Stromal Rigidity Stress Accelerates Pancreatic Intraepithelial Neoplasia Progression and Chromosomal Instability via Nuclear Protein Tyrosine Kinase 2 Localization.

Because the mechanotransduction by stromal stiffness stimulates the rupture and repair of the nuclear envelope in pancreatic progenitor cells, accumulated genomic aberrations are under selection in the tumor microenvironment. Analysis of cell growth, micronuclei, and phosphorylated Ser-139 residue of the histone variant H2AX (γH2AX) foci linked to mechanotransduction pressure in vivo during serial orthotopic passages of mouse KrasLSL-G12D/+;Trp53flox/flox;Pdx1-Cre (KPC) cancer cells in the tumor and in migrating through the size-restricted 3-µm micropores. To search for pancreatic cancer cell-of-origin, analysis of single-cell data sets revealed that the extracellular matrix shaped an alternate route of acinar-ductal transdifferentiation of acinar cells into topoisomerase II α (TOP2A)-overexpressing cancer cells and derived subclusters with copy number amplifications in MYC-PTK2 (protein tyrosine kinase 2) locus and PIK3CA. High-PTK2 expression is associated with 171 differentially methylated CpG loci, 319 differentially expressed genes, and poor overall survival in The Cancer Genome Atlas-Pancreatic Adenocarcinoma cohort. Abolished RGD-integrin signaling by disintegrin KG blocked the PTK2 phosphorylation, increased cancer apoptosis, decreased vav guanine nucleotide exchange factor 1 (VAV1) expression, and prolonged overall survival in the KPC mice. Reduction of α-smooth muscle actin deposition in the CD248 knockout KPC mice remodeled the tissue stroma and down-regulated TOP2A expression in the epithelium. In summary, stromal stiffness induced the onset of cancer cells-of-origin by ectopic TOP2A expression, and the genomic amplification of MYC-PTK2 locus via alternative transdifferentiation of pancreatic progenitor cells is the vulnerability useful for disintegrin KG treatment.

Oral bacteria accelerate pancreatic cancer development in mice.

OBJECTIVE: Epidemiological studies highlight an association between pancreatic ductal adenocarcinoma (PDAC) and oral carriage of the anaerobic bacterium Porphyromonas gingivalis, a species highly linked to periodontal disease. We analysed the potential for P. gingivalis to promote pancreatic cancer development in an animal model and probed underlying mechanisms. DESIGN: We tracked P. gingivalis bacterial translocation from the oral cavity to the pancreas following administration to mice. To dissect the role of P. gingivalis in PDAC development, we administered bacteria to a genetically engineered mouse PDAC model consisting of inducible acinar cell expression of mutant Kras (Kras +/LSL-G12D; Ptf1a-CreER, iKC mice). These mice were used to study the cooperative effects of Kras mutation and P. gingivalis on the progression of pancreatic intraepithelial neoplasia (PanIN) to PDAC. The direct effects of P. gingivalis on acinar cells and PDAC cell lines were studied in vitro. RESULTS: P. gingivalis migrated from the oral cavity to the pancreas in mice and can be detected in human PanIN lesions. Repetitive P. gingivalis administration to wild-type mice induced pancreatic acinar-to-ductal metaplasia (ADM), and altered the composition of the intrapancreatic microbiome. In iKC mice, P. gingivalis accelerated PanIN to PDAC progression. In vitro, P. gingivalis infection induced acinar cell ADM markers SOX9 and CK19, and intracellular bacteria protected PDAC cells from reactive oxygen species-mediated cell death resulting from nutrient stress. CONCLUSION: Taken together, our findings demonstrate a causal role for P. gingivalis in pancreatic cancer development in mice.

NEMO/NF-κB signaling functions as a double-edged sword in PanIN formation versus progression to pancreatic cancer.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is marked by a dismal survival rate, lacking effective therapeutics due to its aggressive growth, late-stage diagnosis, and chemotherapy resistance. Despite debates on NF-κB targeting for PDAC treatment, no successful approach has emerged. METHODS: To elucidate the role of NF-κB, we ablated NF-κB essential modulator (NEMO), critical for conventional NF-κB signaling, in the pancreata of mice that develop precancerous lesions (KC mouse model). Secretagogue-induced pancreatitis by cerulein injections was utilized to promote inflammation and accelerate PDAC development. RESULTS: NEMO deletion reduced fibrosis and inflammation in young KC mice, resulting in fewer pancreatic intraepithelial neoplasias (PanINs) at later stages. Paradoxically, however, NEMO deletion accelerated the progression of these fewer PanINs to PDAC and reduced median lifespan. Further, analysis of tissue microarrays from human PDAC sections highlighted the correlation between reduced NEMO expression in neoplastic cells and poorer prognosis, supporting our observation in mice. Mechanistically, NEMO deletion impeded oncogene-induced senescence (OIS), which is normally active in low-grade PanINs. This blockage resulted in fewer senescence-associated secretory phenotype (SASP) factors, reducing inflammation. However, blocked OIS fostered replication stress and DNA damage accumulation which accelerated PanIN progression to PDAC. Finally, treatment with the DNA damage-inducing reagent etoposide resulted in elevated cell death in NEMO-ablated PDAC cells compared to their NEMO-competent counterparts, indicative of a synthetic lethality paradigm. CONCLUSIONS: NEMO exhibited both oncogenic and tumor-suppressive properties during PDAC development. Caution is suggested in therapeutic interventions targeting NF-κB, which may be detrimental during PanIN progression but beneficial post-PDAC development.

The karyometric signature is altered in fallopian tubes with serous tubal intraepithelial carcinoma.

OBJECTIVE: Recent evidence suggests that the fimbriated end of the fallopian tube harbors the precursor cells for many high-grade ovarian cancers, opening the door for development of better screening methods that directly assess the fallopian tube in women at risk for malignancy. Previously we have shown that the karyometric signature is abnormal in the fallopian tube epithelium in women at hereditary risk of ovarian cancer. In this study, we sought to determine whether the karyometric signature in serous tubal intraepithelial carcinoma (STIC) is significantly different from normal, and whether an abnormal karyometric signature can be detected in histologically normal tubal epithelial cells adjacent to STIC lesions. METHODS: The karyometric signature was measured in epithelial cells from the proximal and fimbriated portion of the fallopian tube in fallopian tube specimens removed from women at: 1) average risk for ovarian cancer undergoing surgery for benign gynecologic indications (n = 37), 2) hereditary risk of ovarian cancer (germline BRCA alterations) undergoing risk-reducing surgery (n = 44), and 3) diagnosed with fimbrial STICs (n = 17). RESULTS: The karyometric signature in tubes with fimbrial STICs differed from that of tubes with benign histology. The degree of karyometric alteration increased with increasing proximity to fimbrial STICs, ranging from moderate in the proximal portion of the tube, to greatest in both normal appearing fimbrial cells near STICs as well as in fimbrial STIC lesions. CONCLUSION: These data demonstrate an abnormal karyometric signature in STICs that may extend beyond the STIC, potentially providing an opportunity for early detection of fallopian tube neoplasia.

Co-existing TP53 and ARID1A mutations promote aggressive endometrial tumorigenesis.

TP53 and ARID1A are frequently mutated across cancer but rarely in the same primary tumor. Endometrial cancer has the highest TP53-ARID1A mutual exclusivity rate. However, the functional relationship between TP53 and ARID1A mutations in the endometrium has not been elucidated. We used genetically engineered mice and in vivo genomic approaches to discern both unique and overlapping roles of TP53 and ARID1A in the endometrium. TP53 loss with oncogenic PIK3CAH1047R in the endometrial epithelium results in features of endometrial hyperplasia, adenocarcinoma, and intraepithelial carcinoma. Mutant endometrial epithelial cells were transcriptome profiled and compared to control cells and ARID1A/PIK3CA mutant endometrium. In the context of either TP53 or ARID1A loss, PIK3CA mutant endometrium exhibited inflammatory pathway activation, but other gene expression programs differed based on TP53 or ARID1A status, such as epithelial-to-mesenchymal transition. Gene expression patterns observed in the genetic mouse models are reflective of human tumors with each respective genetic alteration. Consistent with TP53-ARID1A mutual exclusivity, the p53 pathway is activated following ARID1A loss in the endometrial epithelium, where ARID1A normally directly represses p53 pathway genes in vivo, including the stress-inducible transcription factor, ATF3. However, co-existing TP53-ARID1A mutations led to invasive adenocarcinoma associated with mutant ARID1A-driven ATF3 induction, reduced apoptosis, TP63+ squamous differentiation and invasion. These data suggest TP53 and ARID1A mutations drive shared and distinct tumorigenic programs in the endometrium and promote invasive endometrial cancer when existing simultaneously. Hence, TP53 and ARID1A mutations may co-occur in a subset of aggressive or metastatic endometrial cancers, with ARID1A loss promoting squamous differentiation and the acquisition of invasive properties.

[Development and validation of predictive models for esophageal squamous cell carcinoma and its precancerous lesions using terminal motif analysis in circulating cell-free DNA].

Objectives: To develop and validate predictive models for esophageal squamous cell carcinoma (ESCC) using circulating cell-free DNA (cfDNA) terminal motif analysis. The goal was to improve the non-invasive detection of early-stage ESCC and its precancerous lesions. Methods: Between August 2021 and November 2022, we prospectively collected plasma samples from 448 individuals at the Department of Endoscopy, Cancer Hospital, Chinese Academy of Medical Sciences for cfDNA extraction, library construction, and sequencing. We analyzed 201 cases of ESCC, 46 high-grade intraepithelial neoplasia (HGIN), 46 low-grade intraepithelial neoplasia (LGIN), 176 benign esophageal lesions, and 29 healthy controls. Participants, including ESCC patients and control subjects, were randomly assigned to a training set (n=284) and a validation set (n=122). The training cohort underwent z-score normalization of cfDNA terminal motif matrices and a selection of distinctive features differentiated ESCC cases from controls. The random forest classifier, Motif-1 (M1), was then developed through principal component analysis, ten-fold cross-validation, and recursive feature elimination. M1's efficacy was then validated in the validation and precancerous lesion sets. Subsequently, individuals with precancerous lesions were included in the dataset and participants were randomly allocated to newly formed training (n=243), validation (n=105), and test (n=150) cohorts. Using the same procedure as M1, we trained the Motif-2 (M2) random forest model with the training cohort. The M2 model's accuracy was then confirmed in the validation cohort to establish the optimal threshold and further tested by performing validation in the test cohort. Results: We developed two cfDNA terminal motif-based predictive models for ESCC and associated precancerous conditions. The first model, M1, achieved a sensitivity of 90.0%, a specificity of 77.4%, and an area under the curve (AUC) of 0.884 in the validation cohort. For LGIN, HGIN, and T1aN0 stage ESCC, M1's sensitivities were 76.1%, 80.4%, and 91.2% respectively. Notably, the sensitivity for jointly predicting HGIN and T1aN0 ESCC reached 85.0%. Both the predictive accuracy and sensitivity increased in line with the cancer's progression (P＜0.001). The second model, M2, exhibited a sensitivity of 87.5%, a specificity of 77.4%, and an AUC of 0.857 in the test cohort. M2's sensitivities for detecting precancerous lesions and ESCC were 80.0% and 89.7%, respectively, and it showed a combined sensitivity of 89.4% for HGIN and T1aN0 stage ESCC. Conclusions: Two predictive models based on cfDNA terminal motif analysis for ESCC and its precancerous lesions are developed. They both show high sensitivity and specificity in identifying ESCC and its precancerous stages, indicating its potential for early ESCC detection.

Clinical validation of methylation biomarkers for optimal detection of high-grade vulvar intraepithelial neoplasia.

The precursor lesions of vulvar squamous cell carcinoma (VSCC) include human papillomavirus (HPV)-associated and HPV-independent squamous neoplasia with a varying cancer risk. Our study aimed to validate the accuracy of previously identified DNA methylation markers for detection of such high-grade vulvar intraepithelial neoplasia (VIN). A large clinical series of 751 vulvar lesions, originally diagnosed as high-grade VIN, were reassessed and categorized into HPV-associated or HPV-independent vulvar disease categories. Together with 113 healthy vulvar controls, all samples were tested for 12 methylation markers with quantitative multiplex methylation-specific PCR (qMSP). Performance of individual markers and selection of an optimal marker panel for detection of high-grade VIN was determined by logistic regression analysis. SST was the best-performing individual marker (AUC 0.90), detecting 80% of high-grade VIN cases, with excellent detection of HPV-independent VIN (95%), known to have the highest cancer risk. Merely 2% of controls tested methylation positive for SST. Selection of a marker panel, including ZNF582, SST and miR124-2, resulted in a comparably high accuracy for detection of high-grade VIN (AUC 0.89). In conclusion, we clinically validated the accuracy of 12 DNA methylation markers for detection of high-grade VIN. SST, as a sole marker or in a panel, provides an optimal diagnostic tool to distinguish high-grade VIN in need of treatment, particularly HPV-independent VIN, from low-grade or reactive vulvar lesions. These findings warrant further prognostic validation of methylation biomarkers for cancer risk stratification of patients with VIN.

Serous Tubal Intraepithelial Carcinoma in a Risk-reducing Salpingo-oophorectomy Specimen From a RAD51D Mutation Carrier: A Case Report.

The RAD51D gene codes a protein-paralog of the RAD51 DNA recombinase, which catalyzes DNA strand exchange during homologous recombination. Similar to BRCA1 / BRCA2 , mutations in RAD51D both predispose to ovarian carcinoma and impart sensitivity to poly (ADP-ribose) polymerase inhibitors in preclinical studies. Based on cancer risk prediction models, RAD51D mutations pose a moderate-to-high risk for ovarian cancer warranting consideration for risk-reducing surgery. We report a case of serous tubal intraepithelial carcinoma in a patient undergoing risk-reducing total hysterectomy with bilateral salpingo-oophorectomy for a RAD51D pathogenic variant. The histopathologic and p53-immunophenotypic features of this lesion are similar to those reported previously in BRCA1 / BRCA2 mutation carriers and those of serous tubal intraepithelial carcinoma associated with sporadic high-grade serous carcinomas. These features include marked increase in nuclear-to-cytoplasmic ratio, loss of cell polarity, absence of ciliation, prominent nucleoli, mitotic activity, epithelial stratification, surface exfoliative changes, and complete loss of p53 staining. Although familial ovarian cancers with mutations in RAD51D -or other genes in the Fanconi anemia pathway-are much less common those with BRCA1 / BRCA2 mutations, our findings support a common phenotype for early serous cancers in this pathway.

Acinar-to-Ductal Metaplasia (ADM): On the Road to Pancreatic Intraepithelial Neoplasia (PanIN) and Pancreatic Cancer.

Adult pancreatic acinar cells show high plasticity allowing them to change in their differentiation commitment. Pancreatic acinar-to-ductal metaplasia (ADM) is a cellular process in which the differentiated pancreatic acinar cells transform into duct-like cells. This process can occur as a result of cellular injury or inflammation in the pancreas. While ADM is a reversible process allowing pancreatic acinar regeneration, persistent inflammation or injury can lead to the development of pancreatic intraepithelial neoplasia (PanIN), which is a common precancerous lesion that precedes pancreatic ductal adenocarcinoma (PDAC). Several factors can contribute to the development of ADM and PanIN, including environmental factors such as obesity, chronic inflammation and genetic mutations. ADM is driven by extrinsic and intrinsic signaling. Here, we review the current knowledge on the cellular and molecular biology of ADM. Understanding the cellular and molecular mechanisms underlying ADM is critical for the development of new therapeutic strategies for pancreatitis and PDAC. Identifying the intermediate states and key molecules that regulate ADM initiation, maintenance and progression may help the development of novel preventive strategies for PDAC.

Target-Specific Nanoparticle Polyplex Down-Regulates Mutant Kras to Prevent Pancreatic Carcinogenesis and Halt Tumor Progression.

Survival from pancreatic cancer is poor because most cancers are diagnosed in the late stages and there are no therapies to prevent the progression of precancerous pancreatic intraepithelial neoplasms (PanINs). Inhibiting mutant KRASG12D, the primary driver mutation in most human pancreatic cancers, has been challenging. The cholecystokinin-B receptor (CCK-BR) is absent in the normal pancreas but becomes expressed in high grade PanIN lesions and is over-expressed in pancreatic cancer making it a prime target for therapy. We developed a biodegradable nanoparticle polyplex (NP) that binds selectively to the CCK-BR on PanINs and pancreatic cancer to deliver gene therapy. PanIN progression was halted and the pancreas extracellular matrix rendered less carcinogenic in P48-Cre/LSL-KrasG12D/+ mice treated with the CCK-BR targeted NP loaded with siRNA to mutant Kras. The targeted NP also slowed proliferation, decreased metastases and improved survival in mice bearing large orthotopic pancreatic tumors. Safety and toxicity studies were performed in immune competent mice after short or long-term exposure and showed no off-target toxicity by histological or biochemical evaluation. Precision therapy with target-specific NPs provides a novel approach to slow progression of advanced pancreatic cancer and also prevents the development of pancreatic cancer in high-risk subjects without toxicity to other tissues.

Epigenetic Alterations in DCIS Progression: What Can lncRNAs Teach Us?

Some transcripts that are not translated into proteins can be encoded by the mammalian genome. Long noncoding RNAs (lncRNAs) are noncoding RNAs that can function as decoys, scaffolds, and enhancer RNAs and can regulate other molecules, including microRNAs. Therefore, it is essential that we obtain a better understanding of the regulatory mechanisms of lncRNAs. In cancer, lncRNAs function through several mechanisms, including important biological pathways, and the abnormal expression of lncRNAs contributes to breast cancer (BC) initiation and progression. BC is the most common type of cancer among women worldwide and has a high mortality rate. Genetic and epigenetic alterations that can be regulated by lncRNAs may be related to early events of BC progression. Ductal carcinoma in situ (DCIS) is a noninvasive BC that is considered an important preinvasive BC early event because it can progress to invasive BC. Therefore, the identification of predictive biomarkers of DCIS-invasive BC progression has become increasingly important in an attempt to optimize the treatment and quality of life of patients. In this context, this review will address the current knowledge about the role of lncRNAs in DCIS and their potential contribution to the progression of DCIS to invasive BC.

AGR2-Dependent Nuclear Import of RNA Polymerase II Constitutes a Specific Target of Pancreatic Ductal Adenocarcinoma in the Context of Wild-Type p53.

BACKGROUND & AIMS: Promoted by pancreatitis, oncogenic KrasG12D triggers acinar cells' neoplastic transformation through acinar-to-ductal metaplasia (ADM) and pancreatic intraepithelial neoplasia. Anterior gradient 2 (Agr2), a known inhibitor of p53, is detected at early stage of pancreatic ductal adenocarcinoma (PDAC) development. RNA polymerase II (RNAPII) is a key nuclear enzyme; regulation of its nuclear localization in mammalian cells represents a potential therapeutic target. METHODS: A mouse model of inflammation-accelerated KrasG12D-driven ADM and pancreatic intraepithelial neoplasia development was used. Pancreas-specific Agr2 ablation was performed to access its role in pancreatic carcinogenesis. Hydrophobic hexapeptides loaded in liposomes were developed to disrupt Agr2-RNAPII complex. RESULTS: We found that Agr2 is up-regulated in ADM-to-pancreatic intraepithelial neoplasia transition in inflammation and KrasG12D-driven early pancreatic carcinogenesis. Genetic ablation of Agr2 specifically blocks this metaplastic-to-neoplastic process. Mechanistically, Agr2 directs the nuclear import of RNAPII via its C-terminal nuclear localization signal, undermining the ATR-dependent p53 activation in ADM lesions. Because Agr2 binds to the largest subunit of RNAPII in a peptide motif-dependent manner, we developed a hexapeptide to interfere with the nuclear import of RNAPII by competitively disrupting the Agr2-RNAPII complex. This novel hexapeptide leads to dysfunction of RNAPII with concomitant activation of DNA damage response in early neoplastic lesions; hence, it dramatically compromises PDAC initiation in vivo. Moreover, the hexapeptide sensitizes PDAC cells and patient-derived organoids harboring wild-type p53 to RNAPII inhibitors and first-line chemotherapeutic agents in vivo. Of note, this therapeutic effect is efficient across various cancer types. CONCLUSIONS: Agr2 is identified as a novel adaptor protein for nuclear import of RNAPII in mammalian cells. Also, we provide genetic evidence defining Agr2-dependent nuclear import of RNAPII as a pharmaceutically accessible target for prevention and treatment in PDAC in the context of wild-type p53.

p53 null phenotype is a "positive result" in urothelial carcinoma in situ.

The concept of a "p53 null phenotype" (complete loss of staining) is well-recognized in the gynecologic pathology literature, implicitly reflecting that this staining pattern represents a TP53 mutation. However, in the genitourinary pathology literature, a p53 null phenotype has only been addressed regarding the prognosis of invasive urothelial carcinoma, and not as a diagnostic biomarker for urothelial carcinoma in situ (CIS). Herein, 25 cases of urothelial carcinoma in situ [diagnoses made on hematoxylin and eosin (H&E) stained sections] showing null pattern p53 staining were retrieved from 22 different patients (16 males and 6 females, age range 52-85 years; average 69.6 years), most commonly showing large cell pleomorphic pattern morphology. One representative tissue block per case was selected for next-generation DNA sequencing (NGS). All 21 cases (100%) passing quality control for NGS showed at least 1 TP53 mutation (majority nonsense or frameshift mutations), including 3 cases with 2 mutations and 3 cases with 3 mutations. Three patients with multiple available samples harbored 1 or more shared TP53 mutations at 2 different time points, indicating clonality of the temporally distinct lesions. Additionally, 2 patients had an additional unique TP53 mutation at a later time point, suggesting intratumoral heterogeneity and/or temporal clonal evolution. While urothelial CIS remains an H&E diagnosis in most cases, a p53 immunostain may be useful in a subset of challenging cases. This study demonstrates that a p53 null phenotype represents an aberrant result in urothelial CIS with supportive molecular analysis showing a previously unknown level of complexity for TP53 mutations among these noninvasive lesions. Adequate recognition of the p53 null phenotype as a "biologically supportive result", similar to strong and diffuse staining with p53, is important and may warrant a formal consensus statement for recommended p53 reporting (i.e., "wild type" versus "aberrant or mutant").

The Origin of Ovarian Cancer Species and Precancerous Landscape.

Unlike other human cancers, in which all primary tumors arise de novo, ovarian epithelial cancers are primarily imported from either endometrial or fallopian tube epithelium. The prevailing paradigm in the genesis of high-grade serous carcinoma (HGSC), the most common ovarian cancer, posits to its development in fallopian tubes through stepwise tumor progression. Recent progress has been made not only in gathering terabytes of omics data but also in detailing the histologic-molecular correlations required for looking into, and making sense of, the tissue origin of HGSC. This emerging paradigm is changing many facets of ovarian cancer research and routine gynecology practice. The precancerous landscape in fallopian tubes contains multiple concurrent precursor lesions, including serous tubal intraepithelial carcinoma (STIC), with genetic heterogeneity providing a platform for HGSC evolution. Mathematical models imply that a prolonged time (decades) elapses from the development of a TP53 mutation, the earliest known molecular alteration, to an STIC, followed by a shorter span (6 years) for progression to an HGSC. Genetic predisposition accelerates the trajectory. This timeline may allow for the early diagnosis of HGSC and STIC, followed by intent-to-cure surgery. This review discusses the recent advances in this tubal paradigm and its biological and clinical implications, alongside the promise and challenge of studying STIC and other precancerous lesions of HGSC.

Incidence, clinicopathological features and genetics of in-situ follicular neoplasia: a comprehensive screening study in a Japanese cohort.

AIMS: In-situ follicular neoplasia (ISFN) is a histologically recognizable neoplastic proliferation of follicular lymphoma (FL)-like B cells confined to the germinal centres. While some ISFNs are associated with overt FL, others are incidentally identified as isolated or pure forms in individuals without evidence of overt FL. The prevalence of incidentally found isolated ISFN is approximately 3% in Europe; however, no screening study has been conducted in Asia. To investigate the incidence and clinicopathological characteristics of ISFNs in the Japanese population, we conducted histopathological screening of the lymph nodes (LNs) resected for solid tumours or inflammatory conditions. METHODS AND RESULTS: We screened for ISFN in 5700 LNs from 340 individuals using immunohistochemistry for BCL2 and identified seven ISFNs, with an incidence of 2.1%. The median age of the individuals with ISFN was 67 years, none of whom developed overt FL, with a median follow-up of 59 months. Next-generation sequencing was performed in five ISFNs, and 10 variants in seven FL-associated genes were identified. The identified variants included HIST1H1E (n = 2), ARID1A (n = 2), KMT2D (n = 1), CARD11 (n = 1), BCL7A (n = 1), CREBBP (n = 1) and TNFRSF14 (n = 1). CONCLUSIONS: The incidence of isolated ISFN in the Japanese population is not significantly different from that in Europe, presumably reflecting the recent increase in FL in Japan. These incidentally found ISFNs have a low potential to transform into overt FL. Although mutations of FL-associated genes are already present in ISFNs, further molecular studies are needed to identify driver genes leading to the transformation of ISFN to overt FL.

A renewed model of pancreatic cancer evolution based on genomic rearrangement patterns.

Pancreatic cancer, a highly aggressive tumour type with uniformly poor prognosis, exemplifies the classically held view of stepwise cancer development. The current model of tumorigenesis, based on analyses of precursor lesions, termed pancreatic intraepithelial neoplasm (PanINs) lesions, makes two predictions: first, that pancreatic cancer develops through a particular sequence of genetic alterations (KRAS, followed by CDKN2A, then TP53 and SMAD4); and second, that the evolutionary trajectory of pancreatic cancer progression is gradual because each alteration is acquired independently. A shortcoming of this model is that clonally expanded precursor lesions do not always belong to the tumour lineage, indicating that the evolutionary trajectory of the tumour lineage and precursor lesions can be divergent. This prevailing model of tumorigenesis has contributed to the clinical notion that pancreatic cancer evolves slowly and presents at a late stage. However, the propensity for this disease to rapidly metastasize and the inability to improve patient outcomes, despite efforts aimed at early detection, suggest that pancreatic cancer progression is not gradual. Here, using newly developed informatics tools, we tracked changes in DNA copy number and their associated rearrangements in tumour-enriched genomes and found that pancreatic cancer tumorigenesis is neither gradual nor follows the accepted mutation order. Two-thirds of tumours harbour complex rearrangement patterns associated with mitotic errors, consistent with punctuated equilibrium as the principal evolutionary trajectory. In a subset of cases, the consequence of such errors is the simultaneous, rather than sequential, knockout of canonical preneoplastic genetic drivers that are likely to set-off invasive cancer growth. These findings challenge the current progression model of pancreatic cancer and provide insights into the mutational processes that give rise to these aggressive tumours.

Image-based detection and targeting of therapy resistance in pancreatic adenocarcinoma.

Pancreatic intraepithelial neoplasia is a pre-malignant lesion that can progress to pancreatic ductal adenocarcinoma, a highly lethal malignancy marked by its late stage at clinical presentation and profound drug resistance. The genomic alterations that commonly occur in pancreatic cancer include activation of KRAS2 and inactivation of p53 and SMAD4 (refs 2-4). So far, however, it has been challenging to target these pathways therapeutically; thus the search for other key mediators of pancreatic cancer growth remains an important endeavour. Here we show that the stem cell determinant Musashi (Msi) is a critical element of pancreatic cancer progression both in genetic models and in patient-derived xenografts. Specifically, we developed Msi reporter mice that allowed image-based tracking of stem cell signals within cancers, revealing that Msi expression rises as pancreatic intraepithelial neoplasia progresses to adenocarcinoma, and that Msi-expressing cells are key drivers of pancreatic cancer: they preferentially harbour the capacity to propagate adenocarcinoma, are enriched in circulating tumour cells, and are markedly drug resistant. This population could be effectively targeted by deletion of either Msi1 or Msi2, which led to a striking defect in the progression of pancreatic intraepithelial neoplasia to adenocarcinoma and an improvement in overall survival. Msi inhibition also blocked the growth of primary patient-derived tumours, suggesting that this signal is required for human disease. To define the translational potential of this work we developed antisense oligonucleotides against Msi; these showed reliable tumour penetration, uptake and target inhibition, and effectively blocked pancreatic cancer growth. Collectively, these studies highlight Msi reporters as a unique tool to identify therapy resistance, and define Msi signalling as a central regulator of pancreatic cancer.

Outcome and Management of Serous Tubal Intraepithelial Carcinoma Following Opportunistic Salpingectomy: Systematic Review and Meta-Analysis.

OBJECTIVE: Serous ovarian cancer is the most common subtype of epithelial ovarian carcinoma-the most prevalent type of ovarian cancer. High-grade serous ovarian carcinoma (HGSOC) is thought to arise from the distal fallopian tube, with a precursor lesion known as serous tubal intraepithelial carcinoma (STIC). STICs are found in the final pathology of a salpingectomy specimen in 10%-20% of women with a BRCA gene mutation and 1%-7% of women without a mutation. However, there is currently no official guideline and a paucity of data on the management of STICs. DATA SOURCES: We performed a systematic review following PRISMA guidelines. Five databases were searched for relevant studies on STICs. STUDY SELECTION: Two independent reviewers performed the abstract and full-text screening and data extraction, with conflicts resolved through discussion with the third reviewer. The risk of bias of each study was assessed using the Newcastle-Ottawa scale. DATA EXTRACTION AND SYNTHESIS: Fourteen articles were included. Ninety-nine patients who were diagnosed with STIC and subsequently followed for a mean period of 55.5 months were included in this analysis. Eighty-three patients (83.9%) were BRCA mutation carriers. After the diagnosis of isolated STIC, 7 patients (7.3%) received chemotherapy and 25 (26%) underwent surgical staging. Three of the 25 patients were diagnosed with HGSOC based on the staging surgery. Nine patients were later diagnosed with HGSOC during follow-up, with an average duration of follow-up of 58.5 months between the diagnosis of STIC and the diagnosis of HGSOC. CONCLUSION: Based on our review of the literature, there is a 10.7% risk of having concurrent HGSOC at the time of STIC diagnosis, and the risk of developing a subsequent HGSOC is 14.5%. BRCA mutation status should be determined in cases of isolated STIC, as 83.9% of patients included in this study were found to carry BRCA mutations. We believe it is necessary to further investigate the role of surgical staging following the diagnosis of STIC.

Intravesical nadofaragene firadenovec gene therapy for BCG-unresponsive non-muscle-invasive bladder cancer: a single-arm, open-label, repeat-dose clinical trial.

BACKGROUND: BCG is the most effective therapy for high-risk non-muscle-invasive bladder cancer. Nadofaragene firadenovec (also known as rAd-IFNa/Syn3) is a replication-deficient recombinant adenovirus that delivers human interferon alfa-2b cDNA into the bladder epithelium, and a novel intravesical therapy for BCG-unresponsive non-muscle-invasive bladder cancer. We aimed to evaluate its efficacy in patients with BCG-unresponsive non-muscle-invasive bladder cancer. METHODS: In this phase 3, multicentre, open-label, repeat-dose study done in 33 centres (hospitals and clinics) in the USA, we recruited patients aged 18 years or older, with BCG-unresponsive non-muscle-invasive bladder cancer and an Eastern Cooperative Oncology Group status of 2 or less. Patients were excluded if they had upper urinary tract disease, urothelial carcinoma within the prostatic urethra, lymphovascular invasion, micropapillary disease, or hydronephrosis. Eligible patients received a single intravesical 75 mL dose of nadofaragene firadenovec (3 × 1011 viral particles per mL). Repeat dosing at months 3, 6, and 9 was done in the absence of high-grade recurrence. The primary endpoint was complete response at any time in patients with carcinoma in situ (with or without a high-grade Ta or T1 tumour). The null hypothesis specified a complete response rate of less than 27% in this cohort. Efficacy analyses were done on the per-protocol population, to include only patients strictly meeting the BCG-unresponsive definition. Safety analyses were done in all patients who received at least one dose of treatment. The study is ongoing, with a planned 4-year treatment and monitoring phase. This study is registered with ClinicalTrials.gov, NCT02773849. FINDINGS: Between Sept 19, 2016, and May 24, 2019, 198 patients were assessed for eligibility. 41 patients were excluded, and 157 were enrolled and received at least one dose of the study drug. Six patients did not meet the definition of BCG-unresponsive non-muscle-invasive bladder cancer and were therefore excluded from efficacy analyses; the remaining 151 patients were included in the per-protocol efficacy analyses. 55 (53·4%) of 103 patients with carcinoma in situ (with or without a high-grade Ta or T1 tumour) had a complete response within 3 months of the first dose and this response was maintained in 25 (45·5%) of 55 patients at 12 months. Micturition urgency was the most common grade 3-4 study drug-related adverse event (two [1%] of 157 patients, both grade 3), and there were no treatment-related deaths. INTERPRETATION: Intravesical nadofaragene firadenovec was efficacious, with a favourable benefit:risk ratio, in patients with BCG-unresponsive non-muscle-invasive bladder cancer. This represents a novel treatment option in a therapeutically challenging disease state. FUNDING: FKD Therapies Oy.

Cancer Risk Stratification of Anal Intraepithelial Neoplasia in Human Immunodeficiency Virus-Positive Men by Validated Methylation Markers Associated With Progression to Cancer.

BACKGROUND: High-grade anal intraepithelial neoplasia (HGAIN; AIN2-3) is highly prevalent in HIV+ men, but only a minority of these lesions progress towards cancer. Currently, cancer progression risk cannot be established; therefore, no consensus exists on whether HGAIN should be treated. This study aimed to validate previously identified host cell DNA methylation markers for detection and cancer risk stratification of HGAIN. METHODS: A large independent cross-sectional series of 345 anal cancer, AIN3, AIN2, AIN1, and normal control biopsies of HIV+ men was tested for DNA methylation of 6 genes using quantitative methylation-specific PCR. We determined accuracy for detection of AIN3 and cancer (AIN3+) by univariable and multivariable logistic regression analysis, followed by leave-one-out cross-validation. Methylation levels were assessed in a series of 10 anal cancer cases with preceding HGAIN at similar anatomic locations, and compared with the cross-sectional series. RESULTS: Methylation levels of all genes increased with increasing severity of disease (P < .05). HGAIN revealed a heterogeneous methylation pattern, with a subset resembling cancer. ZNF582 showed highest accuracy (AUC = 0.88) for AIN3+ detection, slightly improved by addition of ASCL1 and SST (AUC = 0.89), forming a marker panel. In the longitudinal series, HGAIN preceding cancer displayed high methylation levels similar to cancers. CONCLUSIONS: We validated the accuracy of 5 methylation markers for the detection of anal (pre-) cancer. High methylation levels in HGAIN were associated with progression to cancer. These markers provide a promising tool to identify HGAIN in need of treatment, preventing overtreatment of HGAIN with a low cancer progression risk.