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Purpose: Classic Hodgkin lymphoma (cHL) is a B-cell lymphoma that occurs primarily in young adults and, less frequently, in elderly individuals. A hallmark of cHL is the exceptional scarcity (1%-5%) of the malignant Hodgkin Reed-Sternberg (HRS) cells within a network of nonmalignant immune cells. Molecular determinants governing the relationship between HRS cells and their proximal microenvironment remain largely unknown., Experimental Design: We performed spatially resolved multiplexed protein imaging and transcriptomic sequencing to characterize HRS cell states, cellular neighborhoods, and gene expression signatures of 23.6 million cells from 36 newly diagnosed Epstein-Barr virus (EBV)-positive and EBV-negative cHL tumors., Results: We show that MHC-I expression on HRS cells is associated with immune-inflamed neighborhoods containing CD8+ T cells, MHC-II+ macrophages, and immune checkpoint expression (i.e., PD1 and VISTA). We identified spatial clustering of HRS cells, consistent with the syncytial variant of cHL, and its association with T-cell-excluded neighborhoods in a subset of EBV-negative tumors. Finally, a subset of both EBV-positive and EBV-negative tumors contained regulatory T-cell-high neighborhoods harboring HRS cells with augmented proliferative capacity., Conclusions: Our study links HRS cell properties with distinct immunophenotypes and potential immune escape mechanisms in cHL., (©2024 The Authors; Published by the American Association for Cancer Research.)

Somatic genetic heterogeneity resulting from post-zygotic DNA mutations is widespread in human tissues and can cause diseases, however few studies have investigated its role in neurodegenerative processes such as Alzheimer's Disease (AD). Here we report the selective enrichment of microglia clones carrying pathogenic variants, that are not present in neuronal, glia/stromal cells, or blood, from patients with AD in comparison to age-matched controls. Notably, microglia-specific AD-associated variants preferentially target the MAPK pathway, including recurrent CBL ring-domain mutations. These variants activate ERK and drive a microglia transcriptional program characterized by a strong neuro-inflammatory response, both in vitro and in patients. Although the natural history of AD-associated microglial clones is difficult to establish in human, microglial expression of a MAPK pathway activating variant was previously shown to cause neurodegeneration in mice, suggesting that AD-associated neuroinflammatory microglial clones may contribute to the neurodegenerative process in patients., Competing Interests: Competing interests. FG has been a paid consultant (no equity) to Third Rock Ventures from 2018 to 2020. Sequencing costs and analysis in this study were covered in part by a SRA between Third Rock venture and MSKCC. This work led to patents PCT/US2022/037893/WO2023004054A1 ‘Methods and compositions for the treatment of alzheimer’s disease’ by MSKCC and PCT/US2018/047964 ‘Kinase mutation-associated neurodegenerative disorders by MSKCC’.

Langerhans cell Histiocytosis (LCH) and Erdheim-Chester disease (ECD) are clonal myeloid disorders, associated with MAP-Kinase activating mutations and an increased risk of neurodegeneration. Surprisingly, we found pervasive PU.1 + microglia mutant clones across the brain of LCH and ECD patients with and without neurological symptoms, associated with microgliosis, reactive astrocytosis, and neuronal loss. The disease predominated in the grey nuclei of the rhombencephalon, a topography attributable to a local proliferative advantage of mutant microglia. Presence of clinical symptoms was associated with a longer evolution of the disease and a larger size of PU.1 + clones (p= 0.0003). Genetic lineage tracing of PU.1 + clones suggest a resident macrophage lineage or a bone marrow precursor origin depending on patients. Finally, a CSF1R-inhibitor depleted mutant microglia and limited neuronal loss in mice suggesting an alternative to MAPK inhibitors. These studies characterize a progressive neurodegenerative disease, caused by clonal proliferation of inflammatory microglia (CPIM), with a decade(s)-long preclinical stage of incipient disease that represent a therapeutic window for prevention of neuronal death., Competing Interests: Conflict of Interest. FG has performed consulting for Third Rock venture in the past. Targeted Sequencing was funded in part by a grant from Third Rock venture. FG and RV are inventors in MSKCC’s United States application or PCT international application number PCT/US2018/047964 filed on 8/24/2018 (KINASE MUTATION-ASSOCIATED NEURODEGENERATIVE DISORDERS)

Neuroendocrine (NE) transformation is a mechanism of resistance to targeted therapy in lung and prostate adenocarcinomas leading to poor prognosis. Up to date, even if patients at high risk of transformation can be identified by the occurrence of Tumor Protein P53 (TP53) and Retinoblastoma Transcriptional Corepressor 1 (RB1) mutations in their tumors, no therapeutic strategies are available to prevent or delay histological transformation. Upregulation of the cell cycle kinase Cell Division Cycle 7 (CDC7) occurred in tumors during the initial steps of NE transformation, already after TP53/RB1 co-inactivation, leading to induced sensitivity to the CDC7 inhibitor simurosertib. CDC7 inhibition suppressed NE transdifferentiation and extended response to targeted therapy in in vivo models of NE transformation by inducing the proteasome-mediated degradation of the MYC Proto-Oncogen (MYC), implicated in stemness and histological transformation. Ectopic overexpression of a degradation-resistant MYC isoform reestablished the NE transformation phenotype observed on targeted therapy, even in the presence of simurosertib. CDC7 inhibition also markedly extended response to standard cytotoxics (cisplatin, irinotecan) in lung and prostate small cell carcinoma models. These results nominate CDC7 inhibition as a therapeutic strategy to constrain lineage plasticity, as well as to effectively treat NE tumors de novo or after transformation. As simurosertib clinical efficacy trials are ongoing, this concept could be readily translated for patients at risk of transformation., (© 2024. The Author(s).)

Abstract: B-cell maturation antigen (BCMA)-targeting therapeutics have dramatically improved outcomes in relapsed/refractory multiple myeloma (RRMM). However, whether the mechanisms of resistance between these therapies are shared and how the identification of such mechanisms before therapy initiation could refine clinical decision-making remains undefined. We analyzed outcomes for 72 RRMM patients treated with teclistamab, a CD3 × BCMA bispecific antibody, 42% (30/72) of whom had prior BCMA-directed therapy exposure. Malignant plasma cell BCMA expression was present in all BCMA therapy-naïve patients. Prior therapy-mediated loss of plasma cell BCMA expression before teclistamab treatment, measured by immunohistochemistry, was observed in 3 patients, none of whom responded to teclistamab, and 1 of whom also did not respond to ciltacabtagene autoleucel. Whole exome sequencing of tumor DNA from 1 patient revealed biallelic loss of TNFRSF17 following treatment with belantamab mafodotin. Low-to-undetectable peripheral blood soluble BCMA levels correlated with the absence of BCMA expression by bone marrow plasma cells. Thus, although rare, loss of BCMA expression following TNFRSF17 gene deletions can occur following any BCMA-directed therapy and prevents response to subsequent anti-BCMA-directed treatments, underscoring the importance of verifying the presence of a target antigen., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Purpose: The impact of the intratumoral microbiome on immune checkpoint inhibitor (ICI) efficacy in patients with non-small-cell lung cancer (NSCLC) is unknown. Preclinically, intratumoral Escherichia is associated with a proinflammatory tumor microenvironment and decreased metastases. We sought to determine whether intratumoral Escherichia is associated with outcome to ICI in patients with NSCLC., Patients and Methods: We examined the intratumoral microbiome in 958 patients with advanced NSCLC treated with ICI by querying unmapped next-generation sequencing reads against a bacterial genome database. Putative environmental contaminants were filtered using no-template controls (n = 2,378). The impact of intratumoral Escherichia detection on overall survival (OS) was assessed using univariable and multivariable analyses. The findings were further validated in an external independent cohort of 772 patients. Escherichia fluorescence in situ hybridization (FISH) and transcriptomic profiling were performed., Results: In the discovery cohort, read mapping to intratumoral Escherichia was associated with significantly longer OS (16 v 11 months; hazard ratio, 0.73 [95% CI, 0.59 to 0.92]; P = .0065) in patients treated with single-agent ICI, but not combination chemoimmunotherapy. The association with OS in the single-agent ICI cohort remained statistically significant in multivariable analysis adjusting for prognostic features including PD-L1 expression ( P = .023). Analysis of an external validation cohort confirmed the association with improved OS in univariable and multivariable analyses of patients treated with single-agent ICI, and not in patients treated with chemoimmunotherapy. Escherichia localization within tumor cells was supported by coregistration of FISH staining and serial hematoxylin and eosin sections. Transcriptomic analysis correlated Escherichia-positive samples with expression signatures of immune cell infiltration., Conclusion: Read mapping to potential intratumoral Escherichia was associated with survival to single-agent ICI in two independent cohorts of patients with NSCLC.

Dysregulated epigenetic states are a hallmark of cancer and often arise from genetic alterations in epigenetic regulators. This includes missense mutations in histones, which, together with associated DNA, form nucleosome core particles. However, the oncogenic mechanisms of most histone mutations are unknown. Here, we demonstrate that cancer-associated histone mutations at arginines in the histone H3 N-terminal tail disrupt repressive chromatin domains, alter gene regulation, and dysregulate differentiation. We find that histone H3R2C and R26C mutants reduce transcriptionally repressive H3K27me3. While H3K27me3 depletion in cells expressing these mutants is exclusively observed on the minor fraction of histone tails harboring the mutations, the same mutants recurrently disrupt broad H3K27me3 domains in the chromatin context, including near developmentally regulated promoters. H3K27me3 loss leads to de-repression of differentiation pathways, with concordant effects between H3R2 and H3R26 mutants despite different proximity to the PRC2 substrate, H3K27. Functionally, H3R26C-expressing mesenchymal progenitor cells and murine embryonic stem cell-derived teratomas demonstrate impaired differentiation. Collectively, these data show that cancer-associated H3 N-terminal arginine mutations reduce PRC2 activity and disrupt chromatin-dependent developmental functions, a cancer-relevant phenotype., (© 2024. The Author(s).)

Systemic Lupus Erythematosus (SLE) is an autoimmune disease, the pathophysiology and genetic basis of which are incompletely understood. Using a forward genetic screen in multiplex families with systemic lupus erythematosus (SLE) we identified an association between SLE and compound heterozygous deleterious variants in the non-receptor tyrosine kinases (NRTKs) ACK1 and BRK. Experimental blockade of ACK1 or BRK increased circulating autoantibodies in vivo in mice and exacerbated glomerular IgG deposits in an SLE mouse model. Mechanistically, non-receptor tyrosine kinases (NRTKs) regulate activation, migration, and proliferation of immune cells. We found that the patients' ACK1 and BRK variants impair efferocytosis, the MERTK-mediated anti-inflammatory response to apoptotic cells, in human induced Pluripotent Stem Cell (hiPSC)-derived macrophages, which may contribute to SLE pathogenesis. Overall, our data suggest that ACK1 and BRK deficiencies are associated with human SLE and impair efferocytosis in macrophages.

Broad-spectrum RAS inhibition has the potential to benefit roughly a quarter of human patients with cancer whose tumours are driven by RAS mutations 1,2 . RMC-7977 is a highly selective inhibitor of the active GTP-bound forms of KRAS, HRAS and NRAS, with affinity for both mutant and wild-type variants 3 . More than 90% of cases of human pancreatic ductal adenocarcinoma (PDAC) are driven by activating mutations in KRAS 4 . Here we assessed the therapeutic potential of RMC-7977 in a comprehensive range of PDAC models. We observed broad and pronounced anti-tumour activity across models following direct RAS inhibition at exposures that were well-tolerated in vivo. Pharmacological analyses revealed divergent responses to RMC-7977 in tumour versus normal tissues. Treated tumours exhibited waves of apoptosis along with sustained proliferative arrest, whereas normal tissues underwent only transient decreases in proliferation, with no evidence of apoptosis. In the autochthonous KPC mouse model, RMC-7977 treatment resulted in a profound extension of survival followed by on-treatment relapse. Analysis of relapsed tumours identified Myc copy number gain as a prevalent candidate resistance mechanism, which could be overcome by combinatorial TEAD inhibition in vitro. Together, these data establish a strong preclinical rationale for the use of broad-spectrum RAS-GTP inhibition in the setting of PDAC and identify a promising candidate combination therapeutic regimen to overcome monotherapy resistance., (© 2024. The Author(s).)

Genomic rearrangements are a hallmark of most childhood tumors, including medulloblastoma, one of the most common brain tumors in children, but their causes remain largely unknown. Here, we show that PiggyBac transposable element derived 5 (Pgbd5) promotes tumor development in multiple developmentally accurate mouse models of Sonic Hedgehog (SHH) medulloblastoma. Most Pgbd5-deficient mice do not develop tumors, while maintaining normal cerebellar development. Ectopic activation of SHH signaling is sufficient to enforce cerebellar granule cell progenitor-like cell states, which exhibit Pgbd5-dependent expression of distinct DNA repair and neurodevelopmental factors. Mouse medulloblastomas expressing Pgbd5 have increased numbers of somatic structural DNA rearrangements, some of which carry PGBD5-specific sequences at their breakpoints. Similar sequence breakpoints recurrently affect somatic DNA rearrangements of known tumor suppressors and oncogenes in medulloblastomas in 329 children. This identifies PGBD5 as a medulloblastoma mutator and provides a genetic mechanism for the generation of oncogenic DNA rearrangements in childhood cancer.

Colitis induced by treatment with immune-checkpoint inhibitors (ICI), termed irColitis, is a substantial cause of morbidity complicating cancer treatment. We hypothesized that abnormal fecal microbiome features would be present at the time of irColitis onset and that restoring the microbiome with fecal transplant from a healthy donor would mitigate disease severity. Herein, we present fecal microbiota profiles from 18 patients with irColitis from a single center, 5 of whom were treated with healthy-donor fecal microbial transplantation (FMT). Although fecal samples collected at onset of irColitis had comparable α-diversity to that of comparator groups with gastrointestinal symptoms, irColitis was characterized by fecal microbial dysbiosis. Abundances of Proteobacteria were associated with irColitis in multivariable analyses. Five patients with irColitis refractory to steroids and biologic anti-inflammatory agents received healthy-donor FMT, with initial clinical improvement in irColitis symptoms observed in four of five patients. Two subsequently exhibited recurrence of irColitis symptoms following courses of antibiotics. Both received a second "salvage" FMT that was, again, followed by clinical improvement of irColitis. In summary, we observed distinct microbial community changes that were present at the time of irColitis onset. FMT was followed by clinical improvements in several cases of steroid- and biologic-agent-refractory irColitis. Strategies to restore or prevent microbiome dysbiosis in the context of immunotherapy toxicities should be further explored in prospective clinical trials., (©2023 American Association for Cancer Research.)

Purpose: This study sought to identify β-catenin targets that regulate desmoid oncogenesis and determine whether external signaling pathways, particularly those inhibited by sorafenib (e.g., PDGFRβ), affect these targets to alter natural history or treatment response in patients., Experimental Design: In vitro experiments utilized primary desmoid cell lines to examine regulation of β-catenin targets. Relevance of results was assessed in vivo using Alliance trial A091105 correlative biopsies., Results: CTNNB1 knockdown inhibited hypoxia-regulated gene expression in vitro and reduced levels of HIF1α protein. ChIP-seq identified ABL1 as a β-catenin transcriptional target that modulated HIF1α and desmoid cell proliferation. Abrogation of either CTNNB1 or HIF1A inhibited desmoid cell-induced VEGFR2 phosphorylation and tube formation in endothelial cell co-cultures. Sorafenib inhibited this activity directly but also reduced HIF1α protein expression and c-Abl activity while inhibiting PDGFRβ signaling in desmoid cells. Conversely, c-Abl activity and desmoid cell proliferation were positively regulated by PDGF-BB. Reduction in PDGFRβ and c-Abl phosphorylation was commonly observed in biopsy samples from patients after treatment with sorafenib; markers of PDGFRβ/c-Abl pathway activation in baseline samples were associated with tumor progression in patients on the placebo arm and response to sorafenib in patients receiving treatment., Conclusions: The β-catenin transcriptional target ABL1 is necessary for proliferation and maintenance of HIF1α in desmoid cells. Regulation of c-Abl activity by PDGF signaling and targeted therapies modulates desmoid cell proliferation, thereby suggesting a reason for variable biologic behavior between tumors, a mechanism for sorafenib activity in desmoids, and markers predictive of outcome in patients., (©2023 The Authors; Published by the American Association for Cancer Research.)

T cells are the central drivers of many inflammatory diseases, but the repertoire of tissue-resident T cells at sites of pathology in human organs remains poorly understood. We examined the site-specificity of T cell receptor (TCR) repertoires across tissues (5 to 18 tissues per patient) in prospectively collected autopsies of patients with and without graft-versus-host disease (GVHD), a potentially lethal tissue-targeting complication of allogeneic hematopoietic cell transplantation, and in mouse models of GVHD. Anatomic similarity between tissues was a key determinant of TCR repertoire composition within patients, independent of disease or transplant status. The T cells recovered from peripheral blood and spleens in patients and mice captured a limited portion of the TCR repertoire detected in tissues. Whereas few T cell clones were shared across patients, motif-based clustering revealed shared repertoire signatures across patients in a tissue-specific fashion. T cells at disease sites had a tissue-resident phenotype and were of donor origin based on single-cell chimerism analysis. These data demonstrate the complex composition of T cell populations that persist in human tissues at the end stage of an inflammatory disorder after lymphocyte-directed therapy. These findings also underscore the importance of studying T cell in tissues rather than blood for tissue-based pathologies and suggest the tissue-specific nature of both the endogenous and posttransplant T cell landscape.

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., (©2023 American Association for Cancer Research.)

The shift in cancer therapy from broadly cytotoxic agents toward "personalized" treatments that target specific alterations in each patient's tumor requires diagnostic pathology approaches that are quantitative and biospecimen-friendly. Novel multiplexed antibody-based imaging technologies can measure single-cell expression of over 60 proteins in intact tumor sections and hold promise for clinical oncology., (©2023 The Authors; Published by the American Association for Cancer Research.)

Late cardiac toxicity is a potentially lethal complication of cancer therapy, yet the pathogenic mechanism remains largely unknown, and few treatment options exist. Here we report DNA-damaging agents such as radiation and anthracycline chemotherapies inducing delayed cardiac inflammation following therapy due to activation of cGAS- and STING-dependent type I interferon signaling. Genetic ablation of cGAS-STING signaling in mice inhibits DNA damage-induced cardiac inflammation, rescues late cardiac functional decline, and prevents death from cardiac events. Treatment with a STING antagonist suppresses cardiac interferon signaling following DNA-damaging therapies and effectively mitigates cardiac toxicity. These results identify a therapeutically targetable, pathogenic mechanism for one of the most vexing treatment-related toxicities in cancer survivors., (© 2022 Shamseddine et al.)

The T cell repertoire of healthy mice and humans harbors self-reactive CD4 + conventional T (T conv ) cells capable of inducing autoimmunity. Using T cell receptor profiling paired with in vivo clonal analysis of T cell differentiation, we identified T conv cell clones that are recurrently enriched in non-lymphoid organs following ablation of Foxp3 + regulatory T (T reg ) cells. A subset of these clones was highly proliferative in the lymphoid organs at steady state and exhibited overt reactivity to self-ligands displayed by dendritic cells, yet were not purged by clonal deletion. These clones spontaneously adopted numerous hallmarks of follicular helper T (T FH ) cells, including expression of Bcl6 and PD-1, exhibited an elevated propensity to localize within B cell follicles at steady state, and produced interferon-γ in non-lymphoid organs following sustained T reg cell depletion. Our work identifies a naturally occurring population of self-reactive T FH -like cells and delineates a previously unappreciated fate for self-specific T conv cells., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

With the combination of KRASG12C and EGFR inhibitors, KRAS is becoming a druggable target in colorectal cancer. However, secondary resistance limits its efficacy. Using cell lines, patient-derived xenografts, and patient samples, we detected a heterogeneous pattern of putative resistance alterations expected primarily to prevent inhibition of ERK signaling by drugs at progression. Serial analysis of patient blood samples on treatment demonstrates that most of these alterations are detected at a low frequency except for KRASG12C amplification, a recurrent resistance mechanism that rises in step with clinical progression. Upon drug withdrawal, resistant cells with KRASG12C amplification undergo oncogene-induced senescence, and progressing patients experience a rapid fall in levels of this alteration in circulating DNA. In this new state, drug resumption is ineffective as mTOR signaling is elevated. However, our work exposes a potential therapeutic vulnerability, whereby therapies that target the senescence response may overcome acquired resistance., Significance: Clinical resistance to KRASG12C-EGFR inhibition primarily prevents suppression of ERK signaling. Most resistance mechanisms are subclonal, whereas KRASG12C amplification rises over time to drive a higher portion of resistance. This recurrent resistance mechanism leads to oncogene-induced senescence upon drug withdrawal and creates a potential vulnerability to senolytic approaches. This article is highlighted in the In This Issue feature, p. 1., (©2022 The Authors; Published by the American Association for Cancer Research.)

Response to immunotherapy across multiple cancer types is approximately 25%, with some tumor types showing increased response rates compared to others (i.e. response rates in melanoma and non-small cell lung cancer (NSCLC) are typically 30-60%). Patients whose tumors are resistant to immunotherapy often lack high levels of pre-existing inflammation in the tumor microenvironment. Increased tumor glycolysis, acting through glucose deprivation and lactic acid accumulation, has been shown to have pleiotropic immune suppressive effects using in-vitro and in-vivo models of disease. To determine whether the immune suppressive effect of tumor glycolysis is observed across human solid tumors, we analyzed glycolytic and immune gene expression patterns in multiple solid malignancies. We found that increased expression of a glycolytic signature was associated with decreased immune infiltration and a more aggressive disease across multiple tumor types. Radiologic and pathologic analysis of untreated estrogen receptor (ER)-negative breast cancers corroborated these observations, and demonstrated that protein expression of glycolytic enzymes correlates positively with glucose uptake and negatively with infiltration of CD3 + and CD8 + lymphocytes. This study reveals an inverse relationship between tumor glycolysis and immune infiltration in a large cohort of multiple solid tumor types., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Cohen, Pareja, Socci, Shen, Doane, Schwartz, Khanin, Morris, Sutton and Blasberg.)

Cancers arising from the bladder urothelium often exhibit lineage plasticity with regions of urothelial carcinoma adjacent to or admixed with regions of divergent histomorphology, most commonly squamous differentiation. To define the biologic basis for and clinical significance of this morphologic heterogeneity, here we perform integrated genomic analyses of mixed histology bladder cancers with separable regions of urothelial and squamous differentiation. We find that squamous differentiation is a marker of intratumoral genomic and immunologic heterogeneity in patients with bladder cancer and a biomarker of intrinsic immunotherapy resistance. Phylogenetic analysis confirms that in all cases the urothelial and squamous regions are derived from a common shared precursor. Despite the presence of marked genomic heterogeneity between co-existent urothelial and squamous differentiated regions, no recurrent genomic alteration exclusive to the urothelial or squamous morphologies is identified. Rather, lineage plasticity in bladder cancers with squamous differentiation is associated with loss of expression of FOXA1, GATA3, and PPARG, transcription factors critical for maintenance of urothelial cell identity. Of clinical significance, lineage plasticity and PD-L1 expression is coordinately dysregulated via FOXA1, with patients exhibiting morphologic heterogeneity pre-treatment significantly less likely to respond to immune checkpoint inhibitors., (© 2022. The Author(s).)

Although p53 inactivation promotes genomic instability 1 and presents a route to malignancy for more than half of all human cancers 2,3 , the patterns through which heterogenous TP53 (encoding human p53) mutant genomes emerge and influence tumorigenesis remain poorly understood. Here, in a mouse model of pancreatic ductal adenocarcinoma that reports sporadic p53 loss of heterozygosity before cancer onset, we find that malignant properties enabled by p53 inactivation are acquired through a predictable pattern of genome evolution. Single-cell sequencing and in situ genotyping of cells from the point of p53 inactivation through progression to frank cancer reveal that this deterministic behaviour involves four sequential phases-Trp53 (encoding mouse p53) loss of heterozygosity, accumulation of deletions, genome doubling, and the emergence of gains and amplifications-each associated with specific histological stages across the premalignant and malignant spectrum. Despite rampant heterogeneity, the deletion events that follow p53 inactivation target functionally relevant pathways that can shape genomic evolution and remain fixed as homogenous events in diverse malignant populations. Thus, loss of p53-the 'guardian of the genome'-is not merely a gateway to genetic chaos but, rather, can enable deterministic patterns of genome evolution that may point to new strategies for the treatment of TP53-mutant tumours., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

The genetic, biologic, and clinical heterogeneity of sarcomas poses a challenge for the identification of therapeutic targets, clinical research, and advancing patient care. Because there are > 100 sarcoma subtypes, in-depth genetic studies have focused on one or a few subtypes. Herein, we report a comparative genetic analysis of 2,138 sarcomas representing 45 pathological entities. This cohort is prospectively analyzed using targeted sequencing to characterize subtype-specific somatic alterations in targetable pathways, rates of whole genome doubling, mutational signatures, and subtype-agnostic genomic clusters. The most common alterations are in cell cycle control and TP53, receptor tyrosine kinases/PI3K/RAS, and epigenetic regulators. Subtype-specific associations include TERT amplification in intimal sarcoma and SWI/SNF alterations in uterine adenosarcoma. Tumor mutational burden, while low compared to other cancers, varies between and within subtypes. This resource will improve sarcoma models, motivate studies of subtype-specific alterations, and inform investigations of genetic factors and their correlations with treatment response., (© 2022. The Author(s).)

Access to clinically relevant small cell lung cancer (SCLC) tissue is limited because surgical resection is rare in metastatic SCLC. Patient-derived xenografts (PDX) and circulating tumor cell-derived xenografts (CDX) have emerged as valuable tools to characterize SCLC. Here, we present a resource of 46 extensively annotated PDX/CDX models derived from 33 patients with SCLC. We perform multi-omic analyses, using targeted tumor next-generation sequencing, RNA-sequencing, and immunohistochemistry to deconvolute the mutational landscapes, global expression profiles, and molecular subtypes of these SCLC models. SCLC subtypes characterized by transcriptional regulators, ASCL1, NEUROD1 and POU2F3 are confirmed in this cohort. A subset of SCLC clinical specimens, including matched PDX/CDX and clinical specimen pairs, confirm that the primary features and genomic and proteomic landscapes of the tumors of origin are preserved in the derivative PDX models. This resource provides a powerful system to study SCLC biology., (© 2022. The Author(s).)

Cancer immunotherapy can result in lasting tumor regression, but predictive biomarkers of treatment response remain ill-defined. Here, we performed single-cell proteomics, transcriptomics, and genomics on matched untreated and IL2 injected metastases from patients with melanoma. Lesions that completely regressed following intralesional IL2 harbored increased fractions and densities of nonproliferating CD8+ T cells lacking expression of PD-1, LAG-3, and TIM-3 (PD-1-LAG-3-TIM-3-). Untreated lesions from patients who subsequently responded with complete eradication of all tumor cells in all injected lesions (individuals referred to herein as "extreme responders") were characterized by proliferating CD8+ T cells with an exhausted phenotype (PD-1+LAG-3+TIM-3+), stromal B-cell aggregates, and expression of IFNγ and IL2 response genes. Loss of membranous MHC class I expression in tumor cells of untreated lesions was associated with resistance to IL2 therapy. We validated this finding in an independent cohort of metastatic melanoma patients treated with intralesional or systemic IL2. Our study suggests that intact tumor-cell antigen presentation is required for melanoma response to IL2 and describes a multidimensional and spatial approach to develop immuno-oncology biomarker hypotheses using routinely collected clinical biospecimens., (©2022 American Association for Cancer Research.)

T cell receptor (TCR) signal strength is a key determinant of T cell responses. We developed a cancer mouse model in which tumor-specific CD8 T cells (TST cells) encounter tumor antigens with varying TCR signal strength. High-signal-strength interactions caused TST cells to up-regulate inhibitory receptors (IRs), lose effector function, and establish a dysfunction-associated molecular program. TST cells undergoing low-signal-strength interactions also up-regulated IRs, including PD1, but retained a cell-intrinsic functional state. Surprisingly, neither high- nor low-signal-strength interactions led to tumor control in vivo, revealing two distinct mechanisms by which PD1hi TST cells permit tumor escape; high signal strength drives dysfunction, while low signal strength results in functional inertness, where the signal strength is too low to mediate effective cancer cell killing by functional TST cells. CRISPR-Cas9-mediated fine-tuning of signal strength to an intermediate range improved anti-tumor activity in vivo. Our study defines the role of TCR signal strength in TST cell function, with important implications for T cell-based cancer immunotherapies., Competing Interests: Disclosures: N.A. Defranoux reports personal fees from Alector outside the submitted work. M.D. Hellman receives institutional research funding from Bristol-Myers Squibb; has been a compensated consultant for Merck, Bristol-Myers Squibb, AstraZeneca, Genentech/Roche, Nektar, Syndax, Mirati, Shattuck Labs, Immunai, Blueprint Medicines, Achilles, and Arcus; received travel support/honoraria from AstraZeneca, Eli Lilly, and Bristol-Myers Squibb; has options from Factorial, Shattuck Labs, Immunai, and Arcus; reported personal fees from Adicet, DaVolterra, Genzyme/Sanofi, Janssen, Immunai, Instil Bio, Mana Therapeutics, Natera, Pact Pharms, Chattuck Labs, and Regenron out side the submitted work; and has a patent filed by his institution related to the use of tumor mutation burden to predict response to immunotherapy (PCT/US2015/062208), which has received licensing fees from PGDx. D.K. Wells is a founder, equity holder, and receives advisory fees from Immunai. No other disclosures were reported., (© 2021 Shakiba et al.)