ABSTRACT
In the folded state, biomolecules exchange between multiple conformational states crucial for their function. However, most structural models derived from experiments and computational predictions only encode a single state. To represent biomolecules accurately, we must move towards modeling and predicting structural ensembles. Information about structural ensembles exists within experimental data from X-ray crystallography and cryo-electron microscopy. Although new tools are available to detect conformational and compositional heterogeneity within these ensembles, the legacy PDB data structure does not robustly encapsulate this complexity. We propose modifications to the macromolecular crystallographic information file (mmCIF) to improve the representation and interrelation of conformational and compositional heterogeneity. These modifications will enable the capture of macromolecular ensembles in a human and machine-interpretable way, potentially catalyzing breakthroughs for ensemble-function predictions, analogous to the achievements of AlphaFold with single-structure prediction.
Subject(s)
Cryoelectron Microscopy , Databases, Protein , Models, Molecular , Protein Conformation , Proteins , Crystallography, X-Ray , Proteins/chemistry , Cryoelectron Microscopy/methods , HumansABSTRACT
In their folded state, biomolecules exchange between multiple conformational states that are crucial for their function. Traditional structural biology methods, such as X-ray crystallography and cryogenic electron microscopy (cryo-EM), produce density maps that are ensemble averages, reflecting molecules in various conformations. Yet, most models derived from these maps explicitly represent only a single conformation, overlooking the complexity of biomolecular structures. To accurately reflect the diversity of biomolecular forms, there is a pressing need to shift toward modeling structural ensembles that mirror the experimental data. However, the challenge of distinguishing signal from noise complicates manual efforts to create these models. In response, we introduce the latest enhancements to qFit, an automated computational strategy designed to incorporate protein conformational heterogeneity into models built into density maps. These algorithmic improvements in qFit are substantiated by superior Rfree and geometry metrics across a wide range of proteins. Importantly, unlike more complex multicopy ensemble models, the multiconformer models produced by qFit can be manually modified in most major model building software (e.g., Coot) and fit can be further improved by refinement using standard pipelines (e.g., Phenix, Refmac, Buster). By reducing the barrier of creating multiconformer models, qFit can foster the development of new hypotheses about the relationship between macromolecular conformational dynamics and function.
Subject(s)
Cryoelectron Microscopy , Models, Molecular , Protein Conformation , Cryoelectron Microscopy/methods , Crystallography, X-Ray/methods , Proteins/chemistry , Software , Algorithms , Computational Biology/methodsABSTRACT
Despite advances and social progress, the exclusion of diverse groups in academia, especially science, technology, engineering, and mathematics (STEM) fields, across the US and Europe persists, resulting in the underrepresentation of diverse people in higher education. There is extensive literature about theory, observation, and evidence-based practices that can help create a more equitable, inclusive, and diverse learning environment. In this article, we propose the implementation of a Diversity, Equity, Inclusion, and Justice (DEIJ) journal club as a strategic initiative to foster education and promote action towards making academia a more equitable institution. By creating a space for people to engage with DEIJ theories* and strategize ways to improve their learning environment, we hope to normalize the practice and importance of analyzing academia through an equity lens. Guided by restorative justice principles, we offer 10 recommendations for fostering community cohesion through education and mutual understanding. This approach underscores the importance of appropriate action and self-education in the journey toward a more diverse, equitable, inclusive, and just academic environment. *Authors' note: We understand that "DEIJ" is a multidisciplinary organizational framework that relies on numerous fields of study, including history, sociology, philosophy, and more. We use this term to refer to these different fields of study for brevity purposes.
Subject(s)
Cultural Diversity , Social Justice , Humans , Periodicals as Topic , Engineering/education , Science/education , Mathematics/education , Universities , Diversity, Equity, InclusionABSTRACT
In their folded state, biomolecules exchange between multiple conformational states that are crucial for their function. Traditional structural biology methods, such as X-ray crystallography and cryogenic electron microscopy (cryo-EM), produce density maps that are ensemble averages, reflecting molecules in various conformations. Yet, most models derived from these maps explicitly represent only a single conformation, overlooking the complexity of biomolecular structures. To accurately reflect the diversity of biomolecular forms, there is a pressing need to shift towards modeling structural ensembles that mirror the experimental data. However, the challenge of distinguishing signal from noise complicates manual efforts to create these models. In response, we introduce the latest enhancements to qFit, an automated computational strategy designed to incorporate protein conformational heterogeneity into models built into density maps. These algorithmic improvements in qFit are substantiated by superior Rfree and geometry metrics across a wide range of proteins. Importantly, unlike more complex multicopy ensemble models, the multiconformer models produced by qFit can be manually modified in most major model building software (e.g. Coot) and fit can be further improved by refinement using standard pipelines (e.g. Phenix, Refmac, Buster). By reducing the barrier of creating multiconformer models, qFit can foster the development of new hypotheses about the relationship between macromolecular conformational dynamics and function.
ABSTRACT
Conformational ensembles underlie all protein functions. Thus, acquiring atomic-level ensemble models that accurately represent conformational heterogeneity is vital to deepen our understanding of how proteins work. Modeling ensemble information from X-ray diffraction data has been challenging, as traditional cryo-crystallography restricts conformational variability while minimizing radiation damage. Recent advances have enabled the collection of high quality diffraction data at ambient temperatures, revealing innate conformational heterogeneity and temperature-driven changes. Here, we used diffraction datasets for Proteinase K collected at temperatures ranging from 313 to 363 K to provide a tutorial for the refinement of multiconformer ensemble models. Integrating automated sampling and refinement tools with manual adjustments, we obtained multiconformer models that describe alternative backbone and sidechain conformations, their relative occupancies, and interconnections between conformers. Our models revealed extensive and diverse conformational changes across temperature, including increased bound peptide ligand occupancies, different Ca2+ binding site configurations and altered rotameric distributions. These insights emphasize the value and need for multiconformer model refinement to extract ensemble information from diffraction data and to understand ensemble-function relationships.
Subject(s)
X-Ray Diffraction , Temperature , Crystallography , Binding Sites , Protein DomainsABSTRACT
Conformational ensembles underlie all protein functions. Thus, acquiring atomic-level ensemble models that accurately represent conformational heterogeneity is vital to deepen our understanding of how proteins work. Modeling ensemble information from X-ray diffraction data has been challenging, as traditional cryo-crystallography restricts conformational variability while minimizing radiation damage. Recent advances have enabled the collection of high quality diffraction data at ambient temperatures, revealing innate conformational heterogeneity and temperature-driven changes. Here, we used diffraction datasets for Proteinase K collected at temperatures ranging from 313 to 363K to provide a tutorial for the refinement of multiconformer ensemble models. Integrating automated sampling and refinement tools with manual adjustments, we obtained multiconformer models that describe alternative backbone and sidechain conformations, their relative occupancies, and interconnections between conformers. Our models revealed extensive and diverse conformational changes across temperature, including increased bound peptide ligand occupancies, different Ca2+ binding site configurations and altered rotameric distributions. These insights emphasize the value and need for multiconformer model refinement to extract ensemble information from diffraction data and to understand ensemble-function relationships.
ABSTRACT
While protein conformational heterogeneity plays an important role in many aspects of biological function, including ligand binding, its impact has been difficult to quantify. Macromolecular X-ray diffraction is commonly interpreted with a static structure, but it can provide information on both the anharmonic and harmonic contributions to conformational heterogeneity. Here, through multiconformer modeling of time- and space-averaged electron density, we measure conformational heterogeneity of 743 stringently matched pairs of crystallographic datasets that reflect unbound/apo and ligand-bound/holo states. When comparing the conformational heterogeneity of side chains, we observe that when binding site residues become more rigid upon ligand binding, distant residues tend to become more flexible, especially in non-solvent-exposed regions. Among ligand properties, we observe increased protein flexibility as the number of hydrogen bonds decreases and relative hydrophobicity increases. Across a series of 13 inhibitor-bound structures of CDK2, we find that conformational heterogeneity is correlated with inhibitor features and identify how conformational changes propagate differences in conformational heterogeneity away from the binding site. Collectively, our findings agree with models emerging from nuclear magnetic resonance studies suggesting that residual side-chain entropy can modulate affinity and point to the need to integrate both static conformational changes and conformational heterogeneity in models of ligand binding.
Proteins are the workhorses of our cells. They are large molecules that 'fold' into specific, often highly complex, three-dimensional configurations. These structures are not static, but rather dynamic and flexible. In other words, proteins can shift between different three-dimensional shapes to perform their tasks within the cell. To perform their roles, many proteins have to bind to small molecule ligands. Many ligands are drugs, which means that their effectiveness depends on their ability to bind to and impact the proteins involved in the disease they are treating. When a ligand binds to a protein, it can reshape the protein. For example, certain conformations of the protein, which were difficult for the protein to be in on its own, may become more stable when the ligand binds. Additionally, upon ligand binding, some parts of the protein may move relative to each other. Previous studies have shown that these movements can affect the interaction between ligand and protein. However, these studies only examined a small number of proteins. Therefore, Wankowicz et al. set out to determine, in greater detail, what happens to protein flexibility upon ligand binding. First, a pipeline was created to model alternative configurations of the protein both with and without ligands attached. These models measured flexibility within protein structures. The models revealed that when ligands bind to proteins, the flexibility of different regions of the protein changes and does so in a consistent way. Proteins that become more rigid in the region interacting with their ligands become less rigid in other, distant regions, and vice versa. In other words, the rest of the protein is able to compensate for any changes in flexibility caused by ligand binding, which may contribute to how well a ligand binds to a protein. This study demonstrates the ability of ligands to affect the entire structure of the proteins they bind to, and therefore sheds new light on the role of proteins' innate conformational flexibility during this process. These results will contribute to our understanding of how the ligands and proteins involved in different cellular processes interact with each other and, potentially, how these interactions can be manipulated.
Subject(s)
Proteins , Binding Sites , Ligands , Protein Binding , Protein Conformation , Proteins/metabolismABSTRACT
High-risk localized prostate cancer (HRLPC) is associated with a substantial risk of recurrence and disease mortality. Recent clinical trials have shown that intensifying anti-androgen therapies administered before prostatectomy can induce pathologic complete responses or minimal residual disease, called exceptional response, although the molecular determinants of these clinical outcomes are largely unknown. Here, we perform whole-exome and transcriptome sequencing on pre-treatment multi-regional tumor biopsies from exceptional responders (ERs) and non-responders (NRs, pathologic T3 or lymph node-positive disease) to intensive neoadjuvant anti-androgen therapies. Clonal SPOP mutation and SPOPL copy-number loss are exclusively observed in ERs, while clonal TP53 mutation and PTEN copy-number loss are exclusively observed in NRs. Transcriptional programs involving androgen signaling and TGF-ß signaling are enriched in ERs and NRs, respectively. These findings may guide prospective validation studies of these molecular features in large HRLPC clinical cohorts treated with neoadjuvant anti-androgens to improve patient stratification.
Subject(s)
Androgen Antagonists/therapeutic use , Nuclear Proteins/drug effects , Prostate-Specific Antigen/drug effects , Prostatic Neoplasms/drug therapy , Repressor Proteins/drug effects , Adaptor Proteins, Vesicular Transport , Antineoplastic Agents, Hormonal/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Humans , Male , Neoadjuvant Therapy/methods , Prostatectomy/methods , Prostatic Neoplasms/pathology , RiskABSTRACT
This paper describes outcomes of the 2019 Cryo-EM Model Challenge. The goals were to (1) assess the quality of models that can be produced from cryogenic electron microscopy (cryo-EM) maps using current modeling software, (2) evaluate reproducibility of modeling results from different software developers and users and (3) compare performance of current metrics used for model evaluation, particularly Fit-to-Map metrics, with focus on near-atomic resolution. Our findings demonstrate the relatively high accuracy and reproducibility of cryo-EM models derived by 13 participating teams from four benchmark maps, including three forming a resolution series (1.8 to 3.1 Å). The results permit specific recommendations to be made about validating near-atomic cryo-EM structures both in the context of individual experiments and structure data archives such as the Protein Data Bank. We recommend the adoption of multiple scoring parameters to provide full and objective annotation and assessment of the model, reflective of the observed cryo-EM map density.
Subject(s)
Cryoelectron Microscopy/methods , Models, Molecular , Crystallography, X-Ray , Protein Conformation , Proteins/chemistryABSTRACT
New X-ray crystallography and cryo-electron microscopy (cryo-EM) approaches yield vast amounts of structural data from dynamic proteins and their complexes. Modeling the full conformational ensemble can provide important biological insights, but identifying and modeling an internally consistent set of alternate conformations remains a formidable challenge. qFit efficiently automates this process by generating a parsimonious multiconformer model. We refactored qFit from a distributed application into software that runs efficiently on a small server, desktop, or laptop. We describe the new qFit 3 software and provide some examples. qFit 3 is open-source under the MIT license, and is available at https://github.com/ExcitedStates/qfit-3.0.
Subject(s)
Algorithms , Models, Molecular , Proteins/chemistry , Software , Cryoelectron Microscopy , Crystallography, X-Ray , LigandsABSTRACT
High-grade T1 (HGT1) bladder cancer is the highest risk subtype of non-muscle-invasive bladder cancer with unpredictable outcome and poorly understood risk factors. Here, we examined the association of somatic mutation profiles with nonrecurrent disease (GO, good outcome), recurrence (R), or progression (PD) in a cohort of HGT1 patients. Exome sequencing was performed on 62 HGT1 and 15 matched normal tissue samples. Both tumor only (TO) and paired analyses were performed, focusing on 95 genes known to be mutated in bladder cancer. Somatic mutations, copy-number alterations, mutation load, and mutation signatures were studied. Thirty-three GO, 10 R, 18 PD, and 1 unknown outcome patients were analyzed. Tumor mutational burden (TMB) was similar to muscle-invasive disease and was highest in GO, intermediate in PD, and lowest in R patients (P = 0.017). DNA damage response gene mutations were associated with higher TMB (P < 0.0001) and GO (P = 0.003). ERCC2 and BRCA2 mutations were associated with GO. TP53, ATM, ARID1A, AHR, and SMARCB1 mutations were more frequent in PD. Focal copy-number gain in CCNE1 and CDKN2A deletion was enriched in PD or R (P = 0.047; P = 0.06). APOBEC (46%) and COSMIC5 (34%) signatures were most frequent. APOBEC-A and ERCC2 mutant tumors (COSMIC5) were associated with GO (P = 0.047; P = 0.0002). pT1b microstaging was associated with a genomic cluster (P = 0.05) with focal amplifications of E2F3/SOX4, PVRL4, CCNE1, and TP53 mutations. Findings were validated using external public datasets. These findings require confirmation but suggest that management of HGT1 bladder cancer may be improved via molecular characterization to predict outcome. SIGNIFICANCE: Detailed genetic analyses of HGT1 bladder tumors identify features that correlate with outcome, e.g., high mutational burden, ERCC2 mutations, and high APOBEC-A/ERCC2 mutation signatures were associated with good outcome.
Subject(s)
Mutation , Urinary Bladder Neoplasms/genetics , Urinary Bladder Neoplasms/pathology , Cyclin E/genetics , Cyclin-Dependent Kinase Inhibitor p16/genetics , Female , Follow-Up Studies , Gene Dosage , Humans , Male , Muscles/pathology , Neoplasm Recurrence, Local/pathology , Oncogene Proteins/genetics , Tumor Suppressor Protein p53/genetics , Urinary Bladder Neoplasms/mortality , Xeroderma Pigmentosum Group D Protein/geneticsABSTRACT
Protein conformations are shaped by cellular environments, but how environmental changes alter the conformational landscapes of specific proteins in vivo remains largely uncharacterized, in part due to the challenge of probing protein structures in living cells. Here, we use deep mutational scanning to investigate how a toxic conformation of α-synuclein, a dynamic protein linked to Parkinson's disease, responds to perturbations of cellular proteostasis. In the context of a course for graduate students in the UCSF Integrative Program in Quantitative Biology, we screened a comprehensive library of α-synuclein missense mutants in yeast cells treated with a variety of small molecules that perturb cellular processes linked to α-synuclein biology and pathobiology. We found that the conformation of α-synuclein previously shown to drive yeast toxicity-an extended, membrane-bound helix-is largely unaffected by these chemical perturbations, underscoring the importance of this conformational state as a driver of cellular toxicity. On the other hand, the chemical perturbations have a significant effect on the ability of mutations to suppress α-synuclein toxicity. Moreover, we find that sequence determinants of α-synuclein toxicity are well described by a simple structural model of the membrane-bound helix. This model predicts that α-synuclein penetrates the membrane to constant depth across its length but that membrane affinity decreases toward the C terminus, which is consistent with orthogonal biophysical measurements. Finally, we discuss how parallelized chemical genetics experiments can provide a robust framework for inquiry-based graduate coursework.
Subject(s)
Saccharomyces cerevisiae/drug effects , alpha-Synuclein/toxicity , Amino Acid Sequence , Humans , Mutation , Parkinson Disease/metabolism , Protein Conformation , Saccharomyces cerevisiae/metabolism , alpha-Synuclein/chemistry , alpha-Synuclein/geneticsABSTRACT
PURPOSE: Molecular properties associated with complete response or acquired resistance to concurrent chemotherapy and radiotherapy (CRT) are incompletely characterized.Experimental Design: We performed integrated whole-exome/transcriptome sequencing and immune infiltrate analysis on rectal adenocarcinoma tumors prior to neoadjuvant CRT (pre-CRT) and at time of resection (post-CRT) in 17 patients [8 complete/partial responders, 9 nonresponders (NR)]. RESULTS: CRT was not associated with increased tumor mutational burden or neoantigen load and did not alter the distribution of established somatic tumor mutations in rectal cancer. Concurrent KRAS/TP53 mutations (KP) associated with NR tumors and were enriched for an epithelial-mesenchymal transition transcriptional program. Furthermore, NR was associated with reduced CD4/CD8 T-cell infiltrates and a post-CRT M2 macrophage phenotype. Absence of any local tumor recurrences, KP/NR status predicted worse progression-free survival, suggesting that local immune escape during or after CRT with specific genomic features contributes to distant progression. CONCLUSIONS: Overall, while CRT did not impact genomic profiles, CRT impacted the tumor immune microenvironment, particularly in resistant cases.
Subject(s)
Biomarkers, Tumor , Drug Resistance, Neoplasm , Radiation Tolerance , Rectal Neoplasms/etiology , Antigens, Neoplasm/immunology , Chemoradiotherapy, Adjuvant , Drug Resistance, Neoplasm/genetics , Gene Expression Profiling , Humans , Immunohistochemistry , Mutation , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Radiation Tolerance/genetics , Rectal Neoplasms/metabolism , Rectal Neoplasms/therapy , Transcriptome , Tumor Microenvironment/genetics , Tumor Microenvironment/immunology , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism , Exome SequencingABSTRACT
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
ABSTRACT
BACKGROUND: Metastatic urothelial carcinoma (mUC) is a genomically diverse disease with known alterations in the mTOR pathway and tyrosine kinases including FGFR. We investigated the efficacy and safety of combination treatment with everolimus and pazopanib (E/P) in genomically profiled patients with mUC. METHODS: mUC patients enrolled on a Phase I dose escalation study and an expansion cohort treated with E/P were included. The primary end point was objective response rate (ORR); secondary end points were safety, duration of response (DOR), progression-free survival (PFS) and overall survival (OS). Patients were assessed for mutations and copy number alterations in 300 relevant cancer-associated genes using next-generation sequencing and findings were correlated with outcomes. Time-to-event data were estimated with Kaplan-Meier methods. RESULTS: Of the 23 patients enrolled overall, 19 had mUC. ORR was 21% (one complete response (CR), three partial responses (PR), eight with stable disease (SD). DOR, PFS and OS were 6.5, 3.6, and 9.1 months, respectively. Four patients with clinical benefit (one CR, two PR, one SD) had mutations in TSC1/TSC2 or mTOR and a 5th patient with PR had a FGFR3-TACC3 fusion. CONCLUSIONS: Combination therapy with E/P is safe in mUC and select patients with alterations in mTOR or FGFR pathways derive significant clinical benefit.
Subject(s)
Carcinoma, Transitional Cell/drug therapy , Everolimus/administration & dosage , High-Throughput Nucleotide Sequencing/methods , Kidney Neoplasms/drug therapy , Pyrimidines/administration & dosage , Sulfonamides/administration & dosage , Carcinoma, Transitional Cell/genetics , DNA Copy Number Variations , Everolimus/therapeutic use , Female , Humans , Indazoles , Kidney Neoplasms/genetics , Male , Microtubule-Associated Proteins/genetics , Mutation , Pyrimidines/therapeutic use , Receptor, Fibroblast Growth Factor, Type 3/genetics , Sequence Analysis, DNA , Sulfonamides/therapeutic use , Survival Analysis , Treatment Outcome , Tuberous Sclerosis Complex 1 Protein/genetics , Tuberous Sclerosis Complex 2 Protein/geneticsABSTRACT
OBJECTIVES: To examine the impact of race on quality of care and overall survival (OS) among patients with muscle invasive bladder cancer (MIBC) treated with radical cystectomy (RC) in the U.S. MATERIALS & METHODS: Our cohort consisted of 12,652 patients receiving RC for MIBC within the National Cancer Database from 2004 to 2012. Patients were stratified by race (Black non-Hispanic vs. White non-Hispanic) and imbalances in patient characteristics mitigated using propensity score weighting. Logistic and Cox regressions examined the impact of race on quality of care metrics (receipt of pelvic lymph node dissection (PLND), lymph node count, hospital volume, length of stay, delay of treatment) and on OS. The difference in OS was expressed as Delta, and stratified by facility-type, hospital volume, and region. RESULTS: Blacks were less likely to receive PLND (odds ratio [OR] 0.70, 95% confidence interval [CI]: 0.55-0.91), or to have a greater number of lymph nodes removed (OR 0.76, 95%CI: 0.64-0.90). They exhibited greater length of stay (OR 1.34, 95%CI: 1.13-1.59), and delay of RC among recipients of neoadjuvant chemotherapy (OR 2.59, 95%CI: 1.77-3.85) (all P ≤ 0.001). Notably, utilization of neoadjuvant chemotherapy in advanced disease stages was more common in blacks (OR 2.82, 95%CI: 1.93-4.13, P < 0.001). Additionally, Black race was associated with inferior OS (Hazard ratio 0.87, 95%CI: 0.79-0.97, P < 0.014). Disparities in OS varied based on facility type and geographical region, but not hospital volume. Specifically, Blacks had worse OS when treated in a community cancer program (Delta 0.42, 95%CI: 0.28-0.57,P < 0.001), or within New England/Middle Atlantic region (Delta 0.16, 95% CI: 0.07-0.24,P < 0.001). CONCLUSION: Black race is an independent predictor of inferior quality of care and OS in patients undergoing RC for MIBC. Survival disparities vary based on geographical region and facility type. Notably, the OS disparity appears to have narrowed in comparison to previous studies.
Subject(s)
Carcinoma, Transitional Cell/mortality , Cystectomy/mortality , Healthcare Disparities/ethnology , Urinary Bladder Neoplasms/mortality , Black or African American , Aged , Carcinoma, Transitional Cell/surgery , Databases, Factual , Female , Humans , Male , Middle Aged , Urinary Bladder Neoplasms/surgery , White PeopleABSTRACT
Nearly all prostate cancer deaths are from metastatic castration-resistant prostate cancer (mCRPC), but there have been few whole-genome sequencing (WGS) studies of this disease state. We performed linked-read WGS on 23 mCRPC biopsy specimens and analyzed cell-free DNA sequencing data from 86 patients with mCRPC. In addition to frequent rearrangements affecting known prostate cancer genes, we observed complex rearrangements of the AR locus in most cases. Unexpectedly, these rearrangements include highly recurrent tandem duplications involving an upstream enhancer of AR in 70%-87% of cases compared with <2% of primary prostate cancers. A subset of cases displayed AR or MYC enhancer duplication in the context of a genome-wide tandem duplicator phenotype associated with CDK12 inactivation. Our findings highlight the complex genomic structure of mCRPC, nominate alterations that may inform prostate cancer treatment, and suggest that additional recurrent events in the non-coding mCRPC genome remain to be discovered.
Subject(s)
Prostatic Neoplasms, Castration-Resistant/pathology , Receptors, Androgen/genetics , Whole Genome Sequencing , Aged , Anilides/therapeutic use , Cyclin-Dependent Kinases/genetics , Cyclin-Dependent Kinases/metabolism , Enhancer Elements, Genetic/genetics , Gene Duplication , Gene Rearrangement , Genes, myc , Genetic Loci , Haplotypes , Humans , Male , Middle Aged , Neoplasm Metastasis , PTEN Phosphohydrolase/genetics , Phenotype , Prostate-Specific Antigen/blood , Prostatic Neoplasms, Castration-Resistant/drug therapy , Prostatic Neoplasms, Castration-Resistant/genetics , Protein Kinase Inhibitors/therapeutic use , Pyridines/therapeutic useABSTRACT
Programmed death 1 (PD-1) and PD ligand 1 (PD-L1) inhibitors have shown activity in metastatic clear cell renal cell carcinoma (ccRCC). Data on the activity of these agents in patients with non-clear cell RCC (nccRCC) or patients with sarcomatoid/rhabdoid differentiation are limited. In this multicenter analysis, we explored the efficacy of PD-1/PD-L1 inhibitors in patients with nccRCC or sarcomatoid/rhabdoid differentiation. Baseline and follow-up demographic, clinical, treatment, and radiographic data were collected. The primary endpoint was objective response rate. Secondary endpoints include time-to-treatment failure (TTF), overall survival (OS), and biomarker correlates. Forty-three patients were included: papillary (n = 14; 33%), chromophobe (n = 10; 23%), unclassified (n = 9; 21%), translocation (n = 3; 7%), and ccRCC with sarcomatoid differentiation (n = 7, 16%). Of those 43 patients, 11 patients (26%) had sarcomatoid and/or rhabdoid differentiation (n = 7 with ccRCC; n = 4 nccRCC). Overall, 8 patients (19%) objectively responded, including 4 patients (13%) who received PD-1/PD-L1 monotherapy. Responses were observed in patients with ccRCC with sarcomatoid and/or rhabdoid differentiation (n = 3/7, 43%), translocation RCC (n = 1/3, 33%), and papillary RCC (n = 4/14, 29%). The median TTF was 4.0 months [95% confidence interval (CI), 2.8-5.5] and median OS was 12.9 months (95% CI, 7.4-not reached). No specific genomic alteration was associated with clinical benefit. Modest antitumor activity for PD-1/PD-L1-blocking agents was observed in some patients with nccRCC. Further prospective studies are warranted to investigate the efficacy of PD-1/PD-L1 blockade in this heterogeneous patient population. Cancer Immunol Res; 6(7); 758-65. ©2018 AACR.
Subject(s)
Antineoplastic Agents, Immunological/therapeutic use , B7-H1 Antigen/antagonists & inhibitors , Carcinoma, Renal Cell/drug therapy , Carcinoma, Renal Cell/pathology , Kidney Neoplasms/drug therapy , Kidney Neoplasms/pathology , Programmed Cell Death 1 Receptor/antagonists & inhibitors , Adult , Aged , Antineoplastic Agents, Immunological/pharmacology , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , B7-H1 Antigen/genetics , B7-H1 Antigen/metabolism , Carcinoma, Renal Cell/metabolism , Carcinoma, Renal Cell/mortality , Cell Line, Tumor , Female , Humans , Kidney Neoplasms/metabolism , Kidney Neoplasms/mortality , Male , Middle Aged , Molecular Targeted Therapy , Prognosis , Programmed Cell Death 1 Receptor/genetics , Programmed Cell Death 1 Receptor/metabolism , Treatment Outcome , Young AdultABSTRACT
BACKGROUND: The genomic landscape of primary clear cell renal cell carcinoma (ccRCC) has been well described. However, little is known about cohort genomic alterations (GA) landscape in ccRCC metastases, or how it compares to primary tumours in aggregate. The genomic landscape of metastases may have biological, clinical, and therapeutic implications. METHODS: We collected targeted next-generation sequencing mutation calls from two independent cohorts and described the metastases GA landscape and descriptively compared it to the GA landscape in primary tumours. RESULTS: The cohort 1 (n = 578) consisted of 349 primary tumours and 229 metastases. Overall, the most common mutations in the metastases were VHL (66.8%), PBRM1 (41.87%), and SETD2 (24.7%). TP53 was more frequently mutated in metastases compared to primary tumours (14.85% versus 8.9%; p = 0.031). No other gene had significant difference in the cohort frequency of mutations between the metastases and primary tumours. Mutation burden was not significantly different between the metastases and primary tumours or between metastatic sites. The second cohort (n = 257) consisted of 177 primary tumours and 80 metastases. No differences in frequency of mutations or mutational burden were observed between primaries and metastases. CONCLUSIONS: These data support the theory that ccRCC primary tumours and metastases encompass a uniform distribution of common genomic alterations tested by next-generation sequencing targeted panels. This study does not address variability between matched primary tumours and metastases or the change in genomic alterations over time and after sequential systemic therapies.
Subject(s)
Carcinoma, Renal Cell/genetics , Carcinoma, Renal Cell/pathology , Kidney Neoplasms/genetics , Kidney Neoplasms/pathology , Adolescent , Adult , Aged , Aged, 80 and over , Child , Cohort Studies , Female , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Genomics/methods , High-Throughput Nucleotide Sequencing/methods , Humans , Male , Middle Aged , Neoplasm Metastasis/genetics , Young AdultABSTRACT
Comprehensive genomic characterization of prostate cancer has identified recurrent alterations in genes involved in androgen signaling, DNA repair, and PI3K signaling, among others. However, larger and uniform genomic analysis may identify additional recurrently mutated genes at lower frequencies. Here we aggregate and uniformly analyze exome sequencing data from 1,013 prostate cancers. We identify and validate a new class of E26 transformation-specific (ETS)-fusion-negative tumors defined by mutations in epigenetic regulators, as well as alterations in pathways not previously implicated in prostate cancer, such as the spliceosome pathway. We find that the incidence of significantly mutated genes (SMGs) follows a long-tail distribution, with many genes mutated in less than 3% of cases. We identify a total of 97 SMGs, including 70 not previously implicated in prostate cancer, such as the ubiquitin ligase CUL3 and the transcription factor SPEN. Finally, comparing primary and metastatic prostate cancer identifies a set of genomic markers that may inform risk stratification.