Chromosomal instability is a hallmark of cancer that results from ongoing errors in chromosome segregation during mitosis. Although chromosomal instability is a major driver of tumour evolution, its ...role in metastasis has not been established. Here we show that chromosomal instability promotes metastasis by sustaining a tumour cell-autonomous response to cytosolic DNA. Errors in chromosome segregation create a preponderance of micronuclei whose rupture spills genomic DNA into the cytosol. This leads to the activation of the cGAS-STING (cyclic GMP-AMP synthase-stimulator of interferon genes) cytosolic DNA-sensing pathway and downstream noncanonical NF-κB signalling. Genetic suppression of chromosomal instability markedly delays metastasis even in highly aneuploid tumour models, whereas continuous chromosome segregation errors promote cellular invasion and metastasis in a STING-dependent manner. By subverting lethal epithelial responses to cytosolic DNA, chromosomally unstable tumour cells co-opt chronic activation of innate immune pathways to spread to distant organs.
Aneuploidy, chromosomal instability, somatic copy-number alterations, and whole-genome doubling (WGD) play key roles in cancer evolution and provide information for the complex task of phylogenetic ...inference. We present MEDICC2, a method for inferring evolutionary trees and WGD using haplotype-specific somatic copy-number alterations from single-cell or bulk data. MEDICC2 eschews simplifications such as the infinite sites assumption, allowing multiple mutations and parallel evolution, and does not treat adjacent loci as independent, allowing overlapping copy-number events. Using simulations and multiple data types from 2780 tumors, we use MEDICC2 to demonstrate accurate inference of phylogenies, clonal and subclonal WGD, and ancestral copy-number states.
The translational challenges in the field of precision oncology are in part related to the biological complexity and diversity of this disease. Technological advances in genomics have facilitated ...large sequencing efforts and discoveries that have further supported this notion. In this review, we reflect on the impact of these discoveries on our understanding of several concepts: cancer initiation, cancer prevention, early detection, adjuvant therapy and minimal residual disease monitoring, cancer drug resistance, and cancer evolution in metastasis. We discuss key areas of focus for improving cancer outcomes, from biological insights to clinical application, and suggest where the development of these technologies will lead us. Finally, we discuss practical challenges to the wider adoption of molecular profiling in the clinic and the need for robust translational infrastructure.
Checkpoint inhibitors (CPIs) augment adaptive immunity. Systematic pan-tumor analyses may reveal the relative importance of tumor-cell-intrinsic and microenvironmental features underpinning CPI ...sensitization. Here, we collated whole-exome and transcriptomic data for >1,000 CPI-treated patients across seven tumor types, utilizing standardized bioinformatics workflows and clinical outcome criteria to validate multivariable predictors of CPI sensitization. Clonal tumor mutation burden (TMB) was the strongest predictor of CPI response, followed by total TMB and CXCL9 expression. Subclonal TMB, somatic copy alteration burden, and histocompatibility leukocyte antigen (HLA) evolutionary divergence failed to attain pan-cancer significance. Dinucleotide variants were identified as a source of immunogenic epitopes associated with radical amino acid substitutions and enhanced peptide hydrophobicity/immunogenicity. Copy-number analysis revealed two additional determinants of CPI outcome supported by prior functional evidence: 9q34 (TRAF2) loss associated with response and CCND1 amplification associated with resistance. Finally, single-cell RNA sequencing (RNA-seq) of clonal neoantigen-reactive CD8 tumor-infiltrating lymphocytes (TILs), combined with bulk RNA-seq analysis of CPI-responding tumors, identified CCR5 and CXCL13 as T-cell-intrinsic markers of CPI sensitivity.
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•Large-scale meta-analysis of >1,000 CPI-treated cases with exome/transcriptome data•Clonal TMB and CXCL9/CXCL13 expression are the strongest predictors of CPI response•A multivariable predictor of CPI response significantly outperforms TMB•9q34 loss and CCND1 amplification are additional determinants of CPI response
A whole-exome and transcriptome meta-analysis of over 1,000 patients treated with immune checkpoint blockade across seven tumor types highlights the potential of multivariable prediction models that consider both tumor- and T-cell-intrinsic mechanisms of response.
The axon initial segment (AIS) is a specialized neuronal subcompartment located at the beginning of the axon that is crucially involved in both the generation of action potentials and the regulation ...of neuronal polarity. We recently showed that prolonged neuronal depolarization produces a distal shift of the entire AIS structure away from the cell body, a change associated with a decrease in neuronal excitability. Here, we used dissociated rat hippocampal cultures, with a major focus on the dentate granule cell (DGC) population, to explore the signaling pathways underlying activity-dependent relocation of the AIS. First, a pharmacological screen of voltage-gated calcium channels (VGCCs) showed that AIS relocation is triggered by activation of L-type Cav1 VGCCs with negligible contribution from any other VGCC subtypes. Additional pharmacological analysis revealed that downstream signaling events are mediated by the calcium-sensitive phosphatase calcineurin; inhibition of calcineurin with either FK506 or cyclosporin A totally abolished both depolarization- and optogenetically-induced activity-dependent AIS relocation. Furthermore, calcineurin activation is sufficient for AIS plasticity, because expression of a constitutively active form of the phosphatase resulted in relocation of the AIS of DGCs without a depolarizing stimulus. Finally, we assessed the role of calcineurin in other forms of depolarization-induced plasticity. Neither membrane resistance changes nor spine density changes were affected by FK506 treatment, suggesting that calcineurin acts via a separate pathway to modulate AIS plasticity. Together, these results emphasize calcineurin as a vital player in the regulation of intrinsic plasticity as governed by the AIS.
As tumors grow, they acquire mutations, some of which create neoantigens that influence the response of patients to immune checkpoint inhibitors. We explored the impact of neoantigen intratumor ...heterogeneity (ITH) on antitumor immunity. Through integrated analysis of ITH and neoantigen burden, we demonstrate a relationship between clonal neoantigen burden and overall survival in primary lung adenocarcinomas. CD8⁺ tumor-infiltrating lymphocytes reactive to clonal neoantigens were identified in early-stage non–small cell lung cancer and expressed high levels of PD-1. Sensitivity to PD-1 and CTLA-4 blockade in patients with advanced NSCLC and melanoma was enhanced in tumors enriched for clonal neoantigens. T cells recognizing clonal neoantigens were detectable in patients with durable clinical benefit. Cytotoxic chemotherapy–induced subclonal neoantigens, contributing to an increased mutational load, were enriched in certain poor responders. These data suggest that neoantigen heterogeneity may influence immune surveillance and support therapeutic developments targeting clonal neoantigens.
Most computational methods that infer somatic copy number alterations (SCNAs) from bulk sequencing of DNA analyse tumour samples individually. However, the sequencing of multiple tumour samples from ...a patient's disease is an increasingly common practice. We introduce Refphase, an algorithm that leverages this multi-sampling approach to infer haplotype-specific copy numbers through multi-sample phasing. We demonstrate Refphase's ability to infer haplotype-specific SCNAs and characterise their intra-tumour heterogeneity, to uncover previously undetected allelic imbalance in low purity samples, and to identify parallel evolution in the context of whole genome doubling in a pan-cancer cohort of 336 samples from 99 tumours.
Pre‐clinical non‐small cell lung cancer (NSCLC) models are poorly representative of the considerable inter‐ and intra‐tumor heterogeneity of the disease in patients. Primary cell‐based in vitro ...models of NSCLC are therefore desirable for novel therapy development and personalized cancer medicine. Methods have been described to generate rapidly proliferating epithelial cell cultures from multiple human epithelia using 3T3‐J2 feeder cell culture in the presence of Y‐27632, a RHO‐associated protein kinase (ROCK) inhibitor, in what are known as “conditional reprograming conditions” (CRC) or 3T3 + Y. In some cancer studies, variations of this methodology have allowed primary tumor cell expansion across a number of cancer types but other studies have demonstrated the preferential expansion of normal epithelial cells from tumors in such conditions. Here, we report our experience regarding the derivation of primary NSCLC cell cultures from 12 lung adenocarcinoma patients enrolled in the Tracking Cancer Evolution through Therapy (TRACERx) clinical study and discuss these in the context of improving the success rate for in vitro cultivation of cells from NSCLC tumors.
What's new?
Traditionally, the primary culture of human cancer cells has been challenging. A new protocol known as 3T3 + Y has allowed cancer cell cultures to be established across multiple cancer types, but reports are inconsistent for non‐small cell lung cancer (NSCLC). Here, the authors sought to expand tumor cells from surgically resected primary human lung adenocarcinoma tumors within the TRACERx clinical study using 3T3 + Y. Contaminating normal epithelial cells were the predominant cell type in early passage cultures, although KRAS‐mutant tumor cells were cultured in one donor culture. The researchers urge caution in the use of NSCLC primary cell cultures and call for further investigation in 3T3 + Y applications.
Esophageal adenocarcinomas are associated with a dismal prognosis. Deciphering the evolutionary history of this disease may shed light on therapeutically tractable targets and reveal dynamic ...mutational processes during the disease course and following neoadjuvant chemotherapy (NAC). We exome sequenced 40 tumor regions from 8 patients with operable esophageal adenocarcinomas, before and after platinum-containing NAC. This revealed the evolutionary genomic landscape of esophageal adenocarcinomas with the presence of heterogeneous driver mutations, parallel evolution, early genome-doubling events, and an association between high intratumor heterogeneity and poor response to NAC. Multiregion sequencing demonstrated a significant reduction in thymine to guanine mutations within a CpTpT context when comparing early and late mutational processes and the presence of a platinum signature with enrichment of cytosine to adenine mutations within a CpC context following NAC. Esophageal adenocarcinomas are characterized by early chromosomal instability leading to amplifications containing targetable oncogenes persisting through chemotherapy, providing a rationale for future therapeutic approaches.
This work illustrates dynamic mutational processes occurring during esophageal adenocarcinoma evolution and following selective pressures of platinum exposure, emphasizing the iatrogenic impact of therapy on cancer evolution. Identification of amplifications encoding targetable oncogenes maintained through NAC suggests the presence of stable vulnerabilities, unimpeded by cytotoxics, suitable for therapeutic intervention.
Chromosomal instability is a major challenge to patient stratification and targeted drug development for high-grade serous ovarian carcinoma (HGSOC). Here we show that somatic copy number alterations ...(SCNAs) in frequently amplified HGSOC cancer genes significantly correlate with gene expression and methylation status. We identify five prevalent clonal driver SCNAs (chromosomal amplifications encompassing MYC, PIK3CA, CCNE1, KRAS and TERT) from multi-regional HGSOC data and reason that their strong selection should prioritise them as key biomarkers for targeted therapies. We use primary HGSOC spheroid models to test interactions between in vitro targeted therapy and SCNAs. MYC chromosomal copy number is associated with in-vitro and clinical response to paclitaxel and in-vitro response to mTORC1/2 inhibition. Activation of the mTOR survival pathway in the context of MYC-amplified HGSOC is statistically associated with increased prevalence of SCNAs in genes from the PI3K pathway. Co-occurrence of amplifications in MYC and genes from the PI3K pathway is independently observed in squamous lung cancer and triple negative breast cancer. In this work, we show that identifying co-occurrence of clonal driver SCNA genes could be used to tailor therapeutics for precision medicine.
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