Lung squamous cell carcinoma (LUSC) represents a major subtype of non-small cell lung cancer with limited treatment options. Previous studies have elucidated the complex genetic landscape of LUSC and ...revealed multiple altered genes and pathways. However, in stark contrast to lung adenocarcinoma, few targetable driver mutations have been established so far and targeted therapies for LUSC remain unsuccessful. Immunotherapy has revolutionized LUSC treatment and is currently approved as the new standard of care. To gain a better understanding of the LUSC biology, improved modeling systems are urgently needed. Preclinical models, particularly those mimicking human disease with an intact tumor immune microenvironment, are an invaluable tool to study cancer development and evaluate new therapeutic targets. Here, we discuss recent advances in LUSC preclinical models, with a focus on genetically engineered mouse models (GEMMs) and organoids, in the context of evolving precision medicine and immunotherapy.
Lung squamous cell carcinoma (LSCC) is a deadly disease for which only a subset of patients responds to immune checkpoint blockade (ICB) therapy. Therefore, preclinical mouse models that recapitulate ...the complex genetic profile found in patients are urgently needed.
We used CRISPR genome editing to delete multiple tumor suppressors in lung organoids derived from Cre-dependent SOX2 knock-in mice. We investigated both the therapeutic efficacy and immunologic effects accompanying combination PD-1 blockade and WEE1 inhibition in both mouse models and LSCC patient-derived cell lines.
We show that multiplex gene editing of mouse lung organoids using the CRISPR-Cas9 system allows for efficient and rapid means to generate LSCCs that closely mimic the human disease at the genomic and phenotypic level. Using this genetically defined mouse model and three-dimensional tumoroid culture system, we show that WEE1 inhibition induces DNA damage that primes the endogenous type I IFN and antigen presentation system in primary LSCC tumor cells. These events promote cytotoxic T-cell-mediated clearance of tumor cells and reduce the accumulation of tumor-infiltrating neutrophils. Beneficial immunologic features of WEE1 inhibition are further enhanced by the addition of anti-PD-1 therapy.
We developed a mouse model system to investigate a novel combinatory approach that illuminates a clinical path hypothesis for combining ICB with DNA damage-inducing therapies in the treatment of LSCC.
BackgroundSmall cell lung cancer (SCLC) is a deadly disease with a 5-year survival of less than 7%. The addition of immunotherapy to chemotherapy was recently approved as first-line treatment; ...however, the improved clinical benefit is modest, highlighting an urgent need for new treatment strategies. Nemvaleukin alfa, a novel engineered interleukin-2 fusion protein currently in phase I–III studies, is designed to selectively expand cytotoxic natural killer (NK) cells and CD8+ T cells. Here, using a novel SCLC murine model, we investigated the effects of a mouse version of nemvaleukin (mNemvaleukin) on tumor growth and antitumor immunity.MethodsA novel Rb1−/−p53−/−p130−/− SCLC model that mimics human disease was generated. After confirming tumor burden by MRI, mice were randomized into four treatment groups: vehicle, mNemvaleukin alone, chemotherapy (cisplatin+etoposide) alone, or the combination of mNemvaleukin and chemotherapy. Tumor growth was measured by MRI and survival was recorded. Tumor-infiltrating lymphocytes and peripheral blood immune cells were analyzed by flow cytometry. Cytokine and chemokine secretion were quantified and transcriptomic analysis was performed to characterize the immune gene signatures.ResultsmNemvaleukin significantly inhibited SCLC tumor growth, which was further enhanced by the addition of chemotherapy. Combining mNemvaleukin with chemotherapy provided the most significant survival benefit. Profiling of tumor-infiltrating lymphocytes revealed mNemvaleukin expanded the total number of tumor-infiltrating NK and CD8+ T cells. Furthermore, mNemvaleukin increased the frequencies of activated and proliferating NK and CD8+ T cells in tumors. Similar immune alterations were observed in the peripheral blood of mNemvaleukin-treated mice. Of note, combining mNemvaleukin with chemotherapy had the strongest effects in activating effector and cytotoxic CD8+ T cells. mNemvaleukin alone, and in combination with chemotherapy, promoted proinflammatory cytokine and chemokine production, which was further confirmed by transcriptomic analysis.ConclusionsmNemvaleukin, a novel cytokine-based immunotherapy, significantly inhibited murine SCLC tumor growth and prolonged survival, which was further enhanced by the addition of chemotherapy. mNemvaleukin alone, and in combination with chemotherapy, drove a strong antitumor immune program elicited by cytotoxic immune cells. Our findings support the evaluation of nemvaleukin alone or in combination with chemotherapy in clinical trials for the treatment of SCLC.
Mechanical forces have been proposed to modulate organ growth, but a molecular mechanism that links them to growth regulation in vivo has been lacking. We report that increasing tension within the ...cytoskeleton increases Drosophila wing growth, whereas decreasing cytoskeletal tension decreases wing growth. These changes in growth can be accounted for by changes in the activity of Yorkie, a transcription factor regulated by the Hippo pathway. The influence of myosin activity on Yorkie depends genetically on the Ajuba LIM protein Jub, a negative regulator of Warts within the Hippo pathway. We further show that Jub associates with α-catenin and that its localization to adherens junctions and association with α-catenin are promoted by cytoskeletal tension. Jub recruits Warts to junctions in a tension-dependent manner. Our observations delineate a mechanism that links cytoskeletal tension to regulation of Hippo pathway activity, providing a molecular understanding of how mechanical forces can modulate organ growth.
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•Cytoskeletal tension regulates Drosophila wing growth through the Hippo pathway•The Ajuba protein Jub is regulated by tension and links tension to Hippo signaling•Jub inhibits Warts and recruits Warts to junctions in a tension-dependent manner•Our observations delineate a molecular mechanism that links tension to growth control
Mechanical force on the cytoskeleton controls organ growth in Drosophila through the tension-dependent recruitment of Jub protein to adherens junctions.
Squamous cell lung cancers (lung squamous cell carcinomas LUSCs) are associated with high mortality and a lack of therapies specific to this disease. Although recurrent molecular aberrations are ...present in LUSCs, efforts to develop targeted therapies against receptor tyrosine kinases, signaling transduction, and cell cycle checkpoints in LUSCs were met with significant challenges. The present therapeutic landscape focuses on epigenetic therapies to modulate the expression of lineage-dependent survival pathways and undruggable oncogenes. Another important therapeutic approach is to exploit metabolic vulnerabilities unique to LUSCs. These novel therapies may synergize with immune checkpoint inhibitors in the right therapeutic context. For example, the recognition that alterations in KEAP1-NFE2L2 in LUSCs affected antitumor immune responses created unique opportunities for targeted, metabolic, and immune combinations. This article provides a perspective on how lessons learned from the past influence the current therapeutic landscape and opportunities for future drug development for LUSCs.
Squamous cell lung cancers (lung squamous cell carcinomas LUSCs) are associated with high mortality and a lack of therapies specific to this disease. Although recurrent molecular aberrations are present in LUSCs, efforts to develop targeted therapies against receptor tyrosine kinases, signaling transduction, and cell cycle checkpoints in LUSCs were met with significant challenges. The present therapeutic landscape focuses on epigenetic therapies to modulate the expression of lineage-dependent survival pathways and undruggable oncogenes. Another important therapeutic approach is to exploit metabolic vulnerabilities unique to LUSCs. These novel therapies may synergize with immune checkpoint inhibitors in the right therapeutic context. For example, the recognition that alterations in KEAP1-NFE2L2 in LUSCs affected antitumor immune responses created unique opportunities for targeted, metabolic, and immune combinations. This article provides a perspective on how lessons learned from the past influence the current therapeutic landscape and opportunities for future drug development for LUSCs.
Mechanical stress can influence cell proliferation in vitro, but whether it makes a significant contribution to growth control in vivo, and how it is modulated and experienced by cells within ...developing tissues, has remained unclear. Here we report that differential growth reduces cytoskeletal tension along cell junctions within faster-growing cells. We propose a theoretical model to explain the observed reduction of tension within faster-growing clones, supporting it by computer simulations based on a generalized vertex model. This reduced tension modulates a biomechanical Hippo pathway, decreasing recruitment of Ajuba LIM protein and the Hippo pathway kinase Warts, and decreasing the activity of the growth-promoting transcription factor Yorkie. These observations provide a specific mechanism for a mechanical feedback that contributes to evenly distributed growth, and we show that genetically suppressing mechanical feedback alters patterns of cell proliferation in the developing Drosophila wing. By providing experimental support for the induction of mechanical stress by differential growth, and a molecular mechanism linking this stress to the regulation of growth in developing organs, our results confirm and extend the mechanical feedback hypothesis.
Tissue growth needs to be properly controlled for organs to reach their correct size and shape, but the mechanisms that control growth during normal development are not fully understood. We report ...here that the activity of the Hippo signaling transcriptional activator Yorkie gradually decreases in the central region of the developing
wing disc. Spatial and temporal changes in Yorkie activity can be explained by changes in cytoskeletal tension and biomechanical regulators of Hippo signaling. These changes in cellular biomechanics correlate with changes in cell density, and experimental manipulations of cell density are sufficient to alter biomechanical Hippo signaling and Yorkie activity. We also relate the pattern of Yorkie activity in older discs to patterns of cell proliferation. Our results establish that spatial and temporal patterns of Hippo signaling occur during wing development, that these patterns depend upon cell-density modulated tissue mechanics and that they contribute to the regulation of wing cell proliferation.
Cyclin-dependent kinase 7 (CDK7) is a central regulator of the cell cycle and gene transcription. However, little is known about its impact on genomic instability and cancer immunity. Using a ...selective CDK7 inhibitor, YKL-5-124, we demonstrated that CDK7 inhibition predominately disrupts cell-cycle progression and induces DNA replication stress and genome instability in small cell lung cancer (SCLC) while simultaneously triggering immune-response signaling. These tumor-intrinsic events provoke a robust immune surveillance program elicited by T cells, which is further enhanced by the addition of immune-checkpoint blockade. Combining YKL-5-124 with anti-PD-1 offers significant survival benefit in multiple highly aggressive murine models of SCLC, providing a rationale for new combination regimens consisting of CDK7 inhibitors and immunotherapies.
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•CDK7 inhibition impairs cell cycle and DNA replication and induces genome instability•CDK7 inhibitor YKL-5-124 activates IFN-γ signaling and induces TNF-α and CXCL9/10•YKL-5-124 provokes a robust immune program, which is further improved by anti-PD-1•Combining YKL-5-124 with anti-PD-1 and chemotherapy improves tumor response
Zhang et al. show that selective CDK7 inhibition with YKL-5-124 disrupts the cell cycle and causes replicative stress, eliciting an inflammatory response. YKL-5-124 in combination with anti-PD-1 therapy reduces tumor growth and increases survival in mouse models of small cell lung cancer.
Nuclear markers for live imaging are useful for counting and tracking cells, visualizing cell division, and examining the regulation of proteins that are controlled via entry or exit from the ...nucleus. Near-infrared fluorescent proteins have advantages over shorter wavelength fluorescent proteins, including reduced phototoxicity, less light scattering, and enabling multicolor live imaging. We have constructed and tested transgenic
expressing Histone H2Av iRFP fusion proteins, and confirmed that they can be used to label nuclei in both fixed and live tissue at multiple stages of development.
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