Kent W. Mouw

Somatic ERCC2 mutations are associated with a distinct genomic signature in urothelial tumors

Alterations in DNA repair pathways are common in tumors and can result in characteristic mutational signatures; however, a specific mutational signature associated with somatic alterations in the nucleotide- excision repair (NER) pathway has not yet been identified. Here we examine the mutational processes operating in urothelial cancer, a tumor type in which the core NER gene ERCC2 is significantly mutated. Analysis of three independent urothelial tumor cohorts demonstrates a strong association between somatic ERCC2 mutations and the activity of a mutational signature characterized by a broad spectrum of base changes. In addition, we note an association between the activity of this signature and smoking that is independent of ERCC2 mutation status, providing genomic evidence of tobacco-related mutagenesis in urothelial cancer. Together, these analyses identify an NER-related mutational signature and highlight the related roles of DNA damage and subsequent DNA repair in shaping tumor mutational landscape.

Second nonocular tumors among survivors of retinoblastoma treated with contemporary photon and proton radiotherapy

The leading cause of death among patients with hereditary retinoblastoma is second malignancy. Despite its high rate of efficacy, radiotherapy (RT) is often avoided due to fear of inducing a secondary tumor. Proton RT allows for significant sparing of nontarget tissue. The current study compared the risk of second malignancy in patients with retinoblastoma who were treated with photon and proton RT.

Shaping the Borrelia burgdorferi genome: crystal structure and binding properties of the DNA-bending protein Hbb

The genome of the Lyme disease‐causing spirochete Borrelia burgdorferi encodes only a single polypeptide from the integration host factor (IHF)/HU or ‘DNABII’ family of nucleoid‐associated proteins – Hbb. DNABII proteins induce large bends in DNA and serve as architectural factors in a variety of prokaryotic cellular processes. We have solved the crystal structure of an Hbb–DNA complex in which the DNA is bent by over 180°. We find that like IHF, Hbb relies exclusively on indirect readout to recognize its cognate site. Additional binding studies show that the sequence preferences of Hbb are related to, yet distinct from those of IHF. Defining these binding characteristics may help to uncover additional roles for Hbb in Borrelia DNA metabolism as well as further our understanding of the mechanism of indirect readout.

Architecture of a serine recombinase-DNA regulatory complex

Summary An essential feature of many site-specific recombination systems is their ability to regulate the direction and topology of recombination. Resolvases from the serine recombinase family assemble an interwound synaptic complex that harnesses negative supercoiling to drive the forward reaction and promote recombination between properly oriented sites. To better understand the interplay of catalytic and regulatory functions within these synaptic complexes, we have solved the structure of the regulatory site synapse in the Sin resolvase system. It reveals an unexpected synaptic interface between helix-turn-helix DNA-binding domains that is also highlighted in a screen for synapsis mutants. The tetramer defined by this interface provides the foundation for a robust model of the synaptic complex, assembled entirely from available crystal structures, that gives insight into how the catalytic activity of Sin and other serine recombinases may be regulated.

A Unique Subset of Epithelial Ovarian Cancers with Platinum Sensitivity and PARP Inhibitor Resistance

Platinum and PARP inhibitor (PARPi) sensitivity commonly coexist in epithelial ovarian cancer (EOC) due to the high prevalence of alterations in the homologous recombination (HR) DNA repair pathway that confer sensitivity to both drugs. In this report, we describe a unique subset of EOC with alterations in another DNA repair pathway, the nucleotide excision repair (NER) pathway, which may exhibit a discordance in sensitivities to these drugs. Specifically, 8% of high-grade serous EOC from The Cancer Genome Atlas dataset exhibited NER alterations, including nonsynonymous or splice site mutations and homozygous deletions of NER genes. Tumors with NER alterations were associated with improved overall survival (OS) and progression-free survival (PFS), compared with patients without NER alterations or BRCA1/2 mutations. Furthermore, patients with tumors with NER alterations had similar OS and PFS as BRCA1/2-mutated patients, suggesting that NER pathway inactivation in EOC conferred enhanced platinum sensitivity, similar to BRCA1/2-mutated tumors. Moreover, two NER mutations (ERCC6-Q524* and ERCC4-A583T), identified in the two most platinum-sensitive tumors, were functionally associated with platinum sensitivity in vitro. Importantly, neither NER alteration affected HR or conferred sensitivity to PARPi or other double-strand break-inducing agents. Overall, our findings reveal a new mechanism of platinum sensitivity in EOC that, unlike defective HR, may lead to a discordance in sensitivity to platinum and PARPi, with potential implications for previously reported and ongoing PARPi trials in this disease.

A mutational signature reveals alterations underlying deficient homologous recombination repair in breast cancer

Biallelic inactivation of BRCA1 or BRCA2 is associated with a pattern of genome-wide mutations known as signature 3. By analyzing ∼1,000 breast cancer samples, we confirmed this association and established that germline nonsense and frameshift variants in PALB2, but not in ATM or CHEK2, can also give rise to the same signature. We were able to accurately classify missense BRCA1 or BRCA2 variants known to impair homologous recombination (HR) on the basis of this signature. Finally, we show that epigenetic silencing of RAD51C and BRCA1 by promoter methylation is strongly associated with signature 3 and, in our data set, was highly enriched in basal-like breast cancers in young individuals of African descent.

DNA Damage and Repair Biomarkers of Immunotherapy Response

DNA-damaging agents are widely used in clinical oncology and exploit deficiencies in tumor DNA repair. Given the expanding role of immune checkpoint blockade as a therapeutic strategy, the interaction of tumor DNA damage with the immune system has recently come into focus, and it is now clear that the tumor DNA repair landscape has an important role in driving response to immune checkpoint blockade. Here, we summarize the mechanisms by which DNA damage and genomic instability have been found to shape the antitumor immune response and describe clinical efforts to use DNA repair biomarkers to guide use of immune-directed therapies.Significance: Only a subset of patients respond to immune checkpoint blockade, and reliable predictive biomarkers of response are needed to guide therapy decisions. DNA repair deficiency is common among tumors, and emerging experimental and clinical evidence suggests that features of genomic instability are associated with response to immune-directed therapies. Cancer Discov; 7(7); 675-93. ©2017 AACR.

Regulatory mutations in Sin recombinase support a structure-based model of the synaptosome.

The resolvase Sin regulates DNA strand exchange by assembling an elaborate interwound synaptosome containing catalytic and regulatory Sin tetramers, and an architectural DNA‐bending protein. The crystal structure of the regulatory tetramer was recently solved, providing new insights into the structural basis for regulation. Here we describe the selection and characterization of two classes of Sin mutations that, respectively, bypass or disrupt the functions of the regulatory tetramer. Activating mutations, which allow the catalytic tetramer to assemble and function independently at site I (the crossover site), were found at ∼20% of residues in the N‐terminal domain. The most strongly activating mutation (Q115R) stabilized a catalytically active synaptic tetramer in vitro. The positions of these mutations suggest that they act by destabilizing the conformation of the ground‐state site I‐bound dimers, or by stabilizing the altered conformation of the active catalytic tetramer. Mutations that block activation by the regulatory tetramer mapped to just two residues, F52 and R54, supporting a functional role for a previously reported crystallographic dimer–dimer interface. We suggest how F52/R54 contacts between regulatory and catalytic subunits might promote assembly of the active catalytic tetramer within the synaptosome.

Crosstalk between the nucleotide excision repair and Fanconi anemia/BRCA pathways.

Cells have evolved multiple distinct DNA repair pathways to efficiently correct a variety of genotoxic lesions, and decades of study have led to an improved understanding of the mechanisms and regulation of these individual pathways. However, there is now an increasing appreciation that extensive crosstalk exists among DNA repair pathways and that this crosstalk serves to increase the efficiency and diversity of response to damage. The Fanconi anemia (FA)/BRCA and nucleotide excision repair (NER) pathways have been shown to share common factors, and often work in concert to repair damage. Genomic studies are now revealing that many tumors harbor somatic mutations in FA/BRCA or NER genes, which may provide a growth advantage, but which could also be exploited therapeutically.

Factors Associated With Long-term Speech and Swallowing Outcomes After Chemoradiotherapy for Locoregionally Advanced Head and Neck Cancer

OBJECTIVE to identify factors that influence patient-centered measures of speech and swallowing function after successful use of chemoradiotherapy to treat cancers of the head and neck. DESIGN patients previously enrolled in a phase 2 trial using induction chemotherapy consisting of carboplatin and paclitaxel followed by chemoradiotherapy with paclitaxel, fluorouracil, hydroxyurea, and 1 of 3 radiation dose levels were assigned speaking and swallowing scores at follow-up ranging from 1 to 4, with 1 representing normal speech or swallowing and 4 representing significant sustained deficits. PATIENTS one hundred eighty-four patients with locoregionally advanced head and neck cancer. MAIN OUTCOME MEASURES speech and swallowing function after chemoradiotherapy. RESULTS of the 222 patients originally enrolled in the trial, 184 were alive and free of locoregional recurrence at the outset of this study. Of these eligible patients, 163 (88.6%) were assigned a speaking score of 1 through 4 at an average of 34.8 (range, 1.5-76.4) months after completion of treatment, whereas 166 patients (90.2%) were assigned a swallowing score of 1 through 4 at an average of 34.5 (range, 1.0-76.4) months after completion of treatment. Most patients (84.7% with speaking scores and 63.3% with swallowing scores) had no residual deficit and were assigned scores of 1. Factors that were associated with worse speaking outcomes included female sex, smoking history, hypopharyngeal or laryngeal primary sites, and poor response to induction chemotherapy; factors associated with worse swallowing outcomes included advanced patient age, poor performance status, primary site, and neck dissection. CONCLUSIONS among patients successfully treated for locoregionally advanced cancers of the head and neck, several factors correlate with speaking and swallowing outcomes. Because advances in therapy have led to improved survival in these patients, understanding and controlling adverse effects of treatment should continue to be an active area of investigation.