Overcoming Barriers to Tumor Genomic Profiling through Direct-to-Patient Outreach.

PURPOSE: To overcome barriers to genomic testing for patients with rare cancers, we initiated a program to offer free clinical tumor genomic testing worldwide to patients with select rare cancer subtypes. EXPERIMENTAL DESIGN: Patients were recruited through social media outreach and engagement with disease-specific advocacy groups, with a focus on patients with histiocytosis, germ cell tumors (GCT), and pediatric cancers. Tumors were analyzed using the MSK-IMPACT next-generation sequencing assay with the return of results to patients and their local physicians. Whole-exome recapture was performed for female patients with GCTs to define the genomic landscape of this rare cancer subtype. RESULTS: A total of 333 patients were enrolled, and tumor tissue was received for 288 (86.4%), with 250 (86.8%) having tumor DNA of sufficient quality for MSK-IMPACT testing. Eighteen patients with histiocytosis have received genomically guided therapy to date, of whom 17 (94%) have had clinical benefit with a mean treatment duration of 21.7 months (range, 6-40+). Whole-exome sequencing of ovarian GCTs identified a subset with haploid genotypes, a phenotype rarely observed in other cancer types. Actionable genomic alterations were rare in ovarian GCT (28%); however, 2 patients with ovarian GCTs with squamous transformation had high tumor mutational burden, one of whom had a complete response to pembrolizumab. CONCLUSIONS: Direct-to-patient outreach can facilitate the assembly of cohorts of rare cancers of sufficient size to define their genomic landscape. By profiling tumors in a clinical laboratory, results could be reported to patients and their local physicians to guide treatment. See related commentary by Desai and Subbiah, p. 2339.

Targeting Aurora B kinase prevents and overcomes resistance to EGFR inhibitors in lung cancer by enhancing BIM- and PUMA-mediated apoptosis.

The clinical success of EGFR inhibitors in EGFR-mutant lung cancer is limited by the eventual development of acquired resistance. We hypothesize that enhancing apoptosis through combination therapies can eradicate cancer cells and reduce the emergence of drug-tolerant persisters. Through high-throughput screening of a custom library of ∼1,000 compounds, we discover Aurora B kinase inhibitors as potent enhancers of osimertinib-induced apoptosis. Mechanistically, Aurora B inhibition stabilizes BIM through reduced Ser87 phosphorylation, and transactivates PUMA through FOXO1/3. Importantly, osimertinib resistance caused by epithelial-mesenchymal transition (EMT) activates the ATR-CHK1-Aurora B signaling cascade and thereby engenders hypersensitivity to respective kinase inhibitors by activating BIM-mediated mitotic catastrophe. Combined inhibition of EGFR and Aurora B not only efficiently eliminates cancer cells but also overcomes resistance beyond EMT.

Noninvasive Detection of Polyclonal Acquired Resistance to FGFR Inhibition in Patients With Cholangiocarcinoma Harboring FGFR2 Alterations.

PURPOSE: Fibroblast growth factor receptor (FGFR) 2 alterations, present in 5%-15% of intrahepatic cholangiocarcinomas (IHC), are targets of FGFR-directed therapies. Acquired resistance is common among patients who respond. Biopsies at the time of acquired resistance to targeted agents may not always be feasible and may not capture the genetic heterogeneity that could exist within a patient. We studied circulating tumor DNA (ctDNA) as a less invasive means of potentially identifying genomic mechanisms of resistance to FGFR-targeted therapies. MATERIALS AND METHODS: Serial blood samples were collected from eight patients with FGFR-altered cholangiocarcinoma for ctDNA isolation and next-generation sequencing (NGS) throughout treatment and at resistance to anti-FGFR-targeted therapy. ctDNA was sequenced using a custom ultra-deep coverage NGS panel, incorporating dual index primers and unique molecular barcodes to enable high-sensitivity mutation detection. RESULTS: Thirty-one acquired mutations in FGFR2, 30/31 located in the kinase domain, were identified at resistance in six of eight patients with detectable ctDNA. Up to 13 independent FGFR2 mutations were detected per patient, indicative of striking genomic concordance among resistant subclones. CONCLUSION: ctDNA could be an effective means to longitudinally monitor for acquired resistance in FGFR2-altered IHC. The numerous acquired genetic alterations in FGFR2 suggest frequent polyclonal mechanisms of resistance that cannot be detected from single-site tissue biopsies.

Tumor fraction-guided cell-free DNA profiling in metastatic solid tumor patients.

BACKGROUND: Cell-free DNA (cfDNA) profiling is increasingly used to guide cancer care, yet mutations are not always identified. The ability to detect somatic mutations in plasma depends on both assay sensitivity and the fraction of circulating DNA in plasma that is tumor-derived (i.e., cfDNA tumor fraction). We hypothesized that cfDNA tumor fraction could inform the interpretation of negative cfDNA results and guide the choice of subsequent assays of greater genomic breadth or depth. METHODS: Plasma samples collected from 118 metastatic cancer patients were analyzed with cf-IMPACT, a modified version of the FDA-authorized MSK-IMPACT tumor test that can detect genomic alterations in 410 cancer-associated genes. Shallow whole genome sequencing (sWGS) was also performed in the same samples to estimate cfDNA tumor fraction based on genome-wide copy number alterations using z-score statistics. Plasma samples with no somatic alterations detected by cf-IMPACT were triaged based on sWGS-estimated tumor fraction for analysis with either a less comprehensive but more sensitive assay (MSK-ACCESS) or broader whole exome sequencing (WES). RESULTS: cfDNA profiling using cf-IMPACT identified somatic mutations in 55/76 (72%) patients for whom MSK-IMPACT tumor profiling data were available. A significantly higher concordance of mutational profiles and tumor mutational burden (TMB) was observed between plasma and tumor profiling for plasma samples with a high tumor fraction (z-score≥5). In the 42 patients from whom tumor data was not available, cf-IMPACT identified mutations in 16/42 (38%). In total, cf-IMPACT analysis of plasma revealed mutations in 71/118 (60%) patients, with clinically actionable alterations identified in 30 (25%), including therapeutic targets of FDA-approved drugs. Of the 47 samples without alterations detected and low tumor fraction (z-score<5), 29 had sufficient material to be re-analyzed using a less comprehensive but more sensitive assay, MSK-ACCESS, which revealed somatic mutations in 14/29 (48%). Conversely, 5 patients without alterations detected by cf-IMPACT and with high tumor fraction (z-score≥5) were analyzed by WES, which identified mutational signatures and alterations in potential oncogenic drivers not covered by the cf-IMPACT panel. Overall, we identified mutations in 90/118 (76%) patients in the entire cohort using the three complementary plasma profiling approaches. CONCLUSIONS: cfDNA tumor fraction can inform the interpretation of negative cfDNA results and guide the selection of subsequent sequencing platforms that are most likely to identify clinically-relevant genomic alterations.

Germ Cell Tumor Molecular Heterogeneity Revealed Through Analysis of Primary and Metastasis Pairs.

PURPOSE: Although primary germ cell tumors (GCTs) have been extensively characterized, molecular analysis of metastatic sites has been limited. We performed whole-exome sequencing and targeted next-generation sequencing on paired primary and metastatic GCT samples in a patient cohort enriched for cisplatin-resistant disease. PATIENTS AND METHODS: Tissue sequencing was performed on 100 tumor specimens from 50 patients with metastatic GCT, and sequencing of plasma cell-free DNA was performed for a subset of patients. RESULTS: The mutational landscape of primary and metastatic pairs from GCT patients was highly discordant (68% of all somatic mutations were discordant). Whereas genome duplication was common and highly concordant between primary and metastatic samples, only 25% of primary-metastasis pairs had ≥ 50% concordance at the level of DNA copy number alterations (CNAs). Evolutionary-based analyses revealed that most mutations arose after CNAs at the respective loci in both primary and metastatic samples, with oncogenic mutations enriched in the set of early-occurring mutations versus variants of unknown significance (VUSs). TP53 pathway alterations were identified in nine cisplatin-resistant patients and had the highest degree of concordance in primary and metastatic specimens, consistent with their association with this treatment-resistant phenotype. CONCLUSION: Analysis of paired primary and metastatic GCT specimens revealed significant molecular heterogeneity for both CNAs and somatic mutations. Among loci demonstrating serial genetic evolution, most somatic mutations arose after CNAs, but oncogenic mutations were enriched in the set of early-occurring mutations as compared with VUSs. Alterations in TP53 were clonal when present and shared among primary-metastasis pairs.

Author Correction: Tumour lineage shapes BRCA-mediated phenotypes.

An Amendment to this paper has been published and can be accessed via a link at the top of the paper.

Efficacy and Determinants of Response to HER Kinase Inhibition in HER2-Mutant Metastatic Breast Cancer.

HER2 mutations define a subset of metastatic breast cancers with a unique mechanism of oncogenic addiction to HER2 signaling. We explored activity of the irreversible pan-HER kinase inhibitor neratinib, alone or with fulvestrant, in 81 patients with HER2-mutant metastatic breast cancer. Overall response rate was similar with or without estrogen receptor (ER) blockade. By comparison, progression-free survival and duration of response appeared longer in ER+ patients receiving combination therapy, although the study was not designed for direct comparison. Preexistent concurrent activating HER2 or HER3 alterations were associated with poor treatment outcome. Similarly, acquisition of multiple HER2-activating events, as well as gatekeeper alterations, were observed at disease progression in a high proportion of patients deriving clinical benefit from neratinib. Collectively, these data define HER2 mutations as a therapeutic target in breast cancer and suggest that coexistence of additional HER signaling alterations may promote both de novo and acquired resistance to neratinib. SIGNIFICANCE: HER2 mutations define a targetable breast cancer subset, although sensitivity to irreversible HER kinase inhibition appears to be modified by the presence of concurrent activating genomic events in the pathway. These findings have implications for potential future combinatorial approaches and broader therapeutic development for this genomically defined subset of breast cancer.This article is highlighted in the In This Issue feature, p. 161.

Phase 2 study of LY3023414 in patients with advanced endometrial cancer harboring activating mutations in the PI3K pathway.

BACKGROUND: PI3K pathway activation is common in endometrial cancer. We evaluated the safety and efficacy of the dual PI3K/mTOR inhibitor, LY3023414, in patients with advanced endometrial cancer harboring activating mutations in the PI3K pathway. METHODS: We conducted a single-arm phase 2 study of monotherapy LY3023414. Eligible patients had advanced endometrial cancer of any grade, prior management with 1-4 cytotoxic lines, and PI3K pathway activation prospectively defined as a loss-of-function PTEN alteration or activating alteration in PIK3CA, AKT1, PIK3R1, PIK3R2, or MTOR. The primary objective was best overall response rate (ORR) per RECIST 1.1. RESULTS: Twenty-eight patients were treated; histologies included endometroid (39%), carcinosarcoma (25%), serous (21%), and mixed (14%). Patients were heavily pretreated, with a median of 2 prior cytotoxic lines (range, 1-3). The most common alterations involved PIK3CA (68%), PTEN (43%), and PIK3R1 (32%). In the 25 efficacy-evaluable patients, the ORR was 16% (90% CI, 7%-100%), and the clinical benefit rate was 28% (90% CI, 16%-100%). Four patients had a confirmed partial response, and 2 responses lasted for >9 months. The median progression-free survival and overall survival were 2.5 months (95% CI, 1.2-3.0) and 9.2 months (95% CI, 5.0-15.9), respectively. The most common all-grade treatment-related adverse events were anemia (71%), hyperglycemia (71%), hypoalbuminemia (68%), and hypophosphatemia (61%). No correlation between molecular alterations and response was observed. CONCLUSION: In patients with heavily pretreated advanced endometrial cancer prospectively selected for tumors with activating PI3K pathway mutations, LY3023414 demonstrated modest single-agent activity and a manageable safety profile.

Resistance to TRK inhibition mediated by convergent MAPK pathway activation.

TRK fusions are found in a variety of cancer types, lead to oncogenic addiction, and strongly predict tumor-agnostic efficacy of TRK inhibition1-8. With the recent approval of the first selective TRK inhibitor, larotrectinib, for patients with any TRK-fusion-positive adult or pediatric solid tumor, to identify mechanisms of treatment failure after initial response has become of immediate therapeutic relevance. So far, the only known resistance mechanism is the acquisition of on-target TRK kinase domain mutations, which interfere with drug binding and can potentially be addressable through second-generation TRK inhibitors9-11. Here, we report off-target resistance in patients treated with TRK inhibitors and in patient-derived models, mediated by genomic alterations that converge to activate the mitogen-activated protein kinase (MAPK) pathway. MAPK pathway-directed targeted therapy, administered alone or in combination with TRK inhibition, re-established disease control. Experimental modeling further suggests that upfront dual inhibition of TRK and MEK may delay time to progression in cancer types prone to the genomic acquisition of MAPK pathway-activating alterations. Collectively, these data suggest that a subset of patients will develop off-target mechanisms of resistance to TRK inhibition with potential implications for clinical management and future clinical trial design.

Tumour lineage shapes BRCA-mediated phenotypes.

Mutations in BRCA1 and BRCA2 predispose individuals to certain cancers1-3, and disease-specific screening and preventative strategies have reduced cancer mortality in affected patients4,5. These classical tumour-suppressor genes have tumorigenic effects associated with somatic biallelic inactivation, although haploinsufficiency may also promote the formation and progression of tumours6,7. Moreover, BRCA1/2-mutant tumours are often deficient in the repair of double-stranded DNA breaks by homologous recombination8-13, and consequently exhibit increased therapeutic sensitivity to platinum-containing therapy and inhibitors of poly-(ADP-ribose)-polymerase (PARP)14,15. However, the phenotypic and therapeutic relevance of mutations in BRCA1 or BRCA2 remains poorly defined in most cancer types. Here we show that in the 2.7% and 1.8% of patients with advanced-stage cancer and germline pathogenic or somatic loss-of-function alterations in BRCA1/2, respectively, selective pressure for biallelic inactivation, zygosity-dependent phenotype penetrance, and sensitivity to PARP inhibition were observed only in tumour types associated with increased heritable cancer risk in BRCA1/2 carriers (BRCA-associated cancer types). Conversely, among patients with non-BRCA-associated cancer types, most carriers of these BRCA1/2 mutation types had evidence for tumour pathogenesis that was independent of mutant BRCA1/2. Overall, mutant BRCA is an indispensable founding event for some tumours, but in a considerable proportion of other cancers, it appears to be biologically neutral-a difference predominantly conditioned by tumour lineage-with implications for disease pathogenesis, screening, design of clinical trials and therapeutic decision-making.

Author Correction: HER kinase inhibition in patients with HER2- and HER3-mutant cancers.

The 'Competing interests' statement of this Article has been updated; please see the accompanying Amendment. The original Article has not been corrected online.

Tracking tumour evolution in glioma through liquid biopsies of cerebrospinal fluid.

Diffuse gliomas are the most common malignant brain tumours in adults and include glioblastomas and World Health Organization (WHO) grade II and grade III tumours (sometimes referred to as lower-grade gliomas). Genetic tumour profiling is used to classify disease and guide therapy1,2, but involves brain surgery for tissue collection; repeated tumour biopsies may be necessary for accurate genotyping over the course of the disease3-10. While the detection of circulating tumour DNA (ctDNA) in the blood of patients with primary brain tumours remains challenging11,12, sequencing of ctDNA from the cerebrospinal fluid (CSF) may provide an alternative way to genotype gliomas with lower morbidity and cost13,14. We therefore evaluated the representation of the glioma genome in CSF from 85 patients with gliomas who underwent a lumbar puncture because they showed neurological signs or symptoms. Here we show that tumour-derived DNA was detected in CSF from 42 out of 85 patients (49.4%) and was associated with disease burden and adverse outcome. The genomic landscape of glioma in the CSF included a broad spectrum of genetic alterations and closely resembled the genomes of tumour biopsies. Alterations that occur early during tumorigenesis, such as co-deletion of chromosome arms 1p and 19q (1p/19q codeletion) and mutations in the metabolic genes isocitrate dehydrogenase 1 (IDH1) or IDH21,2, were shared in all matched ctDNA-positive CSF-tumour pairs, whereas growth factor receptor signalling pathways showed considerable evolution. The ability to monitor the evolution of the glioma genome through a minimally invasive technique could advance the clinical development and use of genotype-directed therapies for glioma, one of the most aggressive human cancers.

Isoform Switching as a Mechanism of Acquired Resistance to Mutant Isocitrate Dehydrogenase Inhibition.

Somatic mutations in cytosolic or mitochondrial isoforms of isocitrate dehydrogenase (IDH1 or IDH2, respectively) contribute to oncogenesis via production of the metabolite 2-hydroxyglutarate (2HG). Isoform-selective IDH inhibitors suppress 2HG production and induce clinical responses in patients with IDH1- and IDH2-mutant malignancies. Despite the promising activity of IDH inhibitors, the mechanisms that mediate resistance to IDH inhibition are poorly understood. Here, we describe four clinical cases that identify mutant IDH isoform switching, either from mutant IDH1 to mutant IDH2 or vice versa, as a mechanism of acquired clinical resistance to IDH inhibition in solid and liquid tumors. SIGNIFICANCE: IDH-mutant cancers can develop resistance to isoform-selective IDH inhibition by "isoform switching" from mutant IDH1 to mutant IDH2 or vice versa, thereby restoring 2HG production by the tumor. These findings underscore a role for continued 2HG production in tumor progression and suggest therapeutic strategies to prevent or overcome resistance.This article is highlighted in the In This Issue feature, p. 1494.

HER kinase inhibition in patients with HER2- and HER3-mutant cancers.

Somatic mutations of ERBB2 and ERBB3 (which encode HER2 and HER3, respectively) are found in a wide range of cancers. Preclinical modelling suggests that a subset of these mutations lead to constitutive HER2 activation, but most remain biologically uncharacterized. Here we define the biological and therapeutic importance of known oncogenic HER2 and HER3 mutations and variants of unknown biological importance by conducting a multi-histology, genomically selected, 'basket' trial using the pan-HER kinase inhibitor neratinib (SUMMIT; clinicaltrials.gov identifier NCT01953926). Efficacy in HER2-mutant cancers varied as a function of both tumour type and mutant allele to a degree not predicted by preclinical models, with the greatest activity seen in breast, cervical and biliary cancers and with tumours that contain kinase domain missense mutations. This study demonstrates how a molecularly driven clinical trial can be used to refine our biological understanding of both characterized and new genomic alterations with potential broad applicability for advancing the paradigm of genome-driven oncology.

AKT Inhibition in Solid Tumors With AKT1 Mutations.

Purpose AKT1 E17K mutations are oncogenic and occur in many cancers at a low prevalence. We performed a multihistology basket study of AZD5363, an ATP-competitive pan-AKT kinase inhibitor, to determine the preliminary activity of AKT inhibition in AKT-mutant cancers. Patients and Methods Fifty-eight patients with advanced solid tumors were treated. The primary end point was safety; secondary end points were progression-free survival (PFS) and response according to Response Evaluation Criteria in Solid Tumors (RECIST). Tumor biopsies and plasma cell-free DNA (cfDNA) were collected in the majority of patients to identify predictive biomarkers of response. Results In patients with AKT1 E17K-mutant tumors (n = 52) and a median of five lines of prior therapy, the median PFS was 5.5 months (95% CI, 2.9 to 6.9 months), 6.6 months (95% CI, 1.5 to 8.3 months), and 4.2 months (95% CI, 2.1 to 12.8 months) in patients with estrogen receptor-positive breast, gynecologic, and other solid tumors, respectively. In an exploratory biomarker analysis, imbalance of the AKT1 E17K-mutant allele, most frequently caused by copy-neutral loss-of-heterozygosity targeting the wild-type allele, was associated with longer PFS (hazard ratio [HR], 0.41; P = .04), as was the presence of coincident PI3K pathway hotspot mutations (HR, 0.21; P = .045). Persistent declines in AKT1 E17K in cfDNA were associated with improved PFS (HR, 0.18; P = .004) and response ( P = .025). Responses were not restricted to patients with detectable AKT1 E17K in pretreatment cfDNA. The most common grade ≥ 3 adverse events were hyperglycemia (24%), diarrhea (17%), and rash (15.5%). Conclusion This study provides the first clinical data that AKT1 E17K is a therapeutic target in human cancer. The genomic context of the AKT1 E17K mutation further conditioned response to AZD5363.

Genomic Biomarkers of a Randomized Trial Comparing First-line Everolimus and Sunitinib in Patients with Metastatic Renal Cell Carcinoma.

BACKGROUND: Metastatic renal cell carcinoma (RCC) patients are commonly treated with vascular endothelial growth factor (VEGF) inhibitors or mammalian target of rapamycin inhibitors. Correlations between somatic mutations and first-line targeted therapy outcomes have not been reported on a randomized trial. OBJECTIVE: To evaluate the relationship between tumor mutations and treatment outcomes in RECORD-3, a randomized trial comparing first-line everolimus (mTOR inhibitor) followed by sunitinib (VEGF inhibitor) at progression with the opposite sequence in 471 metastatic RCC patients. DESIGN, SETTING, AND PARTICIPANTS: Targeted sequencing of 341 cancer genes at ∼540× coverage was performed on available tumor samples from 258 patients; 220 with clear cell histology (ccRCC). OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Associations between somatic mutations and median first-line progression free survival (PFS1L) and overall survival were determined in metastatic ccRCC using Cox proportional hazards models and log-rank tests. RESULTS AND LIMITATIONS: Prevalent mutations (≥ 10%) were VHL (75%), PBRM1 (46%), SETD2 (30%), BAP1 (19%), KDM5C (15%), and PTEN (12%). With first-line everolimus, PBRM1 and BAP1 mutations were associated with longer (median [95% confidence interval {CI}] 12.8 [8.1, 18.4] vs 5.5 [3.1, 8.4] mo) and shorter (median [95% CI] 4.9 [2.9, 8.1] vs 10.5 [7.3, 12.9] mo) PFS1L, respectively. With first-line sunitinib, KDM5C mutations were associated with longer PFS1L (median [95% CI] of 20.6 [12.4, 27.3] vs 8.3 [7.8, 11.0] mo). Molecular subgroups of metastatic ccRCC based on PBRM1, BAP1, and KDM5C mutations could have predictive values for patients treated with VEGF or mTOR inhibitors. Most tumor DNA was obtained from primary nephrectomy samples (94%), which could impact correlation statistics. CONCLUSIONS: PBRM1, BAP1, and KDM5C mutations impact outcomes of targeted therapies in metastatic ccRCC patients. PATIENT SUMMARY: Large-scale genomic kidney cancer studies reported novel mutations and heterogeneous features among individual tumors, which could contribute to varied clinical outcomes. We demonstrated correlations between somatic mutations and treatment outcomes in clear cell renal cell carcinoma, supporting the value of genomic classification in prospective studies.

Allele-specific splicing effects on DKKL1 and ZNF419 transcripts in HeLa cells.

Allele-specific splicing is the production of different RNA isoforms from different alleles of a gene. Altered splicing patterns such as exon skipping can have a dramatic effect on the final protein product yet have traditionally proven difficult to predict. We investigated the splicing effects of a set of nine single nucleotide polymorphisms (SNPs) which are predicted to have a direct impact on mRNA splicing, each in a different gene. Predictions were based on SNP location relative to splice junctions and intronic/exonic splicing elements, combined with an analysis of splice isoform expression data from public sources. Of the nine genes tested, six SNPs led to direct impacts on mRNA splicing as determined by the splicing reporter minigene assay and RT-PCR in human HeLa cells, of which four were allele-specific effects. These included previously unreported alternative splicing patterns in the genes ZNF419 and DKKL1. Notably, the SNP in ZNF419, a transcription factor, leads to the deletion of a DNA-binding domain from the protein and is associated with an expression QTL, while the SNP in DKKL1 leads to shortened transcripts predicted to produce a truncated protein. We conclude that the impact of SNP mutations on mRNA splicing, and its biological relevance, can be predicted by integrating SNP position with available data on relative isoform abundance in human cell lines.

Phase II Trial and Correlative Genomic Analysis of Everolimus Plus Bevacizumab in Advanced Non-Clear Cell Renal Cell Carcinoma.

Purpose The decreased effectiveness of single-agent targeted therapies in advanced non-clear cell renal cell carcinoma (ncRCC) compared with clear cell renal cell carcinoma (RCC) supports the study of combination regimens. We evaluated the efficacy of everolimus plus bevacizumab in patients with metastatic ncRCC. Patients and Methods In this single-center phase II trial, treatment-naive patients received everolimus 10 mg oral once per day plus bevacizumab 10 mg/kg intravenously every 2 weeks. The primary end point was progression-free survival (PFS) at 6 months. Correlative analyses explored candidate tissue biomarkers through next-generation sequencing. Results Thirty-five patients were enrolled with the following histologic subtypes: chromophobe (n = 5), papillary (n = 5), and medullary (n = 2) RCC and unclassified RCC (uRCC, n = 23). The majority of patients had papillary growth as a major component (n = 14). For 34 evaluable patients, median PFS, overall survival, and objective response rate (ORR) were 11.0 months, 18.5 months, and 29%, respectively. PFS varied by histology ( P < .001), and ORR was higher in patients with significant papillary (seven of 18) or chromophobe (two of five) elements than for others (one of 11). Presence of papillary features were associated with benefit, including uRCC, where it correlated with ORR (43% v 11%), median PFS (12.9 v 1.9 months), and overall survival (28.2 v 9.3 months; P < .001). Several genetic alterations seemed to segregate by histology. In particular, somatic mutations in ARID1A were seen in five of 14 patients with papillary features but not in other RCC variants. All five patients achieved treatment benefit. Conclusion The study suggests efficacy for this combination in patients with ncRCC characterized by papillary features. Distinct mutational profiles among ncRCCs vary according to specific histology.

Evaluating Cancer of the Central Nervous System Through Next-Generation Sequencing of Cerebrospinal Fluid.

PURPOSE: Cancer spread to the central nervous system (CNS) often is diagnosed late and is unresponsive to therapy. Mechanisms of tumor dissemination and evolution within the CNS are largely unknown because of limited access to tumor tissue. MATERIALS AND METHODS: We sequenced 341 cancer-associated genes in cell-free DNA from cerebrospinal fluid (CSF) obtained through routine lumbar puncture in 53 patients with suspected or known CNS involvement by cancer. RESULTS: We detected high-confidence somatic alterations in 63% (20 of 32) of patients with CNS metastases of solid tumors, 50% (six of 12) of patients with primary brain tumors, and 0% (zero of nine) of patients without CNS involvement by cancer. Several patients with tumor progression in the CNS during therapy with inhibitors of oncogenic kinases harbored mutations in the kinase target or kinase bypass pathways. In patients with glioma, the most common malignant primary brain tumor in adults, examination of cell-free DNA uncovered patterns of tumor evolution, including temozolomide-associated mutations. CONCLUSION: The study shows that CSF harbors clinically relevant genomic alterations in patients with CNS cancers and should be considered for liquid biopsies to monitor tumor evolution in the CNS.

ALCAM is indirectly modulated by miR-125b in MCF7 cells.

MicroRNA (miRNA) deregulation is associated with various cancers. Among an expanding list of cancer-related miRNAs, deregulation of miR-125b has been well documented in many cancers including breast. Based on current knowledge, miR-125b is considered to be a tumor suppressor in breast cancers. While important messenger RNA (mRNA) targets have been defined for miR-125b, here, we aimed to further investigate direct/indirect consequences of miR-125b expression in breast cancer cells by using a transcriptome approach. Upon miR-125b expression, a total of 138 cancer-related genes were found to be differentially expressed in breast cancer cells. While only a few of these were predicted to be direct mRNA targets, majority of the gene expression changes were potentially downstream and indirect effects of miR-125b expression. Among these, activated leukocyte antigen molecule (ALCAM) mRNA and protein levels were found to be highly significantly increased upon miR-125b expression. Given the tumor suppressor role of miR-125b in our model system, upon silencing of ALCAM expression, cell proliferation rate re-increased in miR-125b-expressing cells. While ALCAM's possible context-dependent roles are not clear in breast cancer, a diverse expression pattern of ALCAM mRNA was detected in a panel of breast cancer patient samples. Differentially expressed/regulated cancer-related genes upon miR-125b expression along with the significant increase of ALCAM are of future interest to understand how deregulated expression of miR-125b may have a tumor suppressor role in breast and other cancers.