immediately cracking the thorny problem of model-to-human tumor type prediction, NCI-60 nevertheless provided the conceptual and methodologic foundation for subsequent, much larger-scale human cancer cell panel screens with detailed molecular annotation and sophisticated informatics. Now used in modern molecular target-based drug discovery, these panels help enable the implementation of contemporary biomarker-led precision oncology. See related article by Alley and colleagues, Cancer Res 1988;48:589-601.
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Antineoplásicos , Neoplasias , Humanos , Animales , Ratones , Evaluación Preclínica de Medicamentos , Estudios de Factibilidad , Detección Precoz del Cáncer , Medicina de Precisión , Antineoplásicos/farmacología , Línea Celular Tumoral , Neoplasias/tratamiento farmacológico , Neoplasias/genéticaRESUMEN
Small-molecule chemical "probes" complement the use of molecular biology techniques to explore, validate, and generate hypotheses on the function of proteins in diseases such as cancer. Unfortunately, the poor selection and use of small-molecule reagents can lead to incorrect conclusions. Here, we illustrate examples of poor chemical tools and suggest best practices for the selection, validation, and use of high-quality chemical probes in cancer research. We also note the complexity associated with tools for novel drug modalities, exemplified by protein degraders, and provide advice and resources to facilitate the independent identification of appropriate small-molecule probes by researchers. SIGNIFICANCE: Validation of biological targets and pathways will be aided by a shared understanding of the criteria of potency, selectivity, and target engagement associated with small-molecule reagents ("chemical probes") that enable that work. Interdisciplinary collaboration between cancer biologists, medicinal chemists, and chemical biologists and the awareness of available resources will reduce misleading data generation and interpretation, strengthen data robustness, and improve productivity in academic and industrial research.
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Neoplasias , Investigación , Humanos , Proteínas , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/metabolismoRESUMEN
Within druggable target space, new small-molecule modalities, particularly covalent inhibitors and targeted degraders, have expanded the repertoire of medicinal chemists. Molecules with such modes of action have a large potential not only as drugs but also as chemical probes. Criteria have previously been established to describe the potency, selectivity, and properties of small-molecule probes that are qualified to enable the interrogation and validation of drug targets. These definitions have been tailored to reversibly acting modulators but fall short in their applicability to other modalities. While initial guidelines have been proposed, we delineate here a full set of criteria for the characterization of covalent, irreversible inhibitors as well as heterobifunctional degraders ("proteolysis-targeting chimeras", or PROTACs) and molecular glue degraders. We propose modified potency and selectivity criteria compared to those for reversible inhibitors. We discuss their relevance and highlight examples of suitable probe and pathfinder compounds.
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Ubiquitina-Proteína Ligasas , ProteolisisRESUMEN
CCT251236 1, a potent chemical probe, was previously developed from a cell-based phenotypic high-throughput screen (HTS) to discover inhibitors of transcription mediated by HSF1, a transcription factor that supports malignancy. Owing to its activity against models of refractory human ovarian cancer, 1 was progressed into lead optimization. The reduction of P-glycoprotein efflux became a focus of early compound optimization; central ring halogen substitution was demonstrated by matched molecular pair analysis to be an effective strategy to mitigate this liability. Further multiparameter optimization led to the design of the clinical candidate, CCT361814/NXP800 22, a potent and orally bioavailable fluorobisamide, which caused tumor regression in a human ovarian adenocarcinoma xenograft model with on-pathway biomarker modulation and a clean in vitro safety profile. Following its favorable dose prediction to human, 22 has now progressed to phase 1 clinical trial as a potential future treatment for refractory ovarian cancer and other malignancies.
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Antineoplásicos , Neoplasias Ováricas , Humanos , Femenino , Factores de Transcripción/metabolismo , Neoplasias Ováricas/patología , Línea Celular Tumoral , Antineoplásicos/farmacologíaRESUMEN
canSAR (https://cansar.ai) is the largest public cancer drug discovery and translational research knowledgebase. Now hosted in its new home at MD Anderson Cancer Center, canSAR integrates billions of experimental measurements from across molecular profiling, pharmacology, chemistry, structural and systems biology. Moreover, canSAR applies a unique suite of machine learning algorithms designed to inform drug discovery. Here, we describe the latest updates to the knowledgebase, including a focus on significant novel data. These include canSAR's ligandability assessment of AlphaFold; mapping of fragment-based screening data; and new chemical bioactivity data for novel targets. We also describe enhancements to the data and interface.
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Antineoplásicos , Descubrimiento de Drogas , Bases del Conocimiento , Investigación Biomédica Traslacional , Humanos , Algoritmos , Neoplasias/tratamiento farmacológico , Neoplasias/genéticaRESUMEN
The molecular chaperone heat shock protein 90 (HSP90) works in concert with co-chaperones to stabilize its client proteins, which include multiple drivers of oncogenesis and malignant progression. Pharmacologic inhibitors of HSP90 have been observed to exert a wide range of effects on the proteome, including depletion of client proteins, induction of heat shock proteins, dissociation of co-chaperones from HSP90, disruption of client protein signaling networks, and recruitment of the protein ubiquitylation and degradation machinery-suggesting widespread remodeling of cellular protein complexes. However, proteomics studies to date have focused on inhibitor-induced changes in total protein levels, often overlooking protein complex alterations. Here, we use size-exclusion chromatography in combination with mass spectrometry (SEC-MS) to characterize the early changes in native protein complexes following treatment with the HSP90 inhibitor tanespimycin (17-AAG) for 8 h in the HT29 colon adenocarcinoma cell line. After confirming the signature cellular response to HSP90 inhibition (e.g., induction of heat shock proteins, decreased total levels of client proteins), we were surprised to find only modest perturbations to the global distribution of protein elution profiles in inhibitor-treated HT29 cells at this relatively early time-point. Similarly, co-chaperones that co-eluted with HSP90 displayed no clear difference between control and treated conditions. However, two distinct analysis strategies identified multiple inhibitor-induced changes, including known and unknown components of the HSP90-dependent proteome. We validate two of these-the actin-binding protein Anillin and the mitochondrial isocitrate dehydrogenase 3 complex-as novel HSP90 inhibitor-modulated proteins. We present this dataset as a resource for the HSP90, proteostasis, and cancer communities (https://www.bioinformatics.babraham.ac.uk/shiny/HSP90/SEC-MS/), laying the groundwork for future mechanistic and therapeutic studies related to HSP90 pharmacology. Data are available via ProteomeXchange with identifier PXD033459.
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Adenocarcinoma , Antineoplásicos , Neoplasias del Colon , Humanos , Proteoma/metabolismo , Adenocarcinoma/tratamiento farmacológico , Neoplasias del Colon/tratamiento farmacológico , Proteínas HSP90 de Choque Térmico , Chaperonas Moleculares , Antineoplásicos/farmacología , Espectrometría de Masas , Cromatografía en GelRESUMEN
We describe the Chemical Probes Portal (https://www.chemicalprobes.org/), an expert review-based public resource to empower chemical probe assessment, selection and use. Chemical probes are high-quality small-molecule reagents, often inhibitors, that are important for exploring protein function and biological mechanisms, and for validating targets for drug discovery. The publication, dissemination and use of chemical probes provide an important means to accelerate the functional annotation of proteins, the study of proteins in cell biology, physiology, and disease pathology, and to inform and enable subsequent pioneering drug discovery and development efforts. However, the widespread use of small-molecule compounds that are claimed as chemical probes but are lacking sufficient quality, especially being inadequately selective for the desired target or even broadly promiscuous in behaviour, has resulted in many erroneous conclusions in the biomedical literature. The Chemical Probes Portal was established as a public resource to aid the selection and best-practice use of chemical probes in basic and translational biomedical research. We describe the background, principles and content of the Portal and its technical development, as well as examples of its applications and use. The Chemical Probes Portal is a community resource and we therefore describe how researchers can be involved in its content and development.
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Sondas Moleculares , Proteínas , Descubrimiento de Drogas , Proteínas/química , Proteínas/metabolismo , Bases de Datos de Compuestos QuímicosRESUMEN
Small-molecule chemical probes are among the most important tools to study the function of proteins in cells and organisms. Regrettably, the use of weak and non-selective small molecules has generated an abundance of erroneous conclusions in the scientific literature. More recently, minimal criteria have been outlined for investigational compounds, encouraging the selection and use of high-quality chemical probes. Here, we briefly recall the milestones and key initiatives that have paved the way to this new era, illustrate examples of recent high-quality chemical probes and provide our perspective on future challenges and developments.
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New discoveries and technologies are driving major advances in our understanding of cancer and underpinning a new era of precision medicine and immunotherapy. However, advances in treatment have been uneven: whereas survival rates for some common cancers are improving rapidly, for others there has been much less progress. In addition, healthcare systems are finding it difficult to provide access to expensive new treatments. There is an urgent need for imaginative policy solutions to incentivise the creation of novel therapies that address the full range of cancer-causing mechanisms. We have worked with organisations across the medical research community to develop consensus statements on how to enhance access to innovative cancer drugs. Here, we present our reflections on these statements.
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Antineoplásicos , Neoplasias , Antineoplásicos/uso terapéutico , Humanos , Factores Inmunológicos , Inmunoterapia , Neoplasias/tratamiento farmacológico , Medicina de PrecisiónRESUMEN
FOLFIRINOX, a combination of chemotherapy drugs (Fluorouracil, Oxaliplatin, Irinotecan -FOI), provides the best clinical benefit in pancreatic ductal adenocarcinoma (PDAC) patients. In this study we explore the role of miRNAs (MIR) as modulators of chemosensitivity to identify potential biomarkers of response. We find that 41 and 84 microRNA inhibitors enhance the sensitivity of Capan1 and MiaPaCa2 PDAC cells respectively. These include a MIR1307-inhibitor that we validate in further PDAC cell lines. Chemotherapy-induced apoptosis and DNA damage accumulation are higher in MIR1307 knock-out (MIR1307KO) versus control PDAC cells, while re-expression of MIR1307 in MIR1307KO cells rescues these effects. We identify binding of MIR1307 to CLIC5 mRNA through covalent ligation of endogenous Argonaute-bound RNAs cross-linking immunoprecipitation assay. We validate these findings in an in vivo model with MIR1307 disruption. In a pilot cohort of PDAC patients undergoing FOLFIRONX chemotherapy, circulating MIR1307 correlates with clinical outcome.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Carcinoma Ductal Pancreático/tratamiento farmacológico , Daño del ADN , Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Neoplasias Pancreáticas/tratamiento farmacológico , Carcinoma Ductal Pancreático/genética , Fluorouracilo/administración & dosificación , Humanos , Irinotecán/administración & dosificación , Estimación de Kaplan-Meier , Leucovorina/administración & dosificación , Terapia Neoadyuvante , Evaluación de Resultado en la Atención de Salud/métodos , Evaluación de Resultado en la Atención de Salud/estadística & datos numéricos , Oxaliplatino/administración & dosificación , Neoplasias Pancreáticas/genéticaRESUMEN
It has been recognized for decades that ERBB signaling is important in prostate cancer, but targeting ERBB receptors as a therapeutic strategy for prostate cancer has been ineffective clinically. However, we show here that membranous HER3 protein is commonly highly expressed in lethal prostate cancer, associating with reduced time to castration resistance (CR) and survival. Multiplex immunofluorescence indicated that the HER3 ligand NRG1 is detectable primarily in tumor-infiltrating myelomonocytic cells in human prostate cancer; this observation was confirmed using single-cell RNA sequencing of human prostate cancer biopsies and murine transgenic prostate cancer models. In castration-resistant prostate cancer (CRPC) patient-derived xenograft organoids with high HER3 expression as well as mouse prostate cancer organoids, recombinant NRG1 enhanced proliferation and survival. Supernatant from murine bone marrow-derived macrophages and myeloid-derived suppressor cells promoted murine prostate cancer organoid growth in vitro, which could be reversed by a neutralizing anti-NRG1 antibody and ERBB inhibition. Targeting HER3, especially with the HER3-directed antibody-drug conjugate U3-1402, exhibited antitumor activity against HER3-expressing prostate cancer. Overall, these data indicate that HER3 is commonly overexpressed in lethal prostate cancer and can be activated by NRG1 secreted by myelomonocytic cells in the tumor microenvironment, supporting HER3-targeted therapeutic strategies for treating HER3-expressing advanced CRPC. SIGNIFICANCE: HER3 is an actionable target in prostate cancer, especially with anti-HER3 immunoconjugates, and targeting HER3 warrants clinical evaluation in prospective trials.
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Anticuerpos Monoclonales Humanizados/farmacología , Biomarcadores de Tumor/metabolismo , Camptotecina/análogos & derivados , Neurregulina-1/metabolismo , Organoides/patología , Neoplasias de la Próstata/patología , Receptor ErbB-3/antagonistas & inhibidores , Animales , Antineoplásicos Inmunológicos/farmacología , Apoptosis , Biomarcadores de Tumor/genética , Camptotecina/farmacología , Proliferación Celular , Estudios de Seguimiento , Humanos , Masculino , Ratones Endogámicos NOD , Ratones SCID , Neurregulina-1/genética , Organoides/efectos de los fármacos , Organoides/metabolismo , Pronóstico , Estudios Prospectivos , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Receptor ErbB-3/genética , Receptor ErbB-3/metabolismo , Tasa de Supervivencia , Células Tumorales Cultivadas , Microambiente Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Most small molecules interact with several target proteins but this polypharmacology is seldom comprehensively investigated or explicitly exploited during drug discovery. Here, we use computational and experimental methods to identify and systematically characterize the kinase cross-pharmacology of representative HSP90 inhibitors. We demonstrate that the resorcinol clinical candidates ganetespib and, to a lesser extent, luminespib, display unique off-target kinase pharmacology as compared with other HSP90 inhibitors. We also demonstrate that polypharmacology evolved during the optimization to discover luminespib and that the hit, leads, and clinical candidate all have different polypharmacological profiles. We therefore recommend the computational and experimental characterization of polypharmacology earlier in drug discovery projects to unlock new multi-target drug design opportunities.
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Descubrimiento de Drogas , Evolución Molecular , Proteínas HSP90 de Choque Térmico/metabolismo , Inhibidores de Proteínas Quinasas/metabolismo , Sitios de Unión , Receptor con Dominio Discoidina 1/antagonistas & inhibidores , Receptor con Dominio Discoidina 1/metabolismo , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Humanos , Isoxazoles/química , Isoxazoles/metabolismo , Simulación del Acoplamiento Molecular , Inhibidores de Proteínas Quinasas/química , Proteínas Proto-Oncogénicas c-abl/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-abl/metabolismo , Resorcinoles/química , Resorcinoles/metabolismo , Triazoles/química , Triazoles/metabolismoRESUMEN
Casein kinase 2 (CK2) is highly expressed in cancer and has been considered a potential therapeutic target. Wells and colleagues developed and characterized the new CK2 inhibitor SGC-CK2-1. Unexpectedly, this potent and highly selective chemical probe does not show broad antiproliferative activity in cancer cells.
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Quinasa de la Caseína II , Neoplasias , Humanos , Neoplasias/tratamiento farmacológico , Inhibidores de Proteínas QuinasasRESUMEN
Anticancer drug development is a costly and protracted activity, and failure at late phases of clinical testing is common. We have previously proposed the Pharmacological Audit Trail (PhAT) intended to improve the efficiency of drug development, with a focus on the use of tumour tissue-based biomarkers. Blood-based 'liquid biopsy' approaches, such as targeted or whole-genome sequencing studies of plasma circulating cell-free tumour DNA (ctDNA) and circulating tumour cells (CTCs), are of increasing relevance to this drug development paradigm. Liquid biopsy assays can provide quantitative and qualitative data on prognostic, predictive, pharmacodynamic and clinical response biomarkers, and can also enable the characterization of disease evolution and resistance mechanisms. In this Perspective, we examine the promise of integrating liquid biopsy analyses into the PhAT, focusing on the current evidence, advances, limitations and challenges. We emphasize the continued importance of analytical validation and clinical qualification of circulating tumour biomarkers through prospective clinical trials.
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Antineoplásicos/uso terapéutico , Desarrollo de Medicamentos , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Algoritmos , Antineoplásicos/aislamiento & purificación , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , ADN Tumoral Circulante/sangre , Auditoría Clínica/métodos , Auditoría Clínica/organización & administración , Desarrollo de Medicamentos/métodos , Desarrollo de Medicamentos/organización & administración , Humanos , Biopsia Líquida , Neoplasias/diagnóstico , Células Neoplásicas Circulantes/patología , PronósticoRESUMEN
Chordomas are rare spinal tumors addicted to expression of the developmental transcription factor brachyury. In chordomas, brachyury is super-enhancer associated and preferentially downregulated by pharmacologic transcriptional CDK inhibition, leading to cell death. To understand the underlying basis of this sensitivity, we dissect the brachyury transcription regulatory network and compare the consequences of brachyury degradation with transcriptional CDK inhibition. Brachyury defines the chordoma super-enhancer landscape and autoregulates through binding its super-enhancer, and its locus forms a transcriptional condensate. Transcriptional CDK inhibition and brachyury degradation disrupt brachyury autoregulation, leading to loss of its transcriptional condensate and transcriptional program. Compared with transcriptional CDK inhibition, which globally downregulates transcription, leading to cell death, brachyury degradation is much more selective, inducing senescence and sensitizing cells to anti-apoptotic inhibition. These data suggest that brachyury downregulation is a core tenet of transcriptional CDK inhibition and motivates developing strategies to target brachyury and its autoregulatory feedback loop.
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Biomarcadores de Tumor/genética , Cordoma/genética , Quinasas Ciclina-Dependientes/genética , Proteínas Fetales/genética , Proteínas de Neoplasias/genética , Neoplasias de la Columna Vertebral/genética , Proteínas de Dominio T Box/genética , Secuencia de Bases , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Supervivencia Celular , Cordoma/metabolismo , Cordoma/patología , Quinasas Ciclina-Dependientes/metabolismo , Proteínas Fetales/metabolismo , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes , Células HEK293 , Histonas/genética , Histonas/metabolismo , Humanos , Queratina-18/genética , Queratina-18/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas de Neoplasias/metabolismo , Proteolisis , Transducción de Señal , Neoplasias de la Columna Vertebral/metabolismo , Neoplasias de la Columna Vertebral/patología , Proteínas de Dominio T Box/metabolismoRESUMEN
High-quality small molecule chemical probes are extremely valuable for biological research and target validation. However, frequent use of flawed small-molecule inhibitors produces misleading results and diminishes the robustness of biomedical research. Several public resources are available to facilitate assessment and selection of better chemical probes for specific protein targets. Here, we review chemical probe resources, discuss their current strengths and limitations, and make recommendations for further improvements. Expert review resources provide in-depth analysis but currently cover only a limited portion of the liganded proteome. Computational resources encompass more proteins and are regularly updated, but have limitations in data availability and curation. We show how biomedical scientists may use these resources to choose the best available chemical probes for their research.
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Inhibidores Enzimáticos/química , Sondas Moleculares/química , Proteínas/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Algoritmos , Animales , Simulación por Computador , Bases de Datos de Compuestos Químicos , Inhibidores Enzimáticos/farmacología , Humanos , Sondas Moleculares/farmacología , Proteoma/química , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-ActividadRESUMEN
canSAR (http://cansar.icr.ac.uk) is the largest, public, freely available, integrative translational research and drug discovery knowledgebase for oncology. canSAR integrates vast multidisciplinary data from across genomic, protein, pharmacological, drug and chemical data with structural biology, protein networks and more. It also provides unique data, curation and annotation and crucially, AI-informed target assessment for drug discovery. canSAR is widely used internationally by academia and industry. Here we describe significant developments and enhancements to the data, web interface and infrastructure of canSAR in the form of the new implementation of the system: canSARblack. We demonstrate new functionality in aiding translation hypothesis generation and experimental design, and show how canSAR can be adapted and utilised outside oncology.
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Biología Computacional/métodos , Bases de Datos Genéticas , Descubrimiento de Drogas/métodos , Bases del Conocimiento , Neoplasias/genética , Investigación Biomédica Traslacional/métodos , Antineoplásicos/química , Antineoplásicos/uso terapéutico , Minería de Datos/métodos , Genómica/métodos , Humanos , Internet , Oncología Médica/métodos , Estructura Molecular , Neoplasias/metabolismo , Proteómica/métodos , Interfaz Usuario-ComputadorRESUMEN
The undruggable nature of oncogenic Myc transcription factors poses a therapeutic challenge in neuroblastoma, a pediatric cancer in which MYCN amplification is strongly associated with unfavorable outcome. Here, we show that CYC065 (fadraciclib), a clinical inhibitor of CDK9 and CDK2, selectively targeted MYCN-amplified neuroblastoma via multiple mechanisms. CDK9 - a component of the transcription elongation complex P-TEFb - bound to the MYCN-amplicon superenhancer, and its inhibition resulted in selective loss of nascent MYCN transcription. MYCN loss led to growth arrest, sensitizing cells for apoptosis following CDK2 inhibition. In MYCN-amplified neuroblastoma, MYCN invaded active enhancers, driving a transcriptionally encoded adrenergic gene expression program that was selectively reversed by CYC065. MYCN overexpression in mesenchymal neuroblastoma was sufficient to induce adrenergic identity and sensitize cells to CYC065. CYC065, used together with temozolomide, a reference therapy for relapsed neuroblastoma, caused long-term suppression of neuroblastoma growth in vivo, highlighting the clinical potential of CDK9/2 inhibition in the treatment of MYCN-amplified neuroblastoma.
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Adenosina/análogos & derivados , Quinasa 2 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 9 Dependiente de la Ciclina/antagonistas & inhibidores , Proteína Proto-Oncogénica N-Myc/biosíntesis , Neuroblastoma/tratamiento farmacológico , Temozolomida/farmacología , Adenosina/farmacología , Línea Celular Tumoral , Quinasa 2 Dependiente de la Ciclina/metabolismo , Quinasa 9 Dependiente de la Ciclina/metabolismo , Elementos de Facilitación Genéticos , Humanos , Proteína Proto-Oncogénica N-Myc/genética , Neuroblastoma/genética , Neuroblastoma/metabolismo , Neuroblastoma/patología , Factor B de Elongación Transcripcional Positiva/genética , Factor B de Elongación Transcripcional Positiva/metabolismo , Transcripción Genética/efectos de los fármacosRESUMEN
The original version of the book was revised: Chapter 11 is now available open access under a CC BY 4.0 license and the copyright holder has been changed to 'The Author(s)'.
RESUMEN
Heat shock protein 90 (Hsp90) is a molecular chaperone that plays an important role in tumour biology by promoting the stabilisation and activity of oncogenic 'client' proteins. Inhibition of Hsp90 by small-molecule drugs, acting via its ATP hydrolysis site, has shown promise as a molecularly targeted cancer therapy. Owing to the importance of Hop and other tetratricopeptide repeat (TPR)-containing cochaperones in regulating Hsp90 activity, the Hsp90-TPR domain interface is an alternative site for inhibitors, which could result in effects distinct from ATP site binders. The TPR binding site of Hsp90 cochaperones includes a shallow, positively charged groove that poses a significant challenge for druggability. Herein, we report the apo, solution-state structure of Hop TPR2A which enables this target for NMR-based screening approaches. We have designed prototype TPR ligands that mimic key native 'carboxylate clamp' interactions between Hsp90 and its TPR cochaperones and show that they block binding between Hop TPR2A and the Hsp90 C-terminal MEEVD peptide. We confirm direct TPR-binding of these ligands by mapping 1H-15N HSQC chemical shift perturbations to our new NMR structure. Our work provides a novel structure, a thorough assessment of druggability and robust screening approaches that may offer a potential route, albeit difficult, to address the chemically challenging nature of the Hop TPR2A target, with relevance to other TPR domain interactors.