Preinvasive Neoplasia Research Articles

Abstract B084: Comparative functional analysis of TP53 alleles in pancreatic ductal adenocarcinoma in vivo

Abstract TP53 is the second most mutated gene in pancreatic ductal adenocarcinoma (PDAC), bearing both truncal loss-of-function and missense point mutations. TP53 encodes a stress-induced transcription factor (p53) capable of activating tumor suppressive pathways mediating cell cycle arrest, apoptosis, senescence, and metabolic reprogramming. Despite high mutational frequency, how TP53 mutants function to suppress pancreatic tumorigenesis in vivo remains unclear. Previous studies indicate that the most frequent point mutations observed in human PDAC (TP53 R175H and R273H; mouse homologues Trp53 R172H and R270H) exhibit both dominant-negative (e.g., response to radiation in leukemia cells) and gain-of-function (e.g., increased liver metastasis in advanced PDAC models) roles compared to p53 null cells in various contexts. To rigorously explore the differential functions of p53 point and deletion mutants in PDAC progression, we performed an isogenic comparative analysis of Trp53 alleles in vivo using Mosaic Analysis with Double Markers (MADM) in mice. MADM uses stochastic mitotic recombination to induce two genotypically distinct daughter cells – created at 1-to-1 ratios – expressing different Trp53 variants dependent on the genotype of the mouse (+/+ vs. -/-, R172H/R172H vs. +/+, R172H/R172H vs. -/-, R270H/R270H vs. +/+, R270H/R270H vs. -/-) and simultaneously label them with unique genetically encoded fluorescent markers (TdTomato vs. GFP). We integrated MADM into a faithful Kras-driven model of pancreatic tumorigenesis (KC: Kras LSL-G12D/+ ; Pdx1-Cre) and used fluorescence microscopy to trace subclonal populations with various Trp53 alleles within the same mouse. By analyzing the ratio of TdTomato+ to GFP+ cells, we studied the functional differences between alleles in driving tumor initiation and expansion at various stages of progression. Strikingly, we found that Trp53 -/- cells exhibited greater expansion in preinvasive pancreatic intraepithelial neoplasias (PanINs) compared to not only Trp53 +/+ cells but also cells harboring either point mutation (Trp53 R172H/R172H or Trp53 R270H/R270H ). While Trp53 R270H/R270H cells predominated over Trp53 +/+ in PanINs, Trp53 R172H/R172H cells were surprisingly indistinguishable from Trp53 +/+ cells, suggesting that p53R172H retains tumor suppressive properties in early PDAC progression. Conversely, both Trp53 R172H/R172H and Trp53 R270H/R270H cells progressed to advanced PDAC with comparable PDAC incidence and survival as Trp53 -/- , arguing that Trp53 R172H is a separation-of-function mutant concordant with a distinct transcriptional profile relative to Trp53 -/- tumor cells. Together, these studies offer the strongest evidence to date for stage-specific functional differences between TP53 mutant variants in cancer progression in vivo and provide a faithful MADM-based PDAC model to define molecular mechanisms (through ongoing transcriptomic, epigenomic, and metabolomic studies) that govern functional differences amongst these diverse alleles. Citation Format: Sherry S. Agabiti, Andy Tang, Timothy Nottoli, Mandar D. Muzumdar. Comparative functional analysis of TP53 alleles in pancreatic ductal adenocarcinoma in vivo [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr B084.

Clinical significance of atypical glandular cells on cytology: 10 years' experience of a colposcopic referral center.

'Atypical glandular cells' (AGC) is an uncommon cytological result of cervical Pap smears which includes a wide of histopathological diagnoses, from benign to premalignant and malignant cervical disorders, endometrial cancer and, occasionally, other genital malignancies. This study aims to provide a comprehensive overview of AGC, assessing risk factors and clinical and histological features in affected patients. A retrospective analysis was conducted on a cohort of 239 women diagnosed with AGC between 2012 and 2022 at the 'Regional Referral Center for Prevention, Diagnosis and Treatment of HPV-related Genital Disorders', Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, Milan, Italy. Following AGC detection, patients underwent colposcopy with endocervical sampling and endometrial assessment via pelvic ultrasound. Selective cases also received endometrial biopsies. Among a total of 190 women who underwent both colposcopy and endometrial assessment, 116 (61%) had negative clinical and histopathological findings. The remainder displayed various abnormalities: 36 women (18.9%) were found to have endometrial or cervical polyps, 23 (12.1%) were diagnosed with preinvasive cervical neoplasia, and 21 (10.9%) with invasive cervical or endometrial disease. Menopause, multiparity, and older age were all significantly associated with endometrial cancer, but none of the abovementioned variables were significantly associated with cervical neoplasia. Our data confirm that AGC may reveal the presence of a wide range of histopathological conditions. Patients diagnosed with AGC should undergo a careful evaluation including both colposcopy with endocervical sampling and an endometrial assessment.

Metaproteomic analysis from cervical biopsies and cytologies identifies proteinaceous biomarkers representing both human and microbial species

The detection of HPV infection and microbial colonization in cervical lesions is currently done through PCR-based viral or bacterial DNA amplification. Our objective was to develop a methodology to expand the metaproteomic landscape of cervical disease and determine if protein biomarkers from both human and microbes could be detected in distinct cervical samples. This would lead to the development of multi-species proteomics, which includes protein-based lateral flow diagnostics that can define patterns of microbes and/or human proteins relevant to disease status. In this study, we collected both non-frozen tissue biopsy and exfoliative non-fixed cytology samples to assess the consistency of detecting human proteomic signatures between the cytology and biopsy samples. Our results show that proteomics using biopsies or cytologies can detect both human and microbial organisms. Across patients, Lumican and Galectin-1 were most highly expressed human proteins in the tissue biopsy, whilst IL-36 and IL-1RA were most highly expressed human proteins in the cytology. We also used mass spectrometry to assess microbial proteomes known to reside based on prior 16S rRNA gene signatures. Lactobacillus spp. was the most highly expressed proteome in patient samples and specific abundant Lactobacillus proteins were identified. These methodological approaches can be used in future metaproteomic clinical studies to interrogate the vaginal human and microbiome structure and metabolic diversity in cytologies or biopsies from the same patients who have pre-invasive cervical intraepithelial neoplasia, invasive cervical cancer, as well as in healthy controls to assess how human and pathogenic proteins may correlate with disease presence and severity.

T cells in testicular germ cell tumors: new evidence of fundamental contributions by rare subsets

BackgroundImmune cell infiltration is heterogeneous but common in testicular germ cell tumors (TGCT) and pre-invasive germ cell neoplasia in situ (GCNIS). Tumor-infiltrating T cells including regulatory T (Treg) and follicular helper T (Tfh) cells are found in other cancer entities, but their contributions to TGCT are unknown.MethodsHuman testis specimens from independent patient cohorts were analyzed using immunohistochemistry, flow cytometry and single-cell RNA sequencing (scRNA-seq) with special emphasis on delineating T cell subtypes.ResultsProfound changes in immune cell composition within TGCT, shifting from macrophages in normal testes to T cells plus B and dendritic cells in TGCT, were documented. In most samples (96%), the CD4+ T cell frequency exceeded that of CD8+ cells, with decreasing numbers from central to peripheral tumor areas, and to tumor-free, contralateral testes. T cells including Treg and Tfh were most abundant in seminoma compared to mixed tumors and embryonal carcinoma.ConclusionDespite considerable heterogeneity between patients, T cell subtypes form a key part of the TGCT microenvironment. The novel finding of rare Treg and Tfh cells in human testis suggests their involvement in TGCT pathobiology, with implications for understanding tumor progression, to assess patients’ prognosis, and as putative targets for personalized immunotherapy.

Abstract 1262: Comparative analysis of TP53 alleles in pancreatic ductal adenocarcinoma

Abstract The transcription factor p53 is a tumor suppressor capable of inducing cell cycle arrest, senescence, and apoptosis. Both truncating loss-of-function mutations and point mutations of the TP53 gene (mouse gene Trp53), which encodes for the p53 protein, are frequently observed across human cancers. While point mutations in p53 result in loss of canonical tumor suppressor function, recent studies suggest that these mutated proteins acquire pro-tumorigenic gain-of-function properties. For example, Trp53R172H mutations (the murine homolog of TP53R175H, the most common human TP53 variant) enhance metastatic spread relative to deletion mutations in mouse models of advanced pancreatic ductal adenocarcinoma (PDAC). To further explore the differential functions of p53 point and deletion mutants in cancer progression, we performed a rigorous comparative analysis of Trp53 alleles in vivo using Mosaic Analysis with Double Markers (MADM) in mice. MADM uses stochastic mitotic recombination to induce two genotypically distinct daughter cells - created at 1-to-1 ratios - expressing different Trp53 variants dependent on the genotype of the mouse (+/+ vs. -/-, R172H/R172H vs. +/+, R172H/R172H vs. -/-) and simultaneously label them with unique genetically encoded fluorescent markers (TdTomato vs. GFP). We integrated MADM into a faithful Kras-driven model of pancreatic tumorigenesis and used fluorescence microscopy to trace subclonal populations with various p53 alleles within the same mouse. By analyzing the ratio of TdTomato+ to GFP+ cells, we studied the functional differences between alleles in driving pancreatic cancer at various stages of progression. We found that Trp53−/− cells exhibited greater tumor initiation of preinvasive pancreatic intraepithelial neoplasia (PanINs) and early PanIN cell expansion compared to Trp53+/+ cells. Surprisingly, Trp53R172H/R172H cells were comparable in number to Trp53+/+ cells in early PanINs but were outcompeted by Trp53−/− cells.In contrast, both Trp53R172H/R172H and Trp53−/− cells were capable of facilitating the transition to advanced PDAC with comparable survival times. These phenotypic differences imply that p53R172H retains tumor suppressive properties in early pancreatic tumorigenesis, but similar to the loss of p53, it is incapable of constraining progression to advanced disease, arguing that p53R172H is a separation of function mutation in vivo. Ongoing studies examining the molecular differences between Trp53 alleles by fluorescence-activated cell sorting and RNA-sequencing will shed further light onto the underlying mechanisms that explain these phenotypic differences. Citation Format: Andy Tang, Sherry Agabiti, Hannah Chung, Mandar Deepak Muzumdar. Comparative analysis of TP53 alleles in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1262.

Abstract 6165: Early detection of pancreatic cancer by hyperpolarized magnetic resonance

Abstract Introduction: Pancreatic cancer is difficult to diagnose due to its asymptomatic presentation at early stages. Therefore, there is an unmet need for non-invasive imaging markers that can help identify the aggressive pancreatic lesions at an early time point. One of the most promising imaging biomarkers is the conversion of hyperpolarized (HP) pyruvate to lactate1-3. With HP metabolic imaging, the conversion will be monitored among different premalignant models at different timepoints as well as tested against pancreatitis, a known confounder of pancreatic cancer in the clinic. Methods: HP [1-13C]pyruvate Magnetic Resonance Spectroscopy (MRS) was employed to study the metabolic processes in tamoxifen inducible genetically engineered mouse models (P48CreERT2;LSL-KrasG12D(iKC)) with pre-invasive pancreatic intraepithelial neoplasia (PanIN) precursor lesions, invasive pancreatic cancer model (P48CreERT2;LSL-KrasG12D; LSL-p53R172H (iKPC)) and control animals (P48CreERT2 (iC)) at 7T Bruker MRI scanner. These mice were imaged before tamoxifen induction at 10-, 20-, and 30-weeks post induction. For the pancreatitis model, iC and iKC mice they were treated with caerulein for three weeks, 7 weeks after tamoxifen induction. The pancreatitis mice were imaged 24 hours after the last caerulein injection. Results/Discussion: HP lactate-to-pyruvate ratio is increased in the cancer models compared to the control model. In the iKPC mouse model, at the 20-week post induction imaging there was a significant increase of the lactate-to-pyruvate ratio compared to the 10-weektime point after induction. The 20-week iKPC time point ratio compared to the iKC and control mouse models was significantly higher, indicating the aggressive nature of the disease. Even in the iKC model there is a slight increase of the lactate-to-pyruvate ratio at 20-weeks post induction compared to both previous time points, pre-induction and 10-week. No change in the lactate-to-pyruvate ratio was not observed with the pancreatitis phenotype. In the future, we plan on imaging at earlier timepoints or at smaller intervals in the iKPC model, to observe the exact moment the metabolism switches to lactate. Conclusion: Our data demonstrates that the metabolic flux to lactate is only increasing in the premalignant models of pancreatic cancer but not in the pancreatitis model, suggesting that the differentiation between early pancreatic cancer and pancreatitis can be achieved by hyperpolarized metabolic imaging. We are currently in the process of translating this in a clinical trial at MD Anderson Cancer Center. Citation Format: Paytience L. Smith, José S. Enriquez, Rian Howell, Olivereen Le Roux, Shivanand Pudakalakatti, Prasanta Dutta, Florencia Mcallister, Pratip Bhattacharya. Early detection of pancreatic cancer by hyperpolarized magnetic resonance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6165.

Abstract 2873: Evolution of immune cell composition and functionality as pancreatic intraepithelial neoplasia progresses to pancreatic ductal adenocarcinoma

Abstract Pancreatic ductal adenocarcinoma (PDAC) is most often diagnosed at an advanced stage. Newly diagnosed patients therefore have a dismal five-year survival rate of 11%. However, PDAC progresses from pre-invasive pancreatic intraepithelial neoplasia (PanIN) over at least a decade. Throughout this transition, the tissue microenvironment becomes increasingly immunosuppressive. Early PanINs may therefore be more amenable to immune-based interception strategies; however, little is known about the pre-malignant lesion immune microenvironment in PDAC. We hypothesized that the identification of the immune landscape of PanINs will elucidate the immuno-dynamic changes that occur during PanIN-to-PDAC progression and identify novel strategies for intercepting PDAC. Here, we use an inducible mouse model to study the evolution of immunosuppression from PanINs to PDAC. We pair spatial molecular profiling of our mouse model with profiling of human tissue in a cohort of patients with normal tissue, chronic pancreatitis, PanIN, and PDAC. To examine the evolution of the immune microenvironment throughout PanIN-to-PDAC progression, we first optimized a tamoxifen-inducible Pdx1-CreERT2 mouse that controls KRASG12D expression and knocks out p53. The impact of KRASG12D expression on the immune cell landscape in PanIN and PDAC lesions was examined by immunohistochemistry (IHC) and RNA in situ hybridization on mouse pancreas. We used imaging mass cytometry (IMC) of 35 immune markers to better classify and quantify the immune cell subtypes. Our analyses thus far reveal increased Tregs as PanINs progress to PDAC. Furthermore, although CD3+ T cells are recruited to tumors, these immune cells are strictly restricted to the immediate edge of the tumor and predominantly consist of Tregs. For our human analyses, FFPE pancreas sections from treatment naïve patients who had undergone surgical resection without neoadjuvant chemotherapy were evaluated. Each section contained regions of normal tissue, chronic pancreatitis, PanIN and PDAC. These were evaluated using IHC, IMC, and spatial transcriptomics to allow us to spatially evaluate the immune populations associated with lesions and PDAC. The relative density and localization of myeloid and lymphoid cell types in both PanIN and PDAC regions revealed an initial influx of CD8+ and CD4+ T cells to PanINs and a progressively immunosuppressive microenvironment in subsequent stages. While PanINs and PDAC both recruited immune cells, the phenotypes of the immune infiltrates were distinct and revealed unique immune pathways that could contribute to immunosuppression as PanINs develop into PDACs. Our proteomic and transcriptomic data from mouse and human pancreas show that mutant KRAS driven premalignant lesions recruit an evolving immune response that readily becomes immunosuppressive as progression to PDAC occurs. Citation Format: Melissa R. Lyman, Jacob T. Mitchell, Luciane T. Kagohara, Benjamin Barrett, Amanda Huff, Sarah Shin, Gabriella Longway, Anuj Gupta, Lalitya Andaloori, Todd D. Armstrong, Daniel Haldar, Robert Anders, Elizabeth Thompson, Nilo Azad, Won Jin Ho, Elizabeth Jaffee, Elana J. Fertig, Neeha Zaidi. Evolution of immune cell composition and functionality as pancreatic intraepithelial neoplasia progresses to pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2873.

Pancreatic Cancer: Changing Epidemiology and New Approaches to Risk Assessment, Early Detection, and Prevention

Pancreatic cancer usually results in poor survival with limited options for treatment, as most affected individuals present with advanced disease. Early detection of preinvasive pancreatic neoplasia and identifying molecular therapeutic targets provide opportunities for extending survival. Although screening for pancreatic cancer is currently not recommended for the general population, emerging evidence indicates that pancreatic surveillance can improve outcomes for individuals in certain high-risk groups. Changes in the epidemiology of pancreatic cancer, experience from pancreatic surveillance, and discovery of novel biomarkers provide a roadmap for new strategies for pancreatic cancer risk assessment, early detection, and prevention.

Neuropeptide Y and its receptors in prostate cancer: associations with cancer invasiveness and perineural spread

PurposeNeuropeptide Y (NPY) is a pleiotropic peptide, which is involved in many biological mechanisms important in regulation of cell growth and survival. The aim of this study was a comprehensive analysis of the NPY system in prostate pathology.MethodsThe study was based on immunohistochemical analysis of NPY and its receptors, Y1R, Y2R and Y5R, in tissue samples from benign prostate (BP), primary prostate cancer (PCa) and PCa bone metastases. Tissue microarray (TMA) technique was employed, with analysis of multiple cores from each specimen. Intensity of the immunoreactivity and expression index (EI), as well as distribution of the immunostaining in neoplastic cells and stromal elements were evaluated. Perineural invasion (PNI) and extraprostatic extension (EPE) were areas of special interests. Moreover, a transwell migration assay on the LNCaP PCa cell line was used to assess the chemotactic properties of NPY.ResultsMorphological analysis revealed homogeneous membrane and cytoplasmic pattern of NPY staining in cancer cells and its membrane localization with apical accentuation in BP glands. All elements of the NPY system were upregulated in pre-invasive prostate intraepithelial neoplasia, PCa and metastases. EI and staining intensity of NPY receptors were significantly higher in PCa then in BP with correlation between Y2R and Y5R. The strength of expression of the NPY system was further increased in the PNI and EPE areas. In bone metastases, Y1R and Y5R presented high expression scores.ConclusionThe results of our study suggest that the NPY system is involved in PCa, starting from early stages of its development to disseminated states of the disease, and participates in the invasion of PCa into the auto and paracrine matter.

Expression of Intermediate Filaments in the Developing Testis and Testicular Germ Cell Cancer.

Cytokeratin and desmin expression have been associated with Sertoli cell maturity and the development of testicular germ cell cancer (TGCC). Thus, the present study aimed to characterize the expression of these intermediate filaments in normal testis development and TGCC. Cytokeratin and desmin were determined by immunohistochemistry and immunofluorescence in human fetal, and adult testis and tissue from patients with pre-invasive germ cell neoplasia in-situ (GCNIS) or invasive TGCC. Desmin was expressed in Sertoli cells of the human fetal testis, and the proportion of desmin expressing Sertoli cells was significantly reduced in the second trimester, compared with the first trimester (31.14% vs. 6.74%, p = 0.0016). Additionally, Desmin was expressed in the majority of Sertoli cells in the adult testis and TGCC samples. Cytokeratin was detected in Sertoli cells of human fetal testis but was not expressed in Sertoli cells of human adult testis. In patients with TGCC, cytokeratin was not expressed in Sertoli cells in tubules with active spermatogenesis but was detected in Sertoli cells in tubules containing GCNIS cells in patients with both pre-invasive and invasive TGCC. In conclusion, desmin was not associated with Sertoli cell maturation or progression to TGCC. However, cytokeratin appeared to be an indicator of impaired Sertoli cell maturation.

Tumor-associated tissue eosinophilia in preinvasive and invasive squamous cell carcinoma of cervix

Background: Differentiating between preinvasive and invasive squamous neoplasms of the cervix can be a challenge in small, superficial fragments of cervical biopsy, particularly when inflammatory infiltrates obscure the epithelial stroma interface and due to tangential sectioning of the acanthotic squamous epithelium. This study evaluates whether thresholds of eosinophils in stroma serve as a morphological marker for the presence of tumor invasion in squamous neoplasms of the cervix. Material and methods: Sixty-five cases of invasive squamous cell carcinoma and preinvasive squamous neoplasia were evaluated. In each case, the number of eosinophils per HPF and per 10 HPF in the tissue adjacent to the neoplastic epithelium was counted. For statistical purposes, the elevated eosinophils were categorized as > 3 eosinophils/HPF, > 5 eosinophils/HPF, and >10/10 HPF. Results: Stromal eosinophils were present in 1/5 cases of LSIL, 3/8 HSIL cases, 15/16 cases of microinvasive squamous cell carcinoma, and 34/36 cases of invasive squamous cell carcinoma, signifying an increase in eosinophil count from a spectrum of preinvasive to invasive carcinoma. Greater than 5 eosinophils/HPF and >10 eosinophils/10 HPF had sensitivity, specificity, and positive predictive values of 55%, 100%, 100%, and 75%, 100%and 100%, respectively. The difference of >5eosinophils/HPF and >10eosinophils/10 HPF between SIL and SCC was statistically significant. Conclusions: Our study suggests that the presence of increased eosinophils in squamous neoplasia of the cervix can be associated with tumor invasion and should prompt a thorough evaluation when evidence of invasion is absent.

Peak density of immature nerve cells occurs with high-grade dysplasia in intraductal papillary mucinous neoplasms of the pancreas.

The development of neural structures within tumors is now considered vital for carcinogenesis. However, the time course of this development in human pre-invasive neoplasia has been incompletely described. Therefore, we performed a detailed analysis of nerves across the neoplastic spectrum in resected intraductal papillary mucinous neoplasms (IPMNs) of the pancreas. Histology and multiplexed immunochemistry demonstrated that nerve density increased from low-grade (LG) to high-grade dysplasia (HG) but did not further increase once invasive IPMN (INV IPMN) was present. Higher nerve density correlated with increasing expression of nerve growth factor (NGF) by the tumor cells. Intra-tumoral nerves were immature and lacked markers of sympathetic, parasympathetic, and sensory lineages. Here, we show for the first time the presence of neural precursor cells (NPCs) within the stroma of pancreatic tumors. The density of these doublecortin (DCX)-positive NPCs increased from LG to HG, but not from HG to INV IPMN. We conclude that peak neural density of tumors is reached in high-grade dysplasia (often termed carcinoma in situ) rather than after invasion. These findings suggest that nerve-tumor interactions are important in IPMN progression and may serve as the basis for future mechanistic studies and novel therapeutic modalities. © 2022 The Pathological Society of Great Britain and Ireland.

Comparison of simulated outcomes between stool- and blood-based colorectal cancer screening tests.

10529 Background: The Centers for Medicare & Medicaid Services (CMS) recommends covering blood-based biomarker tests with proposed minimum performance thresholds for colorectal cancer (CRC) screening test every 3 years for average-risk, asymptomatic Medicare beneficiaries ages 50–85 years. A blood-based test is a non-invasive screening method to detect CRC but is limited by low sensitivity to detect adenomas. Using the CRC-AIM microsimulation model, predicted life-years gained (LYG) and CRC incidence and mortality reduction were compared between a blood-based test meeting the CMS minimum thresholds and stool-based screening tests (fecal immunochemical test [FIT], fecal occult blood test [FOBT], and multitarget stool DNA [mt-sDNA]). Methods: Outcomes of blood- and stool-based tests were simulated for average-risk individuals free of diagnosed CRC at age 40 and screened between ages 45–75 years per USPSTF recommendations. Per CMS proposed criteria, CRC sensitivity and specificity for a blood-based test were set at 74% and 90%, respectively, and adenoma sensitivity was set at 10%. Published adenoma and CRC sensitivity and specificity were used for each stool test. For the primary analysis, adherence was assumed to be 100%. For secondary analysis, adherence was set at 30–70%, in 10% increments. Outcomes were per 1000 individuals. Results: At 100% adherence, LYG was 229 for a blood test and ≥305 for stool tests, corresponding to at least 25% lower LYG for a blood test relative to all stool tests (Table). Absolute CRC incidence and mortality reductions were at least 19% and 18% lower, respectively, for a blood test vs all stool tests. Secondary analysis indicates that at identical adherence rates that are less than 100%, a blood test had lower LYG vs all stool tests (Table). When the adherence of any test was 50%–70%, a blood test resulted in at least 20% lower LYG vs any stool test, and absolute reductions in CRC incidence and mortality were at least 9% and 10% lower, respectively, vs any stool test. Conclusions: This model suggests that if blood-based CRC screening tests do not sufficiently detect advanced adenomas, clinical outcomes will be inferior to stool-based testing due to lack of cancer prevention. Further discovery efforts to identify blood-based markers associated with both invasive and preinvasive neoplasia are needed to address this deficiency. [Table: see text]

Epstein-Barr Virus, But Not Human Papillomavirus, Is Associated With Preinvasive and Invasive Ocular Surface Squamous Neoplasias in Zambian Patients.

BackgroundThe etiopathogenesis of ocular surface squamous neoplasia (OSSN) is not fully understood. We assessed the frequency of oncogenic viruses in OSSN by immunohistochemistry (IHC) and polymerase chain reaction (PCR) for human papillomavirus (HPV), Epstein–Barr virus (EBV), Merkel cell polyomavirus (MCPyV), Kaposi sarcoma virus, and adenovirus. Cases from Zambia were prospectively enrolled using a cross-sectional study design between November 2017 and March 2020.MethodsDemographic and clinical data [age, sex, HIV status, antiretroviral therapy (ART) history, CD4 count, plasma viral load] and tumor biopsies were collected from 243 consenting patients. Tumor samples were bisected, and half was used for DNA isolation, while the other half was formalin fixed and paraffin embedded (FFPE) for histopathology analysis. The expressions of latent EBV nuclear antigen 1 (EBNA1), CDKN2A/p16INK4A (p16), and MCPyV large T-antigen (LT) were tested by IHC. Multiplex PCR was used to detect 16 HPV genotypes and four other DNA tumor viruses [Kaposi’s sarcoma-associated herpesvirus (KSHV), EBV, MCPyV, and adenovirus]. Relationships between HIV status, viral DNA and protein expression, and tumor grades were determined by statistical analysis.ResultsOSSN tumors from patients were 29.6% preinvasive and 70.4% invasive. Patients presented with unilateral tumors that were 70.4% late stage (T3/T4). OSSN patients were HIV positive (72.8%). IHC on 243 FFPE biopsies resulted in the detection of EBNA1 (EBV), p16 high-risk HPV (HR-HPV), and MCPyV LT expression in 89.0%, 4.9%, and 0.0%, respectively. EBNA1 was expressed in all grades of preinvasive [cornea–conjunctiva intraepithelial neoplasia (CIN)1, 100%; CIN2, 85.7%; CIN3, 95.8%; and carcinoma in situ (CIS), 83.8%] and in invasive (89.2%) OSSN. PCR on 178 samples detected EBV, HR-HPV, and MCPyV in 80.3%, 9.0%, and 13.5% of tumors, respectively. EBV was detected in all grades of preinvasive and invasive OSSN. EBV detection was associated with high HIV viral loads (p = 0.022). HR-HPV was detected in 0.0% CIN1, 0.0% CIN2, 5.6% CIN3, 13.0% CIS, and 7.0% invasive OSSN.ConclusionsOur findings of EBV DNA and EBNA1 protein in all the grades of preinvasive and especially invasive OSSN are consistent with a potential causal role for EBV in OSSN. A role of HPV in OSSN was not clearly established in this study.

A COMPARATIVE EVALUATION BETWEEN CERVICAL CYTOLOGY AND VIRAL BIOMARKERS IN EARLY DETECTION OF CERVICAL INTRAEPITHELIAL LESIONS.

Cervical cancer is a public health problem in developing countries like India, so much so that India alone accounts for one-quarter of the worldwide burden of cervical cancers.[1] It is the one of the leading causes of cancer mortality, accounting for 17% of all cancer deaths among women aged between 30 and 69 years. Every year in India, 122,844 women are diagnosed with cervical cancer and 67,477 die from the disease.[2] Cervical cancer is a leading form of cancer among women living in low resource regions of the world and often kills women at young age when they are still raising families. Cervical cancer due to its slow progression from precancerous lesions to frank malignancy and easy accessibility to examination, is highly amenable to screening. The three screening modalities are Cervical Cytology Screening (Pap Smear), Visual Inspection, and HPV DNA test. Squamous intraepithelial lesions are viewed as precancerous lesions exhibiting many of the morphological characteristics of invasive carcinomas, identication of these entities is the focus of cervical screening. Persistence of high-risk HPV infection after initial infection is highly predictive of a lifetime risk of pre-invasive and invasive cervical neoplasia. The HPV genotype seems to be the most important factor in persistence, with HPV-16 and HPV-18 being the most likely to persist. The aim of this study was to evaluate concurrently the efcacy of Cervical cytology, HPV PCR methods for risk assessment in an attempt to dene the most efcient combination, so as to know which one is more applicable and accurate in cervical cancer screening strategies. The present prospective observational study was conducted in Department of Pathology, NSCB medical college and hospital, Jabalpur, M.P. from 1st February 2019 to 31st July 2020, with the primary aim of to assess cytomorphological incidence of various lesions of uterine cervix by Bethesda system and to study the biomarker HPV-DNA testing by PCR in case of abnormal cervical ndings and to correlate the ndings and results of cytology with the histopathology reports. This study had a total population of 140 women who attended the gynaecology OPD and fullled the inclusion criteria were enrolled in study. Per speculum examination was done and Pap smear examination was taken followed by colposcopy guided cervical biopsy in the symptomatic females. Out of 140 ,60 samples were tested for high HPV 16 and 18 using RTPCR. The maximum numbers of the patients were in third decade of life, followed by fourth decade. Maximum numbers of cases were reported as inammatory smear. ASCUS and HSIL was seen commonly in the age group of 31 to 40 years. Most common presenting complaint was white discharge 55.71 % .Erosive cervicitis (41.4%) was the most common histopathological nding. HPV DNAtesting showed only 6.7 % positivity. 16.67 % Of squamous intraepithelial lesions were positive for HPV and the most prevalent HPV genotype was found to be HPVtype 16. Conclusion: Pap smear evaluation and HPVtyping are most efcient combination for mass cervical screening programmes

Molecular Alterations in Gastric Preneoplastic Lesions and Early Gastric Cancer.

Prognosis of gastric cancer is dramatically improved by early diagnosis. Correa’s cascade correlates the expression of some molecular markers with the progression of preneoplastic lesions toward carcinoma. This article reviews the diagnostic and prognostic values of molecular markers in complete (MUC2) and incomplete (MUC2, MUC5AC, and MUC6) intestinal metaplasia, gastric dysplasia/intra-epithelial neoplasia, and early gastric cancer. In particular, considering preinvasive neoplasia and early gastric cancer, some studies have demonstrated a correlation between molecular alterations and prognosis, for example, mucins phenotype in gastric dysplasia, and GATA6, TP53 mutation/LOH and MUC6 in early gastric cancer. Moreover, this review considers novelties from the literature regarding the (immuno)histochemical characterization of diffuse-type/signet ring cell gastric cancer, with particular attention to clinical outcomes of patients. The aim of this review is the evaluation of the state of the art regarding suitable biomarkers used in the pre-surgical phase, which can distinguish patients with different prognoses and help decide the best therapeutic strategy.

Vulvar Intraepithelial Neoplasia: A Review of the Disease and Current Management.

Vulvar intraepithelial neoplasia (VIN) represents an increasingly common, yet challenging diagnosis that shares many common risk factors with cervical intraepithelial neoplasia. However, unlike cervical intraepithelial neoplasia, effective screening and treatment strategies are much less defined for patients with VIN. The objective of this article is to review the underlying risk factors leading to the development of VIN, identify special populations at risk for VIN, and outline acceptable treatment strategies. This literature review was performed primarily using PubMed. Vulvar intraepithelial neoplasia can be separated into usual VIN (uVIN) and differentiated VIN (dVIN). The more common uVIN is related to underlying human papillomavirus infection, whereas dVIN occurs in the setting of other vulvar inflammatory conditions such as lichen sclerosis. Differentiated VIN carries a higher risk of progression to invasive malignancy. Extramammary Paget disease is a rare intraepithelial adenocarcinoma unrelated to uVIN and dVIN, although management is similar. Vulvar intraepithelial neoplasia is a preinvasive neoplasia of the vulva with few robust strategies for surveillance or management. Careful examination with targeted biopsy is warranted for suspicious lesions, and a combination of surgical and medical management can be tailored for individual patient needs.

Cervical cancer - statistical analysis data for 2016 and 2017 in Republic of North Macedonia

The exceptional accessibility of the cervix (cervix uteri) and direct physical examination have allowed intensive research into the nature of the malignant lesions of the cervix. Studies have shown that many of these tumors have no explosive but gradual onset. Their pre-invasive antecedents may persist for two-way surfaces or in situ disease for several years, although this may change, at least in some patients. There is convincing evidence that the cytological screening programs are effective in reducing mortality from cervical cancer. Screening has not only reduced the incidence and mortality rate but has also found many women with pre-invasive neoplasia (which is the role of screening rather than diagnosing cancer). From the results of the Institute of Public Health of the Republic of Northern Macedonia for two years 2016 and 2017, it is concluded that the cervical cancer mortality rate in 2016 is significantly higher than in 2017, where the screening program is more developed and higher and there’s more coverage of at-risk patient group (50+ years). In 2016, 87.4% had normal results, while 12.6% had cellular abnormalities, compared to 2017 - 76.2% had normal results, while 23.8% had cellular abnormalities. The mortality rate in 2017 from cervical cancer is 30 which is lower than in 2016. This situation points to the necessity of increasing the coverage of women in the Program for screening and promoting the implementation of the Program by the Ministry of Health as soon as possible.

Abstract LB-077: Dendritic cell dysfunction precedes tumor formation during pancreatic oncogenesis

Abstract Dysfunction of type 1 conventional dendritic cells (cDC1s) - the dendritic cell subtype crucial for spontaneous anti-tumor T cell immunity - has long been hypothesized to develop secondarily to invasive cancer. Here, we reveal that cDC1 dysfunction instead develops in the earliest stages of cancer development upon formation of preinvasive pancreatic intraepithelial neoplasias (PanINs) in the KrasG12D Trp53R172H Pdx1-Cre-driven (KPC) mouse model of pancreatic ductal adenocarcinoma. This results in systemically increased cDC1 apoptosis and insufficient upregulation of T cell-polarizing cytokines during cDC1 maturation. CD8+ T cell activation following vaccination is subsequently impaired in PanIN- and tumor-bearing mice. Yet, combination therapy with CD40 agonist and Flt3 ligand in tumor-bearing mice returns cDC1 abundance to normal levels and induces an IFNg response signature, repairing cDC1 maturation and restoring full CD8+ T cell activation. This study therefore reveals the unexpectedly early and systemic onset of cDC1 dysregulation while elaborating therapeutically tractable strategies towards cDC1 repair. Citation Format: Jeffrey H. Lin, Robert H. Vonderheide. Dendritic cell dysfunction precedes tumor formation during pancreatic oncogenesis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr LB-077.

Type 1 conventional dendritic cells are systemically dysregulated early in pancreatic carcinogenesis.

Type 1 conventional dendritic cells (cDC1s) are typically thought to be dysregulated secondarily to invasive cancer. Here, we report that cDC1 dysfunction instead develops in the earliest stages of preinvasive pancreatic intraepithelial neoplasia (PanIN) in the KrasLSL-G12D/+ Trp53LSL-R172H/+ Pdx1-Cre-driven (KPC) mouse model of pancreatic cancer. cDC1 dysfunction is systemic and progressive, driven by increased apoptosis, and results in suboptimal up-regulation of T cell-polarizing cytokines during cDC1 maturation. The underlying mechanism is linked to elevated IL-6 concomitant with neoplasia. Neutralization of IL-6 in vivo ameliorates cDC1 apoptosis, rescuing cDC1 abundance in tumor-bearing mice. CD8+ T cell response to vaccination is impaired as a result of cDC1 dysregulation. Yet, combination therapy with CD40 agonist and Flt3 ligand restores cDC1 abundance to normal levels, decreases cDC1 apoptosis, and repairs cDC1 maturation to drive superior control of tumor outgrowth. Our study therefore reveals the unexpectedly early and systemic onset of cDC1 dysregulation during pancreatic carcinogenesis and suggests therapeutically tractable strategies toward cDC1 repair.