Preclinical evidence for the use of anti-Trop-2 antibody-drug conjugate Sacituzumab govitecan in cerebral metastasized castration-resistant prostate cancer.

PURPOSE: Improved survival rates have been observed in castration-resistant prostate cancer (CRPC) due to advancements in treatment options. However, individuals with brain metastases still have limited therapeutic options and an unfavorable prognosis. Therefore, there is an urgent need to explore new therapeutic avenues, such as antibody-drug conjugates (ADCs), which have demonstrated significant clinical activity against active brain metastases in solid tumors. Our objective was to determine the expression levels of the ADC targets Trop-2 and NECTIN-4 in cerebral metastasized CRPC (mCRPC). METHODS: Immunohistochemical staining of Trop-2 and NECTIN-4 with evaluation of H-score was performed in CRPC brain metastases (n = 31). Additionally, we examined Trop-2 protein expression in prostate cancer cell lines and studied their responsiveness to the anti-Trop-2 ADC Sacituzumab govitecan (SG) in vitro. RESULTS: Our analysis revealed that most patients exhibited moderate to strong Trop-2 expression [n = 27/31 with H-score ≥100, median H-score 220 (IQR 180-280)], while NECTIN-4 was absent in all cerebral metastases. Mechanistically, we demonstrated that the efficacy of SG depends on Trop-2 expression levels in vitro. Overexpression of Trop-2 in Trop-2-negative PC-3 cells led to sensitization to SG, whereas CRISPR-Cas9-mediated knockdown of Trop-2 in Trop-2-expressing DU-145 cells conferred resistance to SG. CONCLUSION: The substantial expression of Trop-2 in cerebral metastases, along with our preclinical in vitro results, supports the efficacy of SG in treating cerebral mCRPC. Thus, our results extend the understanding of the potential of ADCs in prostate cancer treatment and provide an additional treatment strategy for the challenging subset of patients with cerebral metastases.

Integration of on-treatment modified Glasgow prognostic score (mGPS) to improve imaging-based prediction of outcomes in patients with non-small cell lung cancer on immune checkpoint inhibition.

INTRODUCTION: A large number of patients with non-small cell lung cancer (NSCLC) on immune checkpoint inhibition (ICI) achieve stable disease (SD) as the best overall response, which is associated with heterogeneous outcomes. In this context, complementary biomarkers that improve outcome prediction are needed. We have recently demonstrated that measuring the on-treatment modified Glasgow prognostic score (mGPS), which is based on the two serum markers C-reactive protein (CRP) and albumin, can improve outcome prediction complementary to radiological staging in metastatic renal cell carcinoma. However, this concept has not been assessed for patients with NSCLC on ICI. METHODS: We assessed the prognostic and predictive value of on-treatment mGPS at week six in patients with NSCLC treated with atezolizumab or docetaxel in the phase 3 OAK trial (NCT02008227) comprising n = 750 patients and validated the findings in the phase 2 BIRCH (NCT02031458, n = 560). RESULTS: On-treatment mGPS assessed at week six demonstrated valuable prognostic information (Hazard Ratio (HR) for mGPS low-risk vs intermediate risk 2.34 (95 % CI 1.76-3.11, p < 0.001) and vs high risk 3.56, (95 % CI 2.57-4.91, p < 0.001) in the atezolizumab-treated subgroup. On-treatment mGPS predicted overall survival more accurately than imaging using RECIST criteria (concordance index: on-treatment mGPS 0.646 (95 % CI 0.615-0.677) vs RECIST 0.606 (95 % CI 0.575-0.637)). On-treatment mGPS provides additional prognostic information to imaging-assessed treatment response at first staging, especially for the patient subgroup with SD. These findings were validated in the BIRCH trial. CONCLUSIONS: We highlight the novel concept of integrating on-treatment mGPS for improved outcome prediction in conjunction with radiological imaging for patients with NSCLC on ICI.

Adipocyte Precursor-Derived NRG1 Promotes Resistance to FGFR Inhibition in Urothelial Carcinoma.

Aberrations of the fibroblast growth factor receptor (FGFR) family members are frequently observed in metastatic urothelial cancer (mUC), and blocking the FGF/FGFR signaling axis is used as a targeted therapeutic strategy for treating patients. Erdafitinib is a pan-FGFR inhibitor, which has recently been approved by the FDA for mUC with FGFR2/3 alterations. Although mUC patients show initial response to erdafitinib, acquired resistance rapidly develops. Here, we found that adipocyte precursors promoted resistance to erdafitinib in FGFR-dependent bladder and lung cancer in a paracrine manner. Moreover, neuregulin 1 (NRG1) secreted from adipocyte precursors was a mediator of erdafitinib resistance by activating human epidermal growth factor receptor 3 (ERBB3; also known as HER3) signaling, and knockdown of NRG1 in adipocyte precursors abrogated the conferred paracrine resistance. NRG1 expression was significantly downregulated in terminally differentiated adipocytes compared with their progenitors. Pharmacologic inhibition of the NRG1/HER3 axis using pertuzumab reversed erdafitinib resistance in tumor cells in vitro and prolonged survival of mice bearing bladder cancer xenografts in vivo. Remarkably, data from single-cell RNA sequencing revealed that NRG1 was enriched in platelet-derived growth factor receptor-A (PDGFRA) expressing inflammatory cancer-associated fibroblasts, which is also expressed on adipocyte precursors. Together, this work reveals a paracrine mechanism of anti-FGFR resistance in bladder cancer, and potentially other cancers, that is amenable to inhibition using available targeted therapies. SIGNIFICANCE: Acquired resistance to FGFR inhibition can be rapidly promoted by paracrine activation of the NRG1/HER3 axis mediated by adipocyte precursors and can be overcome by the combination of pertuzumab and erdafitinib treatment. See related commentary by Kolonin and Anastassiou, p. 648.

A replicating LCMV-based vaccine for the treatment of solid tumors.

Harnessing the immune system to eradicate tumors requires identification and targeting of tumor antigens, including tumor-specific neoantigens and tumor-associated self-antigens. Tumor-associated antigens are subject to existing immune tolerance, which must be overcome by immunotherapies. Despite many novel immunotherapies reaching clinical trials, inducing self-antigen-specific immune responses remains challenging. Here, we systematically investigate viral-vector-based cancer vaccines encoding a tumor-associated self-antigen (TRP2) for the treatment of established melanomas in preclinical mouse models, alone or in combination with adoptive T cell therapy. We reveal that, unlike foreign antigens, tumor-associated antigens require replication of lymphocytic choriomeningitis virus (LCMV)-based vectors to break tolerance and induce effective antigen-specific CD8+ T cell responses. Immunization with a replicating LCMV vector leads to complete tumor rejection when combined with adoptive TRP2-specific T cell transfer. Importantly, immunization with replicating vectors leads to extended antigen persistence in secondary lymphoid organs, resulting in efficient T cell priming, which renders previously "cold" tumors open to immune infiltration and reprograms the tumor microenvironment to "hot." Our findings have important implications for the design of next-generation immunotherapies targeting solid cancers utilizing viral vectors and adoptive cell transfer.

Pretreatment albumin is a prognostic and predictive biomarker for response to atezolizumab across solid tumors.

Objectives: Reliable predictive biomarkers for response to immune checkpoint inhibition (ICI) are lacking. Pretreatment serum albumin, a known prognostic and predictive factor in ICI-treated patients, has been proposed as a potential pharmacokinetic surrogate marker for anti-PD1/PD-L1 antibodies, as it shares a homeostatic pathway with IgG. However, this hypothesis is currently based on theoretical considerations and limited evidence from retrospective data. Therefore, we comprehensively investigated the prognostic and predictive value of pretreatment albumin and its relationship with anti-PD-L1 IgG levels. Methods: We analysed pretreatment albumin and atezolizumab serum levels and clinical response in four trials (IMvigor210, IMvigor211, IMmotion151 and OAK) of patients with metastatic lung-, renal- or urothelial cancer who received atezolizumab alone or in combination. Results: A total of 3391 patients were analysed. Correlation between serum albumin and atezolizumab levels was weak (Pearson's coefficient 0.23). We found a strong prognostic value for pretreatment serum albumin across all trials. Both atezolizumab serum levels and serum albumin were independently correlated with overall survival. Importantly, in the three randomised phase III clinical trials, the survival benefit for immunotherapy compared with the active comparator arm was limited to patients with pretreatment serum albumin > 35 g L-1. Conclusion: Our data do not support the hypothesis that albumin serves as a surrogate for atezolizumab pharmacokinetics. However, we show that albumin on its own exerts strong prognostic value for patients treated with immunotherapy. As benefit from immunotherapy was limited to patients with normal/elevated serum albumin levels, baseline albumin could potentially be used as a predictive marker for immune checkpoint inhibition.

On-treatment Modified Glasgow Prognostic Score Provides Predictive Information Complementary to Radiological Staging in Metastatic Urothelial Carcinoma on Immunotherapy.

In the immunotherapy era it is difficult to predict patient prognosis on the basis of radiological staging alone, especially for the subgroup with stable disease (SD), which encompasses a wide range of clinical outcomes. Thus, there is need for reliable and, ideally, cost-efficient biomarkers to improve the accuracy of outcome prediction. We evaluated the on-treatment modified Glasgow Prognostic Score (mGPS)-a known predictor of outcomes in several cancers that is based on serum C-reactive protein and albumin-in patients with metastatic urothelial carcinoma (mUC) treated with immune checkpoint inhibition (ICI) in the phase 2 IMvigor210 and phase 3 IMvigor211 trials. On-treatment mGPS provides valuable prognostic information complementary to radiological staging, particularly for patients with SD. In IMvigor210, on-treatment mGPS predicts outcomes as early as 6 wk after ICI initiation, considerably before the first routine staging typically performed after 10-12 wk. Our study suggests that on-treatment mGPS complements radiological imaging in predicting outcomes for patients with mUC undergoing ICI. PATIENT SUMMARY: For patients with metastatic bladder cancer receiving immunotherapy, it is difficult to predict treatment outcomes from imaging scans alone. Our study results suggest that a score called the modified Glasgow Prognostic Score based on just two proteins (C-reactive protein and albumin) measured in blood can accurately predict outcomes. Use of the mGPS along with imaging scans may be better in predicting the survival benefit from immunotherapy.

Integrating On-Treatment Modified Glasgow Prognostic Score and Imaging to Predict Response and Outcomes in Metastatic Renal Cell Carcinoma.

Importance: In the era of immuno-oncology, imaging alone seems to be insufficient to capture treatment responses, as patients with stable disease treated with immunotherapy have a wide range of clinical outcomes. There is an unmet need for complementary (ideally cost-efficient) markers that enable assessment of therapy response and outcomes in conjunction with imaging. Objectives: To examine whether longitudinal changes in the modified Glasgow prognostic score (mGPS), which is based on C-reactive protein and albumin, can predict responses and outcomes in patients with metastatic renal cell carcinoma (mRCC). Design, Setting, and Participants: This post hoc analysis, conducted from October 2022 to April 2023, evaluated the prognostic and predictive performance of on-treatment mGPS in patients with mRCC being treated with atezolizumab (plus bevacizumab) or sunitinib in 2 randomized clinical trials: the phase 3 IMmotion151 study (discovery cohort) and the phase 2 IMmotion150 study (validation cohort). Main Outcomes and Measures: Outcomes were investigator-assessed progression-free survival (PFS) per Response Evaluation Criteria in Solid Tumors (RECIST), version 1.1 and overall survival (OS) for survival analyses. To compare the prognostic value of the on-treatment mGPS with radiologic staging, we used RECIST assessed by the Independent Review Committee (IRC-RECIST) to ensure high data quality. Results: Of the 915 patients with mRCC in the IMmotion151 discovery cohort, baseline mGPS was available for 861 patients and on-treatment mGPS for 691. The IMmotion150 validation cohort included 305 patients with mRCC, and on-treatment mGPS could be evaluated for 199. In the IMmotion150 study, on-treatment mGPS predicted outcomes as early as 6 weeks following therapy initiation, thereby opening a window for early therapy adjustments. In both clinical trials, on-treatment mGPS provided valuable prognostic information regardless of imaging-assessed treatment response at first staging. Of note, in the disease control subgroup, on-treatment mGPS exhibited superior and independent prognostic information compared with IRC-RECIST (available for 611 patients; C-index, 0.651 [95% CI, 0.588-0.714] for the mGPS during treatment vs 0.574 [95% CI, 0.528-0.619] for IRC-RECIST). Conclusions and Relevance: These data support the concept of integrating on-treatment mGPS for more holistic and patient-centered therapy monitoring in addition to radiologic staging to improve clinical care at a low cost for patients with mRCC.

Stressed out: NKp46 binds ecto-calreticulin.

In a recent article, Sen Santara et al. demonstrated that the activating natural killer (NK) cell receptor NKp46 binds to externalized calreticulin (ecto-CRT), leading to NK cell degranulation and target cell killing. They show that endoplasmic reticulum stress-induced ecto-CRT serves as a danger-associated molecular pattern, helping NK cells identify and eliminate infected, malignant, stressed or senescent cells.

Oncogenic drivers dictate immune control of acute myeloid leukemia.

Acute myeloid leukemia (AML) is a genetically heterogeneous, aggressive hematological malignancy induced by distinct oncogenic driver mutations. The effect of specific AML oncogenes on immune activation or suppression is unclear. Here, we examine immune responses in genetically distinct models of AML and demonstrate that specific AML oncogenes dictate immunogenicity, the quality of immune response and immune escape through immunoediting. Specifically, expression of NrasG12D alone is sufficient to drive a potent anti-leukemia response through increased MHC Class II expression that can be overcome with increased expression of Myc. These data have important implications for the design and implementation of personalized immunotherapies for patients with AML.

Early C-reactive protein kinetics predicts immunotherapy response in non-small cell lung cancer in the phase III OAK trial.

Static biomarkers like programmed death-ligand 1 (PD-L1) are insufficient to accurately predict response to immune checkpoint inhibition. Therefore, on-treatment biomarkers, which measure immediate therapy-associated changes, are currently shifting into the focus of immuno-oncology. A prime example of a simple predictive on-treatment biomarker is the early C-reactive protein (CRP) kinetics with its predictive CRP flare-response phenomenon. Here, we were able to confirm the predictive value of CRP flare-response kinetics in the pivotal phase III OAK trial (NCT02008227), which compared atezolizumab with docetaxel in patients with non-small cell lung cancer. Of note, CRP flare-response predicted favorable outcomes only in the immune checkpoint inhibition-treated subgroup, which suggests that it is an immunotherapy-specific phenomenon. In conclusion, we have for the first time validated the high predictive value of early CRP kinetics in a pivotal phase III trial, justifying the broad use of this cost-effective and easy-to-implement on-treatment biomarker to optimize therapy monitoring for patients with non-small cell lung cancer.

Neutrophil Conversion to a Tumor-Killing Phenotype Underpins Effective Microbial Therapy.

The inflammatory microenvironment of solid tumors creates a protumorigenic milieu that resembles chronic inflammation akin to a subverted wound healing response. Here, we investigated the effect of converting the tumor microenvironment from a chronically inflamed state to one of acute microbial inflammation by injecting microbial bioparticles directly into tumors. Intratumoral microbial bioparticle injection led to rapid and dramatic changes in the tumor immune composition, the most striking of which was a substantial increase in the presence of activated neutrophils. In situ photoconversion and intravital microscopy indicated that tumor neutrophils transiently switched from sessile producers of VEGF to highly motile neutrophils that clustered to make neutrophil-rich domains in the tumor. The neutrophil clusters remodeled tumor tissue and repressed tumor growth. Single-cell transcriptional analysis of microbe-stimulated neutrophils showed a profound shift in gene expression towards heightened activation and antimicrobial effector function. Microbe-activated neutrophils also upregulated chemokines known to regulate neutrophil and CD8+ T-cell recruitment. Microbial therapy also boosted CD8+ T-cell function and enhanced the therapeutic benefit of checkpoint inhibitor therapy in tumor-bearing mice and provided protection in a model of tumor recurrence. These data indicate that one of the major effector mechanisms of microbial therapy is the conversion of tumor neutrophils from a wound healing to an acutely activated cytotoxic phenotype, highlighting a rationale for broader deployment of microbial therapy in the treatment of solid cancers. SIGNIFICANCE: Intratumoral injection of microbial bioparticles stimulates neutrophil antitumor functions, suggesting pathways for optimizing efficacy of microbial therapies and paving the way for their broader utilization in the clinic.

CDCP1 expression is frequently increased in aggressive urothelial carcinoma and promotes urothelial tumor progression.

The prognosis of patients with advanced urothelial carcinoma (UC) remains poor and improving treatment continues to be a major medical need. CUB domain containing protein 1 (CDCP1) is a known oncogene in various types of solid cancers and its overexpression is associated with impaired prognosis. However, its role in UC remains undetermined. Here we assessed the clinical relevance of CDCP1 in two cohorts of UC at different stages of the disease. Immunohistochemistry showed that CDCP1 is highly expressed in advanced UC, which significantly correlates with shorter overall survival. Importantly, the basal/squamous UC subtype showed significantly enriched CDCP1 at the mRNA and protein levels. The functional role of CDCP1 overexpression was assessed taking advantage of ex vivo organoids derived from the CDCP1pcLSL/+ transgenic mouse model. Furthermore, CDCP1 knockout UC cell lines were generated using CRISPR/Cas9 technology. Interestingly, CDCP1 overexpression significantly induced the activation of MAPK/ERK pathways in ex vivo organoids and increased their proliferation. Similarly, CDCP1 knockout in UC cell lines reduced their proliferation and migration, concomitant with MAPK/ERK pathway activity reduction. Our results highlight the relevance of CDCP1 in advanced UC and demonstrate its oncogenic role, suggesting that targeting CDCP1 could be a rational therapeutic strategy for the treatment of advanced UC.

Single-cell RNA sequencing identifies a population of human liver-type ILC1s.

Group 1 innate lymphoid cells (ILCs) comprise a heterogeneous family of cytotoxic natural killer (NK) cells and ILC1s. We identify a population of "liver-type" ILC1s with transcriptional, phenotypic, and functional features distinct from those of conventional and liver-resident NK cells as well as from other previously described human ILC1 subsets. LT-ILC1s are CD49a+CD94+CD200R1+, express the transcription factor T-BET, and do not express the activating receptor NKp80 or the transcription factor EOMES. Similar to NK cells, liver-type ILC1s produce IFN-γ, TNF-α, and GM-CSF; however, liver-type ILC1s also produce IL-2 and lack perforin and granzyme-B. Liver-type ILC1s are expanded in cirrhotic liver tissues, and they can be produced from blood-derived ILC precursors in vitro in the presence of TGF-ß1 and liver sinusoidal endothelial cells. Cells with similar signature and function can also be found in tonsil and intestinal tissues. Collectively, our study identifies and classifies a population of human cross-tissue ILC1s.

T cell-derived interleukin-22 drives the expression of CD155 by cancer cells to suppress NK cell function and promote metastasis.

Although T cells can exert potent anti-tumor immunity, a subset of T helper (Th) cells producing interleukin-22 (IL-22) in breast and lung tumors is linked to dismal patient outcome. Here, we examined the mechanisms whereby these T cells contribute to disease. In murine models of lung and breast cancer, constitutional and T cell-specific deletion of Il22 reduced metastases without affecting primary tumor growth. Deletion of the IL-22 receptor on cancer cells decreases metastasis to a degree similar to that seen in IL-22-deficient mice. IL-22 induced high expression of CD155, which bound to the activating receptor CD226 on NK cells. Excessive activation led to decreased amounts of CD226 and functionally impaired NK cells, which elevated the metastatic burden. IL-22 signaling was also associated with CD155 expression in human datasets and with poor patient outcomes. Taken together, our findings reveal an immunosuppressive circuit activated by T cell-derived IL-22 that promotes lung metastasis.

Targeting inflamed and non-inflamed melanomas: biological background and clinical challenges.

Immune checkpoint inhibitors (ICIs) have demonstrated impressive antitumor activity in patients with advanced and early stage melanoma, thus improving long-term survival outcomes. However, most patients derive limited benefit from immunotherapy, due to the development of primary, adaptive, or acquired resistance mechanisms. Immunotherapy resistance is a complex phenomenon that depends on genetic and epigenetic mechanisms which, in turn, drive the interplay between cancer cells and the tumor microenvironment (TME). Immunologically "cold" (i.e. non-inflamed) tumors lack or have few tumor infiltrating lymphocytes (TILs) as a result of low tumor mutational burden (TMB), defective antigen presentation, or physical barriers to lymphocyte migration, resulting in a minimal benefit from immunotherapy. In contrast, in most cases immunologically "hot" (i.e. inflamed) tumors display high TMB, implying a higher load of neoantigens and increased programmed cell death ligand 1 (PD-L1) expression, with a consequently higher rate of TILs. However, the presence of TILs does not necessarily denote the tumor as immunologically "hot", since the presence of tumor-specific CD8+ T cells persistently exposed to antigenic stimulation induces a dysfunctional state called "exhaustion", which leads to a reduced response to immunotherapy. In recent years, efforts have been made to characterize mechanisms of resistance to immunotherapy, and to investigate strategies to overcome treatment resistance. Indeed, predictors of response and toxicity to immunotherapy are still lacking and, to date, there are no reliable predictive biomarkers to select patients according to baseline clinical, histological, or genomic characteristics. In this review, we will focus on the morphologic and immunohistochemical characteristics of the TME, and on the molecular determinants of resistance to immunotherapy, differentiating between inflamed and non-inflamed melanomas. Then, we will provide a thorough overview of preclinical data on genetic and epigenetic mechanisms with a potential impact on the immune response and patient outcome. Finally, we will focus our attention on the role of potential biomarkers in determining disease response to immunotherapy, in the adjuvant and metastatic setting, providing an insight into current and future research in this field.

IFN-λ Diminishes the Severity of Viral Bronchiolitis in Neonatal Mice by Limiting NADPH Oxidase-Induced PAD4-Independent NETosis.

Infants with attenuated type III IFN (IFN-λ) responses are at increased risk of severe lower respiratory tract infection (sLRI). The IL-28Rα-chain and IL-10Rß-chain form a heterodimeric receptor complex, necessary for IFN-λ signaling. Therefore, to better understand the immunopathogenic mechanisms through which an IFN-λlo microenvironment predisposes to a sLRI, we inoculated neonatal wild-type and IL-28R-deficient (IL-28R -/-) mice with pneumonia virus of mice, a rodent-specific pneumovirus. Infected IL-28R -/- neonates displayed an early, pronounced, and persistent neutrophilia that was associated with enhanced reactive oxygen species (ROS) production, NETosis, and mucus hypersecretion. Targeted deletion of the IL-28R in neutrophils was sufficient to increase neutrophil activation, ROS production, NET formation, and mucus production in the airways. Inhibition of protein-arginine deiminase type 4 (PAD4), a regulator of NETosis, had no effect on myeloperoxidase expression, citrullinated histones, and the magnitude of the inflammatory response in the lungs of infected IL-28R -/- mice. In contrast, inhibition of ROS production decreased NET formation, cellular inflammation, and mucus hypersecretion. These data suggest that IFN-λ signaling in neutrophils dampens ROS-induced NETosis, limiting the magnitude of the inflammatory response and mucus production. Therapeutics that promote IFN-λ signaling may confer protection against sLRI.

Plasticity of NK cells in Cancer.

Natural killer (NK) cells are crucial to various facets of human immunity and function through direct cytotoxicity or via orchestration of the broader immune response. NK cells exist across a wide range of functional and phenotypic identities. Murine and human studies have revealed that NK cells possess substantial plasticity and can alter their function and phenotype in response to external signals. NK cells also play a critical role in tumor immunity and form the basis for many emerging immunotherapeutic approaches. NK cells can directly target and lyse malignant cells with their inherent cytotoxic capabilities. In addition to direct targeting of malignant cells, certain subsets of NK cells can mediate antibody-dependent cellular cytotoxicity (ADCC) which is integral to some forms of immune checkpoint-blockade immunotherapy. Another important feature of various NK cell subsets is to co-ordinate anti-tumor immune responses by recruiting adaptive and innate leukocytes. However, given the diverse range of NK cell identities it is unsurprising that both pro-tumoral and anti-tumoral NK cell subsets have been described. Here, NK cell subsets have been shown to promote angiogenesis, drive inflammation and immune evasion in the tumor microenvironment. To date, the signals that drive tumor-infiltrating NK cells towards the acquisition of a pro- or anti-tumoral function are poorly understood. The notion of tumor microenvironment-driven NK cell plasticity has substantial implications for the development of NK-based immunotherapeutics. This review will highlight the current knowledge of NK cell plasticity pertaining to the tumor microenvironment. Additionally, this review will pose critical and relevant questions that need to be addressed by the field in coming years.