Cell-Free DNA Sequencing Is Clinically Useful in Pancreatobiliary Cancer
See article, p. 1040.
Sequencing of tumor DNA and cfDNA from 26 patients with advanced tumors was prospectively analyzed.
cfDNA sequencing detects tumor-derived mutations with high sensitivity and specificity.
Profiling of cfDNA enables increased identification of potentially actionable mutations.
The development of targeted therapies in pancreatobiliary cancers has been hampered by the difficulty in obtaining biopsies with sufficient tumor cell content for genomic sequencing analyses. As an alternative approach, Zill and colleagues prospectively assessed the feasibility and efficacy of cell-free DNA (cfDNA) sequencing in 26 patients with advanced pancreatic or biliary carcinomas. Next-generation sequencing (NGS) of a panel of 54 genes was performed to evaluate the concordance between cfDNA and tumor biopsy–derived DNA profiling. In comparison with tumor biopsy sequencing, which detected mutations in 16 (62%) patients, cfDNA testing identified mutations in 22 (85%) patients, including several patients with insufficient biopsy samples. NGS analysis of cfDNA detected mutations in the five most frequently mutated genes with 100% specificity and greater than 90% sensitivity and diagnostic accuracy compared with tumor biopsy testing; among the 17 patients with matched plasma and tumor biopsy sequencing results, the concordance between the two approaches was greater than 90% across all genes. Furthermore, cfDNA sequencing resulted in increased identification of potentially actionable mutations compared with tissue-based NGS, including clinically relevant mutations in the BRAF, FGFR2, and EGFR oncogenes, the last of which was associated with durable clinical benefit in a patient treated with the EGFR inhibitor erlotinib. These findings demonstrate the feasibility and accuracy of cfDNA sequencing in detecting tumor-derived mutations in advanced pancreatobiliary cancers and suggest that this approach may improve personalized treatment of this deadly disease.
Inhibition of TRK Fusions Provides Clinical Benefit
See article, p. 1049.
The TRK inhibitor LOXO-101 inhibits the growth of TRK fusion–positive cells in vitro and in vivo.
An oncogenic LMNA–NTRK1 fusion gene was detected in a patient with metastatic soft-tissue sarcoma.
LOXO-101 treatment induced rapid and substantial tumor regression in this patient.
Fusions involving the tropomyosin-related kinase (TRK) family of neurotrophin receptor tyrosine kinases have been identified in various human cancers and suggested to function as oncogenic drivers. The orally available, highly selective small-molecule TRK family inhibitor LOXO-101 is currently being tested in a phase I dose-escalation study in patients with advanced solid tumors. Doebele and colleagues report the first clinical response of a patient to LOXO-101 as part of this trial. Preclinical studies demonstrated that LOXO-101 inhibited the proliferation of cancer cell lines harboring oncogenic TRK fusions in a dose-dependent manner and resulted in suppression of tumor growth in vivo, consistent with previous studies. A 41-year-old woman who presented with undifferentiated soft-tissue sarcoma and lung metastases was found to harbor a gene fusion consisting of the 5′ region of lamin A/C (LMNA) and the 3′ region of neurotrophic tyrosine kinase receptor type 1 (NTRK1, encoding TRKA). Proximity ligation assays demonstrated that the LMNA–NTRK1 fusion protein was functional and exhibited oncogenic signaling in patient-derived tumor cells. After experiencing tumor progression after initial treatment with sorafenib and chemotherapy followed by surgical resection, the patient was enrolled in the LOXO-101 phase I trial. Administration of LOXO-101 resulted in rapid and substantial tumor regression and significant improvement in exertional dyspnea and oxygen saturation in the absence of drug-related adverse events. These results clinically validate this TRK fusion as a molecular driver and therapeutic target.
Enhancer-Associated Genomic Rearrangements Drive Lymphomagenesis
See article, p. 1058.
PEAR-ChIP simultaneously identifies genomic rearrangements and active enhancers.
Lymphoma subtype–specific enhancers in the MYC locus drive oncogene expression.
The BCL6 locus acts as a donor or recipient of enhancers in a context-specific manner.
B-cell lymphomas (BCL) frequently harbor recurrent genomic rearrangements, many of which result in aberrantly high expression of oncogenes due to translocations or amplifications involving enhancer elements. Histone H3 Lys27 acetylation (H3K27ac) is a mark of active enhancers and an attribute of immunoglobulin loci that undergo physiologic or oncogenic rearrangement in BCLs. To detect enhancer-associated genomic rearrangements that occur within acetylated elements, Ryan, Drier, and colleagues developed Pinpointing Enhancer-Associated Rearrangements by Chromatin Immunoprecipitation (PEAR-ChIP), which integrates paired-end rearrangement analysis with H3K27ac ChIP-sequencing. PEAR-ChIP analysis of lymphoma cell lines and patient biopsies identified known genomic rearrangements as well as previously unidentified enhancer tandem duplications that may act as gain-of-function oncogenic alterations, including tandem duplication of a BCL6-interacting super-enhancer. Activation of BCL6 enhancers was driven by recruitment of the p300 acetyltransferase by the transcription factor myocyte enhancer factor 2B (MEF2B). PEAR-ChIP also identified lymphoma subtype–specific enhancers that interact with the MYC promoter and contained known lymphoma risk variants. Furthermore, the known recurrent t(3;8) BCL6–MYC rearrangement resulted in activation of MYC expression by BCL6 enhancers, demonstrating that an oncogene locus can act as either a recipient or a donor of activating regulatory elements in different rearrangement events. These findings reveal mechanisms by which enhancers mediate activation of MYC and BCL6 in lymphoma and provide insight into the activity of lymphoma subtype–specific enhancers in oncogenic loci.
Loss of p15 in BRAF-Mutant Nevi Promotes the Transition to Melanoma
See article, p. 1072.
p15 is highly upregulated in benign nevus melanocytes but is reduced in malignant melanoma.
BRAFV600E-induced TGFβ leads to p15 upregulation and p15-mediated growth arrest in nevi.
Loss of p15 in melanocytic nevi overrides BRAFV600E-driven arrest to promote melanoma progression.
The BRAFV600E mutation is a common initiating event in benign melanocytic nevi, but the mechanisms that restrain BRAF signaling to prevent progression to melanoma are unknown. The cyclin-dependent kinase (CDK) inhibitor gene clusters CDKN2A and CDKN2B, which encode p16INK4A/p14ARF and p15INK4B, respectively, are often co-deleted in melanoma, but the role of p15 in tumorigenesis remains unclear. McNeal and colleagues found that BRAFV600E–expressing benign human nevus melanocytes were growth arrested and had elevated p15 expression compared with normal melanocytes; however, p15 was downregulated in primary melanoma samples compared with adjacent nevi. Upregulation of p15 in BRAFV600E-expressing nevi resulted from induction of TGFβ downstream of the MAPK pathway. Expression of p15 in primary normal melanocytes halted proliferation, whereas the effects of p16 overexpression were modest. Furthermore, p15 knockdown or CDKN2B deletion in BRAFV600E-expressing melanocytes partially reversed BRAFV600E-induced growth arrest, an effect that was further augmented by combined loss of CDKN2A/B. To determine the functional relevance of p15 in tissue, the authors developed an in vivo three-dimensional human tissue xenograft model using BRAFV600E-positive nevus melanocytes, normal keratinocytes, and native human dermis. In this setting, p15 loss, in combination with other common melanoma-associated genetic events, was sufficient to reverse the growth arrest of BRAFV600E-expressing nevus melanocytes and promote melanoma progression. These results indicate that BRAFV600E induces growth arrest in nevus melanocytes through TGFβ-dependent upregulation of p15, the loss of which is often critical for the transition from benign nevi to malignant melanoma.
Lineage Tracing Reveals Polyclonal Seeding Patterns in PDAC Metastasis
See article, p. 1086.
Premalignant progression of PDAC is associated with a decrease in clonal diversity.
PDAC metastases frequently arise due to polyclonal seeding by multiple tumor subclones.
Monoclonal or polyclonal outgrowth of metastatic lesions differs according to the metastatic site.
The prognosis for patients with pancreatic ductal adenocarcinoma (PDAC) is poor, due in part to the high frequency of metastasis observed at diagnosis. Although sequencing efforts have highlighted clonal heterogeneity in PDAC, metastatic seeding patterns and clonal evolution during metastasis remain relatively unknown. In order to study metastatic clonality in vivo, Maddipati and Stanger used multiplexed labeling to track differentially colored tumor subpopulations in an autochthonous mouse model of PDAC driven by expression of oncogenic Kras and heterozygous loss of Trp53. In contrast to precursor lesions that consisted of different-colored cells, premalignant lesions were primarily characterized as monochromatic, suggesting that tumor progression provides a selective pressure that decreases tumor heterogeneity. Analysis of metastases to the peritoneal wall and diaphragm revealed multicolored polyclonal lesions, which were associated with the presence of bichromatic cellular aggregates in ascites fluid. In vivo cell-mixing experiments showed that injection of multicolored tumor cell clusters drove robust formation of polychromatic metastatic lesions in the diaphragm and lung, providing further evidence that PDAC metastases often arise from polyclonal seeding. However, although polychromatic micrometastases were observed in the liver and lung, large metastases in these tissues were mainly monochromatic, suggestive of selective clonal outgrowth at certain metastatic sites. Together, these data suggest that heterotypic interactions between tumor subclones drive polyclonal seeding and metastatic site–dependent cellular outgrowth that contribute to PDAC progression.
Regulatory T-cell Responses to Inflammation Promote Tumorigenesis
See article, p. 1098.
Tregs and IL17-producing cells are increased after ETBF colonization prior to tumorigenesis.
Initial formation of ETBF-driven colorectal tumors is Treg-dependent.
Tregs promote protumorigenic Th17 differentiation and inhibit Th1 responses via consumption of IL2.
Chronic intestinal inflammation induces an IL17 immune response that drives the development of colitis and colorectal cancer. Intestinal microbiota, such as the bacterium enterotoxigenic Bacteroides fragilis (ETBF), modulate intestinal immune responses and have been implicated in IL17-driven colon tumorigenesis, but the mechanisms underlying the regulation of the procarcinogenic IL17 response have not yet been elucidated. Regulatory T cells (Treg) limit excessive inflammatory responses to maintain intestinal immune homeostasis, prompting Geis and colleagues to investigate the role of Tregs in ETBF-driven colorectal tumorigenesis using ETBF-colonized ApcMin/+ (ApcMin) mice. An increase in the Treg cell population occurred in the distal colon early after ETBF colonization of ApcMin mice and preceded tumorigenesis. Treg depletion resulted in increased colonic inflammation, but unexpectedly reduced early microadenoma formation, which was accompanied by a decrease in Th17 cells and IL17 production and an increase in the IFNγ-producing Th1 cell population. This reduction in neoplastic growth was not due to enhanced IFNγ-mediated antitumor immune responses in the absence of Tregs, but was instead dependent on cell-extrinsic regulation of Th17 differentiation by Tregs. Treg consumption of IL2, which increased IL17, resulted in an increase in Th17 polarization and a subsequent decrease in the Th1 population following ETBF colonization. Taken together, these results show that Tregs drive Th17 polarization to establish protumorigenic colitis and are necessary for the early stages of tumorigenesis in colitis-associated colorectal cancer.
Note: In This Issue is written by Cancer Discovery editorial staff. Readers are encouraged to consult the original articles for full details.