The Hallmarks of Precancer.

Research on precancers, as defined as at-risk tissues and early lesions, is of high significance given the effectiveness of early intervention. We discuss the need for risk stratification to prevent overtreatment, an emphasis on the role of genetic and epigenetic aging when considering risk, and the importance of integrating macroenvironmental risk factors with molecules and cells in lesions and at-risk normal tissues for developing effective intervention and health policy strategies.

Molecular cartography uncovers evolutionary and microenvironmental dynamics in sporadic colorectal tumors.

Colorectal cancer exhibits dynamic cellular and genetic heterogeneity during progression from precursor lesions toward malignancy. Analysis of spatial multi-omic data from 31 human colorectal specimens enabled phylogeographic mapping of tumor evolution that revealed individualized progression trajectories and accompanying microenvironmental and clonal alterations. Phylogeographic mapping ordered genetic events, classified tumors by their evolutionary dynamics, and placed clonal regions along global pseudotemporal progression trajectories encompassing the chromosomal instability (CIN+) and hypermutated (HM) pathways. Integrated single-cell and spatial transcriptomic data revealed recurring epithelial programs and infiltrating immune states along progression pseudotime. We discovered an immune exclusion signature (IEX), consisting of extracellular matrix regulators DDR1, TGFBI, PAK4, and DPEP1, that charts with CIN+ tumor progression, is associated with reduced cytotoxic cell infiltration, and shows prognostic value in independent cohorts. This spatial multi-omic atlas provides insights into colorectal tumor-microenvironment co-evolution, serving as a resource for stratification and targeted treatments.

Characterizing molecular subtypes of high-risk non-muscle-invasive bladder cancer in African American patients.

BACKGROUND: We sought to determine whether differences in subtype distribution and differentially expressed genes exist between African Americans (AAs) and European Americans (EAs) in patients with high-risk nonmuscle-invasive bladder cancer (NMIBC). METHODS: We performed a retrospective cohort study including 26 patients (14 AAs and 12 EAs) from the University of Texas Medical Branch and the Durham Veterans Affair Health Care System from 2010 to 2020 among treatment naïve, high-risk NMIBC. Profiled gene expressions were performed using the UROMOL classification system. RESULTS: UROMOL racial subtype distributions were similar with class 2a being most common with 10 genes commonly upregulated in AAs compared to EAs including EFEMP1, S100A16, and MCL1 which are associated with progression to muscle-invasive bladder cancer, mitomycin C resistance, and bacillus Calmette-Guérin durability, respectively. We used single nuclei analysis to map the malignant cell heterogeneity in urothelial cancer which 5 distinct malignant epithelial subtypes whose presence has been associated with different therapeutic response prediction abilities. We mapped the expression of the 10 genes commonly upregulated by race as a function of the 5 malignant subtypes. This showed borderline (P = 0.056) difference among the subtypes suggesting AAs and EAs may be expected to have different therapeutic responses to treatments for bladder cancer. AAs were enriched with immune-related, inflammatory, and cellular regulation pathways compared to EAs, yet appeared to have reduced levels of the aggressive C3/CDH12 bladder tumor cell population. CONCLUSIONS: While premature, gene expression differed between AAs and EAs, supporting potential race-based etiologies for muscle-invasion, response to treatments, and transcriptome pathway regulations.

Cabozantinib plus Nivolumab Phase I Expansion Study in Patients with Metastatic Urothelial Carcinoma Refractory to Immune Checkpoint Inhibitor Therapy.

PURPOSE: This study investigated the efficacy and tolerability of cabozantinib plus nivolumab (CaboNivo) in patients with metastatic urothelial carcinoma (mUC) that progressed on checkpoint inhibition (CPI). PATIENTS AND METHODS: A phase I expansion cohort of patients with mUC who received prior CPI was treated with cabozantinib 40 mg/day and nivolumab 3 mg/kg every 2 weeks until disease progression/unacceptable toxicity. The primary goal was objective response rate (ORR) per RECIST v.1.1. Secondary objectives included progression-free survival (PFS), duration of response (DoR), overall survival (OS), safety, and tolerability. RESULTS: Twenty-nine out of 30 patients enrolled were evaluable for efficacy. Median follow-up was 22.2 months. Most patients (86.7%) received prior chemotherapy and all patients received prior CPI (median seven cycles). ORR was 16.0%, with one complete response and three partial responses (PR). Among 4 responders, 2 were primary refractory, 1 had a PR, and 1 had stable disease on prior CPI. Median DoR was 33.5 months [95% confidence interval (CI), 3.7-33.5], median PFS was 3.6 months (95% CI, 2.1-5.5), and median OS was 10.4 months (95% CI, 5.8-19.5). CaboNivo decreased immunosuppressive subsets such as regulatory T cells (Tregs) and increased potential antitumor immune subsets such as nonclassical monocytes and effector T cells. A lower percentage of monocytic myeloid-derived suppressor cells (M-MDSC) and polymorphonuclear MDSCs, lower CTLA-4 and TIM-3 expression on Tregs, and higher effector CD4+ T cells at baseline were associated with better PFS and/or OS. CONCLUSIONS: CaboNivo was clinically active, well tolerated, and favorably modulated peripheral blood immune subsets in patients with mUC refractory to CPI.

Inhibitory DAMPs in immunogenic cell death and its clinical implications.

Dying (or dead) cells are increasingly recognized to impose significant biological influence within their tissues of residence-exerting paracrine effects through proteins and metabolites that are expressed or secreted during cellular demise. For example, certain molecules function as potent mitogens, promoting the repopulation of neighboring epithelial cells. And other myriad of factors-classified as damage-associated molecular patterns (DAMPs)-function as "find me" (attractant), "eat me" (engulfment), or "danger" (activation) signals for recruiting and activating effector immune cells (e.g., dendritic cells) to initiate inflammation. Since the discovery of immunogenic cell death (ICD), the current dogma posits DAMPs as immunological adjuvants for innate immune cell mobilization and activation, which ultimately leads to the antitumoral cross-priming of CD8+ T cells. However, what is currently unknown is how these immunostimulatory DAMPs are counteracted to avoid immune-overactivation. Our recent work builds on these fundamentals and introduces prostaglandin E2 (PGE2) as an 'inhibitory' DAMP-a new variable to the ICD equation. Prostaglandin E2 functions as an immunosuppressive counterpoise of adjuvant DAMPs; and thus, mechanistically precludes ICD. Furthermore, the long-debated immunogenicity of gemcitabine chemotherapy was revealed to be contingent on inhibitory DAMP blockade and not due to its inability to promote DAMP expression (i.e., calreticulin) as previously reported. These findings were intriguing. First, despite the presence of gemcitabine-induced hallmark DAMPs, the inhibitory DAMP (i.e., PGE2) was sufficient to hinder the ICD-induced antitumoral immune response (Fig. 1a). And second, rather than pharmacologically substantiating immunostimulatory DAMPs as conventionally approached, the mitigation of the inhibitory DAMP-tipping the immunostimulatory and inhibitory DAMP balance in favor of immunostimulatory DAMPs-was sufficient to render the cell death immunogenic and converted gemcitabine into an ICD-inducing therapy (Fig. 1b). In this microreview, we extrapolate our findings and implicate the value of inhibitory DAMP(s) in drug discovery, its use for clinical prognosis, and as target(s) for therapeutic intervention.

Plasminogen activator inhibitor-2 (PAI-2) overexpression supports bladder cancer development in PAI-1 knockout mice in N-butyl-N- (4-hydroxybutyl)-nitrosamine- induced bladder cancer mouse model.

BACKGROUND: Accumulating evidence suggests that plasminogen activator inhibitor-1 (PAI-1) plays an important role in bladder tumorigenesis by regulating cell cycle. However, it remains unclear whether and how inhibition of PAI-1 suppresses bladder tumorigenesis. METHODS: To elucidate the therapeutic effect of PAI-1 inhibition, we tested its tumorigenicity in PAI-1 knockout (KO) mice exposed to a known bladder carcinogen. RESULTS: PAI-1 deficiency did not inhibit carcinogen-induced bladder cancer in mice although carcinogen-exposed wild type mice significantly increased PAI-1 levels in bladder tissue, plasma and urine. We found that PAI-1 KO mice exposed to carcinogen tended to upregulate protein C inhibitor (PAI-3), urokinase-type plasminogen activator (uPA) and tissue-type PA (tPA), and significantly increased PAI-2, suggesting a potential compensatory function of these molecules when PAI-1 is abrogated. Subsequent studies employing gene expression microarray using mouse bladder tissues followed by post hoc bioinformatics analysis and validation experiments by qPCR and IHC demonstrated that SERPING1 is further downregulated in PAI-1 KO mice exposed to BBN, suggesting that SERPING1 as a potential missing factor that regulate PAI-2 overexpression (compensation pathway). CONCLUSIONS: These results indicate that serpin compensation pathway, specifically PAI-2 overexpression in this model, supports bladder cancer development when oncoprotein PAI-1 is deleted. Further investigations into PAI-1 are necessary in order to identify true potential targets for bladder cancer therapy.

A Consensus Molecular Classification of Muscle-invasive Bladder Cancer.

BACKGROUND: Muscle-invasive bladder cancer (MIBC) is a molecularly diverse disease with heterogeneous clinical outcomes. Several molecular classifications have been proposed, but the diversity of their subtype sets impedes their clinical application. OBJECTIVE: To achieve an international consensus on MIBC molecular subtypes that reconciles the published classification schemes. DESIGN, SETTING, AND PARTICIPANTS: We used 1750 MIBC transcriptomic profiles from 16 published datasets and two additional cohorts. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: We performed a network-based analysis of six independent MIBC classification systems to identify a consensus set of molecular classes. Association with survival was assessed using multivariable Cox models. RESULTS AND LIMITATIONS: We report the results of an international effort to reach a consensus on MIBC molecular subtypes. We identified a consensus set of six molecular classes: luminal papillary (24%), luminal nonspecified (8%), luminal unstable (15%), stroma-rich (15%), basal/squamous (35%), and neuroendocrine-like (3%). These consensus classes differ regarding underlying oncogenic mechanisms, infiltration by immune and stromal cells, and histological and clinical characteristics, including outcomes. We provide a single-sample classifier that assigns a consensus class label to a tumor sample's transcriptome. Limitations of the work are retrospective clinical data collection and a lack of complete information regarding patient treatment. CONCLUSIONS: This consensus system offers a robust framework that will enable testing and validation of predictive biomarkers in future prospective clinical trials. PATIENT SUMMARY: Bladder cancers are heterogeneous at the molecular level, and scientists have proposed several classifications into sets of molecular classes. While these classifications may be useful to stratify patients for prognosis or response to treatment, a consensus classification would facilitate the clinical use of molecular classes. Conducted by multidisciplinary expert teams in the field, this study proposes such a consensus and provides a tool for applying the consensus classification in the clinical setting.

Prognostic Significance of Lymphocyte Infiltration and a Stromal Immunostaining of a Bladder Cancer Associated Diagnostic Panel in Urothelial Carcinoma.

We set out to expand on our previous work in which we reported the epithelial expression pattern of a urine-based bladder cancer-associated diagnostic panel (A1AT, ANG, APOE, CA9, IL8, MMP9, MMP10, PAI1, SDC1, and VEGFA). Since many of the analytes in the bladder cancer-associated diagnostic signature were chemokines, cytokines, or secreted proteins, we set out to report the stromal staining pattern of the diagnostic signature as well as CD3+ (T-cell) cell and CD68+ (macrophage) cell staining in human bladder tumors as a snapshot of the tumor immune landscape. Immunohistochemical staining was performed on 213 tumor specimens and 74 benign controls. Images were digitally captured and quantitated using Aperio (Vista, CA). The expression patterns were correlated with tumor grade, tumor stage, and outcome measures. We noted a positive correlation of seven of the 10 proteins (excluding A1AT and IL8 which had a negative association and VEGFA had no association) in bladder cancer. The overexpression of MMP10 was associated with higher grade disease, while overexpression of MMP10, PAI1, SDC1 and ANG were associated with high stage bladder cancer and CA9 was associated with low stage bladder cancer. Increased tumor infiltration of CD68+ cells were associated with higher stage disease. Overall survival was significantly reduced in bladder cancer patients' whose tumors expressed eight or more of the 10 proteins that comprise the bladder cancer diagnostic panel. These findings confirm that the chemokines, cytokines, and secreted proteins in a urine-based diagnostic panel are atypically expressed, not only in the epithelial component of bladder tumors, but also in the stromal component of bladder tumors and portends a worse overall survival. Thus, when assessing immunohistochemical staining, it is important to report staining patterns within the stroma as well as the entire stroma itself.

A fully Bayesian latent variable model for integrative clustering analysis of multi-type omics data.

Identification of clinically relevant tumor subtypes and omics signatures is an important task in cancer translational research for precision medicine. Large-scale genomic profiling studies such as The Cancer Genome Atlas (TCGA) Research Network have generated vast amounts of genomic, transcriptomic, epigenomic, and proteomic data. While these studies have provided great resources for researchers to discover clinically relevant tumor subtypes and driver molecular alterations, there are few computationally efficient methods and tools for integrative clustering analysis of these multi-type omics data. Therefore, the aim of this article is to develop a fully Bayesian latent variable method (called iClusterBayes) that can jointly model omics data of continuous and discrete data types for identification of tumor subtypes and relevant omics features. Specifically, the proposed method uses a few latent variables to capture the inherent structure of multiple omics data sets to achieve joint dimension reduction. As a result, the tumor samples can be clustered in the latent variable space and relevant omics features that drive the sample clustering are identified through Bayesian variable selection. This method significantly improve on the existing integrative clustering method iClusterPlus in terms of statistical inference and computational speed. By analyzing TCGA and simulated data sets, we demonstrate the excellent performance of the proposed method in revealing clinically meaningful tumor subtypes and driver omics features.

Bladder Cancer Molecular Taxonomy: Summary from a Consensus Meeting.

The advent of Omics technologies has been key to the molecular subclassification of urothelial bladder cancer. Several groups have used different strategies to this aim, with partially overlapping findings. The meeting at the Spanish National Cancer Research Center-CNIO was held to discuss such classifications and reach consensus where appropriate. After updated presentations on the work performed by the teams attending the meeting, a consensus was reached regarding the existence of a group of Basal-Squamous-like tumors - designated BASQ - charaterized the high expression of KRT5/6 and KRT14 and low/undetectable expression of FOXA1 and GATA3. An additional tumor subgroup with urothelial differentiation features was recognized whose optimal molecular definition is required. For other subtypes described, more work is needed to determine how robust they are and how to best define them at the molecular level.

RUNX1 is essential for mesenchymal stem cell proliferation and myofibroblast differentiation.

Myofibroblasts are a key cell type in wound repair, cardiovascular disease, and fibrosis and in the tumor-promoting microenvironment. The high accumulation of myofibroblasts in reactive stroma is predictive of the rate of cancer progression in many different tumors, yet the cell types of origin and the mechanisms that regulate proliferation and differentiation are unknown. We report here, for the first time to our knowledge, the characterization of normal human prostate-derived mesenchymal stem cells (MSCs) and the TGF-ß1-regulated pathways that modulate MSC proliferation and myofibroblast differentiation. Human prostate MSCs combined with prostate cancer cells expressing TGF-ß1 resulted in commitment to myofibroblasts. TGF-ß1-regulated runt-related transcription factor 1 (RUNX1) was required for cell cycle progression and proliferation of progenitors. RUNX1 also inhibited, yet did not block, differentiation. Knockdown of RUNX1 in prostate or bone marrow-derived MSCs resulted in cell cycle arrest, attenuated proliferation, and constitutive differentiation to myofibroblasts. These data show that RUNX1 is a key transcription factor for MSC proliferation and cell fate commitment in myofibroblast differentiation. This work also shows that the normal human prostate gland contains tissue-derived MSCs that exhibit multilineage differentiation similar to bone marrow-derived MSCs. Targeting RUNX1 pathways may represent a therapeutic approach to affect myofibroblast proliferation and biology in multiple disease states.

Three differentiation states risk-stratify bladder cancer into distinct subtypes.

Current clinical judgment in bladder cancer (BC) relies primarily on pathological stage and grade. We investigated whether a molecular classification of tumor cell differentiation, based on a developmental biology approach, can provide additional prognostic information. Exploiting large preexisting gene-expression databases, we developed a biologically supervised computational model to predict markers that correspond with BC differentiation. To provide mechanistic insight, we assessed relative tumorigenicity and differentiation potential via xenotransplantation. We then correlated the prognostic utility of the identified markers to outcomes within gene expression and formalin-fixed paraffin-embedded (FFPE) tissue datasets. Our data indicate that BC can be subclassified into three subtypes, on the basis of their differentiation states: basal, intermediate, and differentiated, where only the most primitive tumor cell subpopulation within each subtype is capable of generating xenograft tumors and recapitulating downstream populations. We found that keratin 14 (KRT14) marks the most primitive differentiation state that precedes KRT5 and KRT20 expression. Furthermore, KRT14 expression is consistently associated with worse prognosis in both univariate and multivariate analyses. We identify here three distinct BC subtypes on the basis of their differentiation states, each harboring a unique tumor-initiating population.

Intraoperative fracture of phacoemulsification sleeve.

BACKGROUND: We describe a case of intraoperative fracture of phacoemulsification sleeve during phacoemulsification surgery. CASE PRESENTATION: Phacoemulsification surgery was performed in the left eye of a 58-year-old lady with grade II nuclear sclerosis & grade I cortical cataract. Towards the end of quadrant removal, there was anterior chamber instability with impaired followability of nuclear fragments. The distal part of the fractured sleeve remained inside the anterior chamber upon removal of the phacoemulsification probe. The retained sleeve was retrieved with a pair of forceps through the corneal incision site, which did not require widening. There was no missing fragments retained intraocularly and the patient had an uneventful recovery with vision of 20/25 at three months post-operatively. CONCLUSION: Phacoemulsification sleeve fracture is an uncommon complication. With early identification of this condition and proper management, major complications can be avoided.

Imaging of STAT3 signaling pathway during mouse embryonic stem cell differentiation.

Signal transducers and activators of transcription 3 (STAT3) is a pleiotropic transcription factor involved in a variety of physiological processes. STAT3 acts as a key transcriptional determinant of mouse embryonic stem (ES) cell self-renewal and plays a pivotal function in early mammalian embryogenesis because the development of many organs requires STAT3 activation. However, little is known about the role of STAT3 function during ES cell differentiation. To answer this question, we built a lentiviral construct with 7-repeat STAT3-binding sequence (enhancer) and minimal TA (promoter) driving renilla luciferase and monomeric red fluorescence protein (Rluc-mRFP), followed by a constitutive cytomegalovirus promoter driving green fluorescent protein as a selection marker. The specificity of our custom-designed 7-repeat STAT3 reporter construct was first confirmed by cotransfection with constitutively active version of STAT3 (STAT3C) into human embryonic kidney 293T cells. Next, a mouse ES cell line stably transduced with STAT3 reporter construct was isolated. This ES cell line showed a tight response in reporter gene expression with leukemia inhibitory factor (LIF) induction and was chosen as a developmental model for the STAT3 functional study. Using serial noninvasive bioluminescence imaging, we showed that the onset of embryoid body (EB) formation involved inhibition of STAT3 activity. However, during differentiation, STAT3 activity steadily increased from day 5 to 14 and was reduced by day 21. STAT3 activity was also confirmed separately by Western blots. Finally, phosphorylation of STAT3 was also found to correspond with cardiomyocyte differentiation. In summary, this is the first study to monitor real-time STAT3 activity during ES cell differentiation. This genetically modified line can be used to study the biological role of STAT3 during ES cell differentiation into different derivatives.

Signal transducer and activator of transcription 3 (Stat3) in epithelial carcinogenesis.

Signal transducer and activator of transcription 3 (Stat3) is one of a family of cytoplasmic proteins that participate in normal cellular responses to cytokines and growth factors as transcription factors. Stat3 modulates various physiological functions including cell survival, cell-cycle regulation, and angiogenesis through regulation of gene expression, and its constitutive activation is associated with a number of human epithelial cancers. Recent studies with skin-specific gain and loss of Stat3 function transgenic mice have shown that Stat3 plays critical roles in skin carcinogenesis. Multistage skin carcinogenesis bioassays performed with these transgenic mice clearly demonstrate that Stat3 is required for both tumor initiation and promotion through regulation of genes involved in survival and proliferation, respectively. Stat3 also plays a role in malignant progression of skin tumors by regulating genes that are involved in angiogenesis and invasion. Further studies have revealed that Stat3 plays a critical role in epidermal cell proliferation and survival following exposure to ultraviolet B (UVB) irradiation. In addition, Stat3 is constitutively active in UVB-induced skin tumors from both mice and humans. Collectively, these studies suggest that Stat3 may be a potential target for both the prevention and treatment of human epithelial cancers including skin cancer.

Evaluation of quality of life in patients with cataract in Hong Kong.

PURPOSE: To evaluate the quality of life in patients in a public hospital in Hong Kong before and after cataract surgery using a new questionnaire. SETTING: Department of Ophthalmology, Tung Wah Eastern Hospital, Hong Kong, China. METHODS: One hundred ten Chinese patients having cataract extraction were evaluated for surgical outcomes and quality of life using a questionnaire modified to reflect the local culture and environment in Hong Kong. The questionnaire consisted of 20 questions divided into 4 domains: distance vision, near vision, social function, and cataract-related symptoms. The test-retest reliability of the questionnaire was assessed in another 15 patients by weighted kappa and internal consistency tested with the Cronbach alpha. Construct validity was tested by correlating visual acuity with quality of life and subjective visual improvement and satisfaction with quality-of-life improvement. The preoperative and 4-month postoperative quality of life were evaluated, and the association with the patients' characteristics, type of surgery performed, presence of systemic or ocular diseases, and subjective patient satisfaction was analyzed. RESULTS: The preoperative visual acuity in the operated eye had a low correlation (0.11) with the quality-of-life score; visual acuity in the better eye had a higher correlation (0.29). Quality-of-life improvement was moderately correlated with visual acuity improvement and patient satisfaction. Overall, quality-of-life scoring improved in all domains after surgery, with an overall effect size of 0.68. Quality-of-life scores improved postoperatively in 83.6% of patients, did not change in 3.6%, and were worse in 12.7%. Visual acuity improved in 94.5%, remained the same in 2.7%, and was worse in 2.7%. There was no difference in quality-of-life improvement between extracapsular cataract extraction and phacoemulsification at 4 months. Patients with systemic or ocular diseases and those who were phakic in the fellow eye had lower quality-of-life improvement; however, this was statistically significant for systemic diseases only. Age and sex did not affect quality of life or its improvement. CONCLUSIONS: The quality of life and surgical outcomes in cataract patients were assessed by a simple questionnaire. The local culture and environment should be taken into consideration in the questionnaire design.