Metastatic progression drives the majority of cancer-related fatalities, and involvement of the central nervous system (CNS) poses especially formidable challenges to patients and clinicians. Brain metastases (BrM), commonly originate from lung, breast and melanoma cancers, and carry disproportionately poor outcomes. Although therapeutic advances have extended survival for many extracranial tumors, BrM incidence continues to climb-underscoring critical knowledge gaps in understanding the unique biology of tumor colonization in the CNS. While definitive evidence remains limited, a growing focus on cancer neuroscience-especially regarding hormone dependent cancer cells in the brain-has begun to reveal that factors normally regulated by sex steroids and neurosteroids may similarly influence the specialized metastatic microenvironment in the CNS. Steroid hormones can permeate the blood-brain barrier (BBB) or be synthesized de novo by astrocytes and other CNS-resident cells, potentially influencing processes such as inflammation, synaptic plasticity, and immune surveillance. However, how these hormonal pathways are co-opted by disseminated cancer cells remains unclear. Here, we review the complex hormonal landscape of the adult brain and examine how neuroendocrine-immune interactions, often regulated by sex hormones, may support metastatic growth. We discuss the interplay between systemic hormones, local steroidogenesis, and tumor adaptation to identify novel therapeutic opportunities.

Small cell lung cancer (SCLC) is known for its high metastatic potential, with most patients demonstrating clinically evident metastases in multiple organs at diagnosis. The factors contributing to this exceptional metastatic capacity have not been defined. To bridge this gap, we compare gene expression in SCLC patient samples who never experienced metastasis or relapse throughout their clinical course, versus primary SCLC patient samples from more typical patients who had metastatic disease at diagnosis. This analysis identifies FOXA2 as a transcription factor strongly associated with SCLC metastasis. Subsequent analyses in experimental models demonstrates that FOXA2 induces a fetal neuroendocrine gene expression program and promotes multi-site metastasis. Moreover, we identify ASCL1, a transcription factor known for its initiating role in SCLC tumorigenesis, as a direct binder of the FOXA2 promoter and regulator of FOXA2 expression. Taken together, these data define the ASCL1-FOXA2 axis as a critical driver of multiorgan SCLC metastasis.

While primary breast cancer (BC) is often effectively managed, metastasis remains the primary cause of BC-related fatalities. Gaps remain in our understanding of the mechanisms regulating cancer cell organotropism with predilection to specific organs. Unraveling mediators of site-specific metastasis could enhance early detection and enable more tailored interventions. Liquid biopsy represents an innovative approach in cancer involving the analysis of biological materials such as circulating tumor DNA and tumor-derived extracellular vesicles (EV) found in body fluids like blood or urine. This offers valuable insights for characterizing and monitoring tumor genomes to advance personalized medicine in metastatic cancers.

We performed in-depth analyses of EV cargo associated with BC metastasis using eight murine cell line models with distinct metastatic potentials and organotropism to the lung, the bone, the liver, and the brain. We characterized the secretome of these cells to identify unique biomarkers specific to metastatic sites.

Small EVs isolated from all cell lines were quantified and validated for established EV markers. Tracking analysis and electron microscopy revealed EV secretion patterns that differed according to cell line. Cell-free (cf)DNA and EV-associated DNA (EV-DNA) were detected from all cell lines with varying concentrations. We detected a TP53 mutation in both EV-DNA and cfDNA. Mass spectrometry-based proteomics analyses identified 698 EV-associated proteins, which clustered according to metastatic site. This analysis highlighted both common EV signatures and proteins involved in cancer progression and organotropism unique to metastatic cell lines. Among these, 327 significantly differentially enriched proteins were quantified with high confidence levels across BC and metastatic BC cells. We found enrichment of specific integrin receptors in metastatic cancer EVs compared to EVs secreted from non-transformed epithelial cells and matched tumorigenic non-metastatic cells. Pathway analyses revealed that EVs derived from parental cancer cells display a cell adhesion signature and are enriched with proteins involved in cancer signaling pathways.

Taken together, the characterization of EV cargo in a unique model of BC organotropism demonstrated that EV-DNA and EV proteomes were informative of normal and cancer states. This work could help to identify BC biomarkers associated with site-specific metastasis and new therapeutic targets.

Bone metastasis is a major cause of cancer death; however, the epigenetic determinants driving this process remain elusive. Here, we report that histone methyltransferase ASH1L is genetically amplified and is required for bone metastasis in men with prostate cancer. ASH1L rewires histone methylations and cooperates with HIF-1α to induce pro-metastatic transcriptome in invading cancer cells, resulting in monocyte differentiation into lipid-associated macrophage (LA-TAM) and enhancing their pro-tumoral phenotype in the metastatic bone niche. We identified IGF-2 as a direct target of ASH1L/HIF-1α and mediates LA-TAMs' differentiation and phenotypic changes by reprogramming oxidative phosphorylation. Pharmacologic inhibition of the ASH1L-HIF-1α-macrophages axis elicits robust anti-metastasis responses in preclinical models. Our study demonstrates epigenetic alterations in cancer cells reprogram metabolism and features of myeloid components, facilitating metastatic outgrowth. It establishes ASH1L as an epigenetic driver priming metastasis and macrophage plasticity in the bone niche, providing a bona fide therapeutic target in metastatic malignancies.

Recurrent/metastatic (R/M) adenoid cystic carcinoma (ACC) is typically slow growing however, some patients have more rapid progression. We sought to classify mutations in chromatin regulating genes in ACC to determine the impact of chromatin regulatory disfunction on clinical outcomes.

Matched clinical-genomic data from 271 pts with non-resectable or R/M ACC were included in this study. 132 pts were recruited locally. 139 ACC pts were included from cBioPortal for analysis. Mutations were classified as pathogenic using standardised bioinformatic pipelines. Analyses was performed to determine the impact of one or more mutations on overall survival from recurrence (OSr) and site of metastases.

KDM6A mutations were seen in 12 % of patients, followed by ARID1A 9 %, EP300 6 %, CREBBP 5 %, KMT2D 5 %, SETD2 2.6 %, an KMT2C 1.8 %. While 15 % of patients harboured activating NOTCH1/2 mutations, which co-occurred with mutations in KDM6A, ARID1A, and CREBBP. NOTCH activation (8.4 v 2.3 years, p= <0.005), KDM6A (6.6 v 4.6 years, p = 0.008) and SETD2 (5.9 v 3.8 years, p = 0.04) mutations were associated with worse OSr. In NOTCH wild-type patients, median OSr for KDM6A 9.2 v 4.8 years (p = 0.07) and SETD2 was 9.8 v 3.2 years (p = 0.04). KDM6A (OR 4.5, 95 % CI 1.7-13.2, p = 0.002) mutations were significantly associated with bone metastasises and ARID1A mutations were associated with bone (OR 3.5, 95 % CI 1.2-11.8, p = 0.025) and liver (OR 3.0, 95 % CI 1.0-9.1, p = 0.053) in NOTCH wild-type patients.

ARID1A, KDM6A and SETD2 loss of function mutations negatively impact clinical outcomes in R/M-ACC irrespective of NOTCH status.

Acquired genetic alterations drive resistance to endocrine and targeted therapies in metastatic breast cancer; however, the underlying processes engendering these alterations are largely uncharacterized. To identify the underlying mutational processes, we utilized a clinically annotated cohort of 3,880 patient samples with tumor-normal sequencing. Mutational signatures associated with apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3) enzymes were prevalent and enriched in post-treatment hormone receptor-positive cancers. These signatures correlated with shorter progression-free survival on antiestrogen plus CDK4/6 inhibitor therapy in hormone receptor-positive metastatic breast cancer. Whole-genome sequencing of breast cancer models and paired primary-metastatic samples demonstrated that active APOBEC3 mutagenesis promoted therapy resistance through characteristic alterations such as RB1 loss. Evidence of APOBEC3 activity in pretreatment samples illustrated its pervasive role in breast cancer evolution. These studies reveal APOBEC3 mutagenesis to be a frequent mediator of therapy resistance in breast cancer and highlight its potential as a biomarker and target for overcoming resistance.

The dynamic regulation of epigenetic states relies on complex macromolecular interactions. PRC2, the methyltransferase complex responsible for depositing H3K27me3, interacts with distinct accessory proteins to form the mutually exclusive subcomplexes PHF1-PRC2.1, MTF2-PRC2.1, PHF19-PRC2.1, and PRC2.2. The functions of these subcomplexes are unclear and thought to be highly redundant. Here we show that PRC2 subcomplexes have distinct roles in epigenetic repression of lineage-specific genes and stem cell differentiation. Using a human pluripotent stem cell model, we engineered a comprehensive set of separation-of-function mutants to dissect the roles of individual protein-protein and DNA-protein interactions. Our results show that PRC2.1 and PRC2.2 deposit H3K27me3 locus-specifically, resulting in opposing outcomes in cardiomyocyte differentiation. We find that MTF2 stimulates PRC2.1-mediated repression in stem cells and cardiac differentiation through its interaction with DNA and H3K36me3, while PHF19 antagonizes it. Furthermore, MTF2-PRC2.1 maintains normal cardiomyocyte function. Together, these results reveal the importance and specificity of individual macromolecular interactions in Polycomb-mediated epigenetic repression in human stem cells and differentiation.

Admixed populations, including the Hispanic/Latino/a community, are underrepresented in cancer genetic/genomic studies. Leveraging the Latino Colorectal Cancer Consortium (LC3) and other existing datasets, we analyzed whole-exome sequencing data on tumor/normal pairs from 718 individuals with colorectal cancer to map somatic mutational features by ethnicity and genetic similarity. Global proportions of African, Asian, European, and Native American genetic ancestries were estimated using ADMIXTURE. Associations between these proportions and somatic mutational features were examined using logistic regression. APC, TP53, and KRAS were the top three mutated genes across all participants and in the subset of Latino individuals in LC3. In analyses examining recurrently mutated genes, tumors from patients of Latino ethnicity had fewer KRAS and PIK3CA mutations compared with tumors from non-Latino patients. Genetic ancestry overall was associated with CDC27 mutation status, and African genetic ancestry was associated with SMAD2 mutation status. In exome-wide analyses, African genetic ancestry was significantly associated with higher odds of mutation in KNCN and TMEM184B. Native American genetic ancestry was associated with a lower frequency of microsatellite instability-high tumors. The SBS11 mutational signature was associated with Native American genetic ancestry as well as Latino ethnicity. In an independent replication dataset, MSK-IMPACT, estimates of association were largely consistent in direction but nonsignificant. A meta-analysis of LC3 and MSK-IMPACT showed that African genetic ancestry was significantly associated with KRAS mutation status and MSI status. This work facilitates precision medicine initiatives by providing insights into the contribution of genetic ancestry to molecular features of colorectal tumors. Significance: Analysis of tumors from various populations can broadly characterize genomic landscapes and enhance precision medicine strategies.

Ampullary carcinoma (AC) is a rare disease with an abysmal prognosis and few treatment options. The molecular landscape and its therapeutic implications remain inadequately understood. This study aims to provide a clinical and genomic characterization of AC and explore opportunities for precision oncology.

We carried out a retrospective analysis of clinical and genomic features in patients with AC treated in our institution. Gene mutations were categorized into molecular pathways, and potentially targetable alterations were classified according to the European Society for Medical Oncology (ESMO) Scale for Clinical Actionability of Molecular Targets (ESCAT). Key molecular findings were validated in an external cohort.

We included 78 patients with a median age of 66 years; 51.6% were women, and most were treated with surgery (81.2%). Histologically, they were classified as pancreaticobiliary (58.3%), intestinal (INT, 33.3%), and mixed (8.3%). The percentages of patients diagnosed at stages I, II, III, and IV disease were 18.8%, 23.4%, 32.8%, and 25.0%, respectively. Of note, the INT subtype was enriched in transforming growth factor-β pathway alterations (25.9% versus 6.1%, P = 0.03). Potentially actionable molecular alterations were found in 52% of the patients. Importantly, KRASWT tumors were enriched in potentially targetable alterations ESCAT I-IIIA both in our cohort (37.2% versus 9.4%, P = 0.006) and external validation cohort (23.0% versus 9.3%, P = 0.01), including 25.6% ERBB2 amplification/mutation, 14.0% homologous recombination deficiency status, and 7.4% microsatellite instability status. Six patients received matched targeted therapies after progression to chemotherapy, with a response rate of 50% and two patients surviving for >1 year.

AC patients are enriched in targetable alterations, especially KRASWT tumors, and could particularly benefit from precision oncology-based approaches.

RBM10 modulates transcriptome-wide cassette exon splicing. Loss-of-function RBM10 mutations are enriched in thyroid cancers with distant metastases. Analysis of transcriptomes and genes mis-spliced by RBM10 loss showed pro-migratory and RHO/RAC signaling signatures. RBM10 loss increases cell velocity. Cytoskeletal and ECM transcripts subject to exon inclusion events included vinculin (VCL), tenascin C (TNC), and CD44. Knockdown of the VCL exon inclusion transcript in RBM10-null cells reduced cell velocity, whereas knockdown of TNC and CD44 exon inclusion isoforms reduced invasiveness. RAC1-GTP levels were increased in RBM10-null cells. Mouse HrasG12V/Rbm1OKO thyrocytes develop metastases that are reversed by RBM10 expression or by combined knockdown of VCL, CD44, and TNC inclusion isoforms. Thus, RBM10 loss generates exon inclusion in transcripts regulating ECM-cytoskeletal interactions, leading to RAC1 activation and metastatic competency. Moreover, a CRISPR-Cas9 screen for synthetic lethality with RBM10 loss identified NFκB effectors as central to viability, providing a therapeutic target for these lethal thyroid cancers.

Small cell lung cancer (SCLC) is a highly malignant lung cancer subtype with a dismal prognosis and limited treatment options. This study aimed to identify new prognostic molecular biomarkers for SCLC and explore their immune-related implications for treatment strategies. We analyzed 200 SCLC samples via whole-exome sequencing (WES) and 313 samples by targeted sequencing. A smoking-related SBS4 mutational signature was linked to poorer prognosis and lower tumor mutational burden (TMB), while the APOBEC-mediated SBS13 signature was associated with better prognosis and higher TMB. We identified a molecular subtype with the worst outcomes and lowest TMB in both cohorts. Among 38 high-frequency mutated genes associated with SCLC prognosis, only UNC13A mutations were beneficial. Patients with UNC13A mutations had favorable immune infiltration and tumor immunogenicity. Additionally, TP53 splice site mutations were related to the worst survival outcomes. In conclusion, we discovered new molecular biomarkers for SCLC prognosis. Our findings on their immunological characteristics offer insights for developing novel SCLC treatment strategies.

To investigate whether genetic analysis of focal barbotage samples obtained at ureterorenoscopy (URS) is possible, and to identify genetic aberrations that might add prognostic information.

This prospective study included barbotage samples from 42 patients with upper urinary tract urothelial carcinoma (UTUC) confirmed at URS. At URS, focal barbotage specimens were collected for cytology and for gene sequencing. Tumour grades were determined from cytology and/or biopsy, or from radical nephroureterectomy samples. Next-generation sequencing using a 385-gene panel was performed and single nucleotide variants (SNVs), deletions/insertions (indels) and copy number aberrations (CNAs) were identified. Manual filtering of the SNVs/indels was performed to identify possible pathogenic mutations.

Of the 42 samples, two failed quality control, therefore, 40 focal barbotage samples were sequenced. We identified known and suspected pathogenic mutations and other genomic aberrations in 36 samples. The most common variants were in TERT (78%), FGFR3 (50%), KMT2D (42%), KDM6A (42%), ARID1A (39%), TP53 (19%) and deletion of 9q (50%). Known pathogenic mutations in FGFR3 were common in grade 1 and 2 tumours, but not present in any grade 3 tumour. No patients with an FGFR3 mutation died during follow-up. TP53 variants or deletions, as well as amplifications of MDM2, were only present in high-grade (HG) tumours or low-grade (LG) tumours in patients who had metastasis/died from urinary tract carcinoma. CNAs were detected in 36/40 barbotage samples, 91% of the HG samples and 69% of the LG samples, including those from all five patients with LG tumours with metastasis or who died from urinary tract cancer.

Focal barbotage samples enable identification of gene mutations and other genetic aberrations that may add important prognostic information to histopathology and cytology. Refined prognostication of UTUC patients already at diagnosis can guide treatment decisions and follow-up programmes.

Genomic profiling of central nervous system (CNS) metastases has the potential to guide treatments. In the present study, we included 584 patients with non-small-cell lung cancer and CNS metastases and performed a comprehensive analysis of cerebrospinal fluid (CSF) circulating tumor DNA (ctDNA) with clinicopathological annotation. CSF ctDNA-positive detection was independently associated with shorter survival than negative detection (hazard ratio (HR) = 1.9, 95% confidence interval (CI) = 1.56-2.39; P < 0.0001). Matched tumor-CSF analysis characterized the CSF private molecular features causing poor survival (HR = 1.64, 95% CI = 1.15-2.32, P = 0.006). A multimetric CSF ctDNA prognostic model integrating CSF ctDNA features and clinical factors was developed for risk-stratifying CNS metastases and validated in an independent cohort. Among patients with treatment histories available, those positive for a driver alteration by CSF ctDNA showed a survival benefit from CSF-matched therapy (HR = 0.78, 95% CI = 0.65-0.92, P = 0.003). Longitudinal monitoring by CSF identified CNS-specific resistant mechanisms and a second matched targeted therapy indicating improved survival (HR = 0.56, 95% CI = 0.35-0.91, P = 0.018). These findings support the clinical value of CSF ctDNA for risk-stratifying CNS metastases and guiding therapy.

Regulatory T cells (Tregs) are critical for maintaining the stability of the immune system and facilitating tumor escape through various mechanisms. Resting T cells are involved in cell-mediated immunity and remain in a resting state until stimulated, while effector T cells promote immune responses. Here, we investigated the roles of two gene signatures, one for resting Tregs (FOXP3 and IL2RA) and another for effector Tregs (FOXP3, CTLA-4, CCR8 and TNFRSF9) in pan-cancer. Using data from The Cancer Genome Atlas (TCGA), The Cancer Proteome Atlas (TCPA) and Gene Expression Omnibus (GEO), we focused on the expression profile of the two signatures, the existence of single nucleotide variants (SNVs) and copy number variants (CNVs), methylation, infiltration of immune cells in the tumor and sensitivity to different drugs. Our analysis revealed that both signatures are differentially expressed across different cancer types, and correlate with patient survival. Furthermore, both types of Tregs influence important pathways in cancer development and progression, like apoptosis, epithelial-to-mesenchymal transition (EMT) and the DNA damage pathway. Moreover, a positive correlation was highlighted between the expression of gene markers in both resting and effector Tregs and immune cell infiltration in adrenocortical carcinoma, while mutations in both signatures correlated with enrichment of specific immune cells, mainly in skin melanoma and endometrial cancer. In addition, we reveal the existence of widespread CNVs and hypomethylation affecting both Treg signatures in most cancer types. Last, we identified a few correlations between the expression of CCR8 and TNFRSF9 and sensitivity to several drugs, including COL-3, Chlorambucil and GSK1070916, in pan-cancer. Overall, these findings highlight new evidence that both Treg signatures are crucial regulators of cancer progression, providing potential clinical outcomes for cancer therapy.

Clonal hematopoiesis of indeterminate potential (CHIP) is an age-related condition associated with increased mortality among patients with cancer. CHIP mutations with high variant-allele frequencies can be detected in tumors, a phenomenon we term tumor-infiltrating clonal hematopoiesis (TI-CH). The frequency of TI-CH and its effect on tumor evolution are unclear.

We characterized CHIP and TI-CH in 421 patients with early-stage non-small-cell lung cancer (NSCLC) from the TRACERx study and in 49,351 patients from the MSK-IMPACT pan-cancer cohort. We studied the association of TI-CH with survival and disease recurrence and evaluated the functional effect of TET2-mutant CHIP on the biologic features of lung tumors.

Among patients with NSCLC, 42% of those with CHIP had TI-CH. TI-CH independently predicted an increased risk of death or recurrence, with an adjusted hazard ratio of 1.80 (95% confidence interval [CI], 1.23 to 2.63) as compared with the absence of CHIP and an adjusted hazard ratio of 1.62 (95% CI, 1.02 to 2.56) as compared with CHIP in the absence of TI-CH. Among patients with solid tumors, 26% of those with CHIP had TI-CH. TI-CH conferred a risk of death from any cause that was 1.17 times (95% CI, 1.06 to 1.29) as high as the risk with CHIP in the absence of TI-CH. TET2 mutations were the strongest genetic predictor of TI-CH; such mutations enhanced monocyte migration to lung tumor cells, fueled a myeloid-rich tumor microenvironment in mice, and resulted in the promotion of tumor organoid growth.

TI-CH increased the risk of disease recurrence or death among patients with NSCLC and the risk of death from any cause among patients with solid tumors. TI-CH remodeled the tumor immune microenvironment and accelerated tumor organoid growth, findings that support a role for an aging-related hematologic clonal proliferation in cancer evolution. (Funded by the Royal Society and others.).

The evolution of metastasis in humans is considerably less well understood than the biology of early carcinogenesis. For over a century, clinicians and scientists have been debating whether metastatic potential is the intrinsic property of a cancer, pre-determined by the molecular characteristics of the tumour founder cell, or whether metastatic capacity evolves in a stepwise fashion as the tumour grows, akin to the multistage accumulation of oncogenic alterations that give rise to the first cancer cell. In this Perspective, I examine how genetic analyses of primary tumours and matched metastases can distinguish between these two competing metastasis evolution models, with particular emphasis on the utility of metastatic randomness - a quantitative measure that reflects whether metastases arise from a random selection of primary tumour subclones or whether they are enriched for descendants of privileged lineages that have acquired pro-metastatic traits. Probable metastasis evolution trajectories in tumours with high and low baseline metastatic capacity are discussed, along with the role of seeding rates and selection at different metastatic host sites. Finally, I argue that trailblazing insights into human metastasis biology are immediately possible if we make a concerted effort to apply existing experimental and theoretical tools to the right patient cohorts.

HER2-low expression has recently gained considerable attention as an actionable biomarker in gastric cancer. However, changes in HER2-low expression between primary and metastatic gastric cancers remain inadequately explored. This study included consecutive patients diagnosed with metastatic gastric cancer with both primary and metastatic tumors, between January 2014 and December 2023. HER2 status was evaluated in both primary and matched metastatic tumors. A total of 332 patients were enrolled, with HER2-negative, HER2-low, and HER2-positive statuses were observed in 226, 81, and 25 primary tumors, respectively, and in 175, 104, and 53 metastatic tumors, respectively. Among the 226 patients with HER2-negative primary tumors, 74 and 23 developed HER2-low and HER2-positive metastatic tumors, respectively. Conversion from HER2-negative primary to HER2-low metastatic gastric cancer was associated with metachronous and non-peritoneal metastasis. Overall, re-biopsy to evaluate HER2 status may be necessary, potentially broadening the patient population eligible for targeted HER2 therapy.

Colorectal carcinoma (CRC) exhibits metastatic organotropism, primarily targeting liver, lung, and rarely the brain. Here, we study chromosomal imbalances (CIs) in cohorts of primary CRCs and metastases. Brain metastases show the highest burden of CIs, including aneuploidies and focal CIs, with enrichment of +12p encoding KRAS. Compared to liver and lung metastases, brain metastases present with increased co-occurrence of KRAS mutation and amplification. CRCs with concurrent KRAS mutation and amplification display significant metabolic reprogramming with upregulation of glycolysis, alongside upregulation of cell cycle pathways, including copy number gains of MDM2 and CDK4. Evolutionary modeling suggests early acquisition of many organotropic CIs enriched in both liver and brain metastases, while brain-enriched CIs preferentially emerge later. Collectively, this study supports a model where cytogenetic events in CRCs favor site-specific metastatic colonization. These site-enriched CI patterns may serve as biomarkers for metastatic potential in precision oncology.

Given the urgent need for alternative strategies to block metastasis progression, we demonstrate that blocking HER2-mediated secondary metastasis improves clinical outcome and establish HER2 as a biomarker for bone metastasis in patients with initial HR+/HER2- breast cancer, which represents ∼70% of all cases.

Brain metastases frequently develop in patients with non-small cell lung cancer (NSCLC) and are a common cause of cancer-related deaths, yet our understanding of the underlying human biology is limited. Here we performed multimodal single-nucleus RNA and T cell receptor, single-cell spatial and whole-genome sequencing of brain metastases and primary tumors of patients with treatment-naive NSCLC. Chromosomal instability (CIN) is a distinguishing genomic feature of brain metastases compared with primary tumors, which we validated through integrated analysis of molecular profiling and clinical data in 4,869 independent patients, and a new cohort of 12,275 patients with NSCLC. Unbiased analyses revealed transcriptional neural-like programs that strongly enriched in cancer cells from brain metastases, including a recurring, CINhigh cell subpopulation that preexists in primary tumors but strongly enriched in brain metastases, which was also recovered in matched single-cell spatial transcriptomics. Using multiplexed immunofluorescence in an independent cohort of treatment-naive pairs of primary tumors and brain metastases from the same patients with NSCLC, we validated genomic and tumor-microenvironmental findings and identified a cancer cell population characterized by neural features strongly enriched in brain metastases. This comprehensive analysis provides insights into human NSCLC brain metastasis biology and serves as an important resource for additional discovery.

The prognostic value of molecular residual disease (MRD) in non-small cell lung cancer (NSCLC) is well established, with treatment-guiding results anticipated. Here, we present updated analyses from our previously published cohort study of 261 patients with NSCLC undergoing complete resection.

A total of 261 patients with stage I to III lung cancer who underwent radical surgery were enrolled. Enrolled patients underwent follow-up blood draws according to the predefined time points after surgery. As of December 31, 2023, with a median follow-up of 43.4 months, 948 postoperative blood samples were collected.

Landmark and longitudinal MRD exhibited positive predictive values of 91.3% and 92.8%, respectively, with a median lead time of 5.2 months. Negative predictive values were 76.5% and 93.2%, respectively. Patients with landmark undetectable MRD could not benefit from adjuvant therapy through the updated follow-up (P = 0.529). Among the 13 patients with recurrent NSCLC and longitudinal undetectable MRD, seven (53.8%) had brain-only metastases, and four (30.8%) had no updated blood samples for over 6 months prior to recurrence. Besides, for those with longitudinal detectable MRD, higher maximum variant allele frequency (>0.55%) and ctDNA level (>13 hGE/mL) were associated with a high risk of short-term recurrence. Additionally, updated follow-up data further support that the peak time for detectable MRD was 18 months after landmark detection.

These findings suggest the significant potential of MRD in guiding personalized treatment for NSCLC. Postoperative longitudinal undetectable MRD can indicate a cured population.

Mismatch repair deficiency leads to high mutation rates and microsatellite instability (MSI-H), associated with immune infiltration and responsiveness to immunotherapies. In early stages, MSI-H tumors generally have a better prognosis and lower metastatic potential than microsatellite-stable (MSS) tumors, especially in colorectal cancer. However, in advanced stages, MSI-H tumors lose this survival advantage for reasons that remain unclear. We developed a syngeneic mouse model of MSI cancer by knocking out the MMR gene Msh2 in the metastatic 4T1 breast cancer cell line. This model mirrored genomic features of MSI-H cancers and showed reduction in metastatic incidence compared to their MSS counterparts. In MSI-H tumors, we observed an enrichment of immune gene-signatures that negatively correlated with metastasis incidence. A hybrid epithelial-mesenchymal signature, related to aggressiveness was detected only in metastatic MSI-H tumors. Interestingly, we identified immature myeloid cells at primary and metastatic sites in MSI-H tumor-bearing mice, suggesting that MMR deficiency elicits specific immune responses beyond T-cell activation.

Ewing sarcoma (EwS) is a rare and aggressive malignancy, which frequently affects children. One of the few recurrent genomic variants in EwS is genomic copy number deletion of CDKN2A; however, the clinical consequences of dysregulation of CDKN2A in EwS are unclear. In this study, we revisit CDKN2A to investigate its role as a potential prognostic biomarker in EwS using data from EwS pre-clinical models as well as clinical samples from patients with EwS. We demonstrate the potential essentiality of CDKN2A dysregulation and sustained downstream CDK4/CCND1 activity. Finally, we present evidence that high expression of CDKN2A is a negative prognostic biomarker at diagnosis in EwS in three independent datasets. Our data may suggest that the role of CDKN2A may change across the clinical context of EwS, however, further study is necessary to validate the function of CDKN2A expression in EwS.

Cutaneous squamous cell carcinoma (cSCC) harbors one of the most mutated genomes. There are limited data on the genomic profile and its predictive potential for response to immunotherapy with PD-1 inhibitors in cSCC.

This study retrospectively reviewed cSCC patients treated with PD-1 inhibitor monotherapy at a single institution. Clinical characteristics, treatment outcomes, PD-L1 expression, tumor mutation burden (TMB), and genomic profile in tumor and blood were analyzed. Logistic regression and a support vector classifier were used to validate identified biomarkers of significance.

Twenty-five patients were evaluable for response and had genomics tested in tumor and/or blood. Of the total, 80% of patients achieved an objective response: 40% complete response (CR), 32% partial response (PR) for more than 6 months, and 8% stable disease (SD) for more than 1 year; 20% of patients progressed on treatment. With a median follow-up of 21 months, progression-free survival (PFS) was 28 months in responders vs. 3 months in non-responders (p = 0.00001). Median PD-L1 was 25% in responders vs. 10% in non-responders (p = 0.39). There was no difference in median TMB between responders and non-responders. Eight gene mutations were significantly more frequent in non-responders than in responders: CDK12 (p = 0.005), CTCF (p = 0.033), CTNNB1 (p = 0.033), IGF1R (p = 0.038), IKBKE (p = 0.016), MLH1 (0.033), QKI (p = 0.016), and TIPARP (p = 0.033). A support vector model of these genes classified responders and non-responders with an accuracy of 0.88 in the training data and 1.0 in the testing data.

PD-1 inhibitor monotherapy produces an impressive response. Eight gene mutations were significantly more frequent in non-responders. PD-L1 and TMB were inconclusive in predicting treatment response to anti-PD-L1.