Keratin 17 is a prognostic and predictive biomarker in pancreatic ductal adenocarcinoma.

OBJECTIVES: To determine the role of keratin 17 (K17) as a predictive biomarker for response to chemotherapy by defining thresholds of K17 expression based on immunohistochemical tests that could be used to optimize therapeutic intervention for patients with pancreatic ductal adenocarcinoma (PDAC). METHODS: We profiled K17 expression, a hallmark of the basal molecular subtype of PDAC, by immunohistochemistry in 2 cohorts of formalin-fixed, paraffin-embedded PDACs (n = 305). We determined a K17 threshold of expression to optimize prognostic stratification according to the lowest Akaike information criterion and explored the potential relationship between K17 and chemoresistance by multivariate predictive analyses. RESULTS: Patients with advanced-stage, low K17 PDACs treated using 5-fluorouracil (5-FU)-based chemotherapeutic regimens had 3-fold longer survival than corresponding cases treated with gemcitabine-based chemotherapy. By contrast, PDACs with high K17 did not respond to either regimen. The predictive value of K17 was independent of tumor mutation status and other clinicopathologic variables. CONCLUSIONS: The detection of K17 in 10% or greater of PDAC cells identified patients with shortest survival. Among patients with low K17 PDACs, 5-FU-based treatment was more likely than gemcitabine-based therapies to extend survival.

Dysregulated pulmonary inflammatory responses exacerbate the outcome of secondary aspergillosis following influenza.

IMPORTANCE: Severe influenza is a risk factor for fatal invasive pulmonary aspergillosis; however, the mechanistic basis for the lethality is unclear. Utilizing an influenza-associated pulmonary aspergillosis (IAPA) model, we found that mice infected with influenza A virus followed by Aspergillus fumigatus had 100% mortality when superinfected during the early stages of influenza but survived at later stages. While superinfected mice had dysregulated pulmonary inflammatory responses compared to controls, they had neither increased inflammation nor extensive fungal growth. Although influenza-infected mice had dampened neutrophil recruitment to the lungs following subsequent challenge with A. fumigatus, influenza did not affect the ability of neutrophils to clear the fungi. Our data suggest that the lethality seen in our model of IAPA is multifactorial with dysregulated inflammation being a greater contributor than uncontrollable microbial growth. If confirmed in humans, our findings provide a rationale for clinical studies of adjuvant anti-inflammatory agents in the treatment of IAPA.

Screening adequacy of unstained thyroid fine needle aspiration samples using a deep learning-based classifier.

Fine needle aspiration (FNA) biopsy of thyroid nodules is a safe, cost-effective, and accurate diagnostic method for detecting thyroid cancer. However, about 10% of initial FNA biopsy samples from patients are non-diagnostic and require repeated FNA, which delays the diagnosis and appropriate care. On-site evaluation of the FNA sample can be performed to filter out non-diagnostic FNA samples. Unfortunately, it involves a time-consuming staining process, and a cytopathologist has to be present at the time of FNA. To bypass the staining process and expert interpretation of FNA specimens at the clinics, we developed a deep learning-based ensemble model termed FNA-Net that allows in situ screening of adequacy of unstained thyroid FNA samples smeared on a glass slide which can decrease the non-diagnostic rate in thyroid FNA. FNA-Net combines two deep learning models, a patch-based whole slide image classifier and Faster R-CNN, to detect follicular clusters with high precision. Then, FNA-Net classifies sample slides to be non-diagnostic if the total number of detected follicular clusters is less than a predetermined threshold. With bootstrapped sampling, FNA-Net achieved a 0.81 F1 score and 0.84 AUC in the precision-recall curve for detecting the non-diagnostic slides whose follicular clusters are less than six. We expect that FNA-Net can dramatically reduce the diagnostic cost associated with FNA biopsy and improve the quality of patient care.

Dysregulated Pulmonary Inflammatory Responses Exacerbate the Outcome of Secondary Aspergillosis Following Influenza.

Inhalation of airborne conidia of the ubiquitous fungus Aspergillus fumigatus commonly occurs but invasive aspergillosis is rare except in profoundly immunocompromised persons. Severe influenza predisposes patients to invasive pulmonary aspergillosis by mechanisms that are poorly defined. Using a post-influenza aspergillosis model, we found that superinfected mice had 100% mortality when challenged with A. fumigatus conidia on days 2 and 5 (early stages) of influenza A virus infection but 100% survival when challenged on days 8 and 14 (late stages). Influenza-infected mice superinfected with A. fumigatus had increased levels of the pro-inflammatory cytokines and chemokines IL-6, TNFα, IFNß, IL-12p70, IL-1α, IL-1ß, CXCL1, G-CSF, MIP-1α, MIP-1ß, RANTES and MCP-1. Surprisingly, on histopathological analysis, superinfected mice did not have greater lung inflammation compared with mice infected with influenza alone. Mice infected with influenza had dampened neutrophil recruitment to the lungs following subsequent challenge with A. fumigatus , but only if the fungal challenge was executed during the early stages of influenza infection. However, influenza infection did not have a major effect on neutrophil phagocytosis and killing of A. fumigatus conidia. Moreover, minimal germination of conidia was seen on histopathology even in the superinfected mice. Taken together, our data suggest that the high mortality rate seen in mice during the early stages of influenza-associated pulmonary aspergillosis is multifactorial, with a greater contribution from dysregulated inflammation than microbial growth. Importance: Severe influenza is a risk factor for fatal invasive pulmonary aspergillosis; however, the mechanistic basis for the lethality is unclear. Utilizing an influenza-associated pulmonary aspergillosis (IAPA) model, we found that mice infected with influenza A virus followed by A. fumigatus had 100% mortality when superinfected during the early stages of influenza but survived at later stages. While superinfected mice had dysregulated pulmonary inflammatory responses compared to controls, they had neither increased inflammation nor extensive fungal growth. Although influenza-infected mice had dampened neutrophil recruitment to the lungs following subsequent challenge with A. fumigatus , influenza did not affect the ability of neutrophils to clear the fungi. Our data suggest that the lethality seen in our model IAPA is multifactorial with dysregulated inflammation being a greater contributor than uncontrollable microbial growth. If confirmed in humans, our findings provide a rationale for clinical studies of adjuvant anti-inflammatory agents in the treatment of IAPA.

COVID-19-induced Esophageal Necrosis Requiring Emergent Total Esophagectomy in a Vaccinated Patient.

Acute esophageal necrosis may be a potential complication of Coronavirus Disease 2019 (COVID-19). COVID-19 has been associated with a variety of sequelae, including acute respiratory distress syndrome, myocarditis, and thromboembolic events. Here, we present a case of a 43-year-old male who was admitted for acute necrotizing pancreatitis and found to have COVID-19 pneumonia. He subsequently developed acute esophageal necrosis requiring a total esophagectomy. Currently, there are at least five other reported cases of esophageal necrosis with concomitant COVID-19 infection. This case is the first requiring esophagectomy. Future studies may establish esophageal necrosis as a known complication of COVID-19.

Predictive Design and Analysis of Drug Transport by Multiscale Computational Models Under Uncertainty.

Computational modeling of drug delivery is becoming an indispensable tool for advancing drug development pipeline, particularly in nanomedicine where a rational design strategy is ultimately sought. While numerous in silico models have been developed that can accurately describe nanoparticle interactions with the bioenvironment within prescribed length and time scales, predictive design of these drug carriers, dosages and treatment schemes will require advanced models that can simulate transport processes across multiple length and time scales from genomic to population levels. In order to address this problem, multiscale modeling efforts that integrate existing discrete and continuum modeling strategies have recently emerged. These multiscale approaches provide a promising direction for bottom-up in silico pipelines of drug design for delivery. However, there are remaining challenges in terms of model parametrization and validation in the presence of variability, introduced by multiple levels of heterogeneities in disease state. Parametrization based on physiologically relevant in vitro data from microphysiological systems as well as widespread adoption of uncertainty quantification and sensitivity analysis will help address these challenges.

Primary Sarcoidosis of the Temporal Bone a Clinical Pathologic Correlation.

Objective: This report describes a case of sarcoidosis that presented as a lytic bone lesion in the squamous part of the temporal bone. Patients: A 64-year-old woman presented with right-sided aural fullness, pulsatile tinnitus, and intermittent otalgia. Interventions: CT and MRI were performed without contrast and suggested an osseodestructive, lytic bone lesion. An excisional biopsy was performed, showing granulomatous infiltration suggestive of osseous sarcoidosis. Main Outcome Measures: Removal of mass and resolution of symptoms. Results: Initial findings from patient imaging suggested a lytic bone lesion. An excisional biopsy was required for diagnosis and was performed with little patient morbidity. Biopsy findings showed granulomatous infiltration suggestive of osseous sarcoidosis. Osseous involvement of sarcoidosis is a rare manifestation and typically occurs secondary to other disease manifestations. After the removal of the mass and a short unrelated course of steroids, the patient's symptoms resolved. Conclusions: Sarcoidosis should be added to the differential diagnosis of lytic bone lesions in the temporal bone.

Deconvolution microscopy: A platform for rapid on-site evaluation of fine needle aspiration specimens that enables recovery of the sample.

CONTEXT: Rapid on-site evaluation (ROSE) optimises the performance of cytology, but requires skilled handling, and smearing can make the material unavailable for some ancillary tests. There is a need to facilitate ROSE without sacrificing part of the sample. OBJECTIVE: We evaluated the image quality of inexpensive deconvolution fluorescence microscopy for optically sectioning non-smeared fine needle aspiration (FNA) tissue fragments. DESIGN: A portion of residual material from 14 FNA samples was stained for 3 min in Hoechst 33342 and Sypro™ Red to label DNA and protein respectively, transferred to an imaging chamber, and imaged at 200× or 400× magnification at 1 micron intervals using a GE DeltaVision inverted fluorescence microscope. A deconvolution algorithm was applied to remove out-of-plane signal, and the resulting images were inverted and pseudocoloured to resemble H&E sections. Five cytopathologists blindly diagnosed 2 to 4 representative image stacks per case (total 70 evaluations), and later compared them to conventional epifluorescent images. RESULTS: Accurate definitive diagnoses were rendered in 45 (64%) of 70 total evaluations; equivocal diagnoses (atypical or suspicious) were made in 21 (30%) of the 70. There were two false positive and two false negative "definite" diagnoses in three cases (4/70; 6%). Cytopathologists preferred deconvolved images compared to raw images (P < 0.01). The imaged fragments were recovered and prepared into a ThinPrep or cell block without discernible alteration. CONCLUSIONS: Deconvolution improves image quality of FNA fragments compared to epifluorescence, often allowing definitive diagnosis while enabling the ROSE material to be subsequently triaged.

Keratin 17 testing in pancreatic cancer needle aspiration biopsies predicts survival.

BACKGROUND: Although pancreatic ductal adenocarcinoma (PDAC) has one of the lowest 5-year survival rates of all cancers, differences in survival exist between patients with clinically identical characteristics. The authors previously demonstrated that keratin 17 (K17) expression in PDAC, measured by RNA sequencing or immunohistochemistry (IHC), is an independent negative prognostic biomarker. Only 20% of cases are candidates for surgical resection, but most patients are diagnosed by needle aspiration biopsy (NAB). The aims of this study were to determine whether there was a correlation in K17 scores detected in matched NABs and surgical resection tissue sections and whether K17 IHC in NAB cell block specimens could be used as a negative prognostic biomarker in PDAC. METHODS: K17 IHC was performed for a cohort of 70 patients who had matched NAB cell block and surgical resection samples to analyze the correlation of K17 expression levels. K17 IHC was also performed in cell blocks from discovery and validation cohorts. Kaplan-Meier and Cox proportional hazards regression models were analyzed to determine survival differences in cases with different levels of K17 IHC expression. RESULTS: K17 IHC expression correlated in matched NABs and resection tissues. NAB samples were classified as high for K17 when ≥80% of tumor cells showed strong (2+) staining. High-K17 cases, including stage-matched cases, had shorter survival. CONCLUSIONS: K17 has been identified as a robust and independent prognostic biomarker that stratifies clinical outcomes for cases that are diagnosed by NAB. Testing for K17 also has the potential to inform clinical decisions for optimization of chemotherapeutic interventions.

Modulating Mechanical and Shape-Memory Properties while Mitigating Degradation-Induced Inflammation of Polylactides by Pendant Aspirin Incorporation.

Synergistically modulating mechanical properties and improving shape-memory performance while mitigating degradation-induced chronic inflammation of polylactide (PLA)-based implants for biomedical applications remain elusive. We test the hypothesis that copolymerizing aspirin-functionalized glycolide with d,l-lactide could enhance the thermal processing, toughness, and shape-memory efficiency of the copolymer while mitigating local inflammatory responses upon its degradation. The content of pendant aspirin was readily modulated by monomer feeds during ring-opening polymerization, and the copolymers with ∼10% or less aspirin pendants exhibited gigapascal-tensile moduli at body temperature and significantly improved fracture toughness and energy dissipation that positively correlated with the aspirin pendant content. The copolymers also exhibited excellent thermal-healing and shape-memory efficacy, achieving a >97% temporary shape fixing ratio at room temperature and facile shape recovery at 50-65 °C. These drastic improvements were attributed to the dynamic hydrophobic aggregations among aspirin pendants that strengthen glassy-state physical entanglement of PLA while readily dissociating under stress/thermal activation. When subcutaneously implanted, the copolymers mitigated degradation-induced inflammation due to concomitant hydrolytic release of aspirin without suppressing early acute inflammatory responses. The incorporation of aspirin pendants in PLA represents a straightforward and innovative strategy to enhance the toughness, shape-memory performance, and in vivo safety of this important class of thermoplastics for biomedical applications.

Keratin 17 Expression Predicts Poor Clinical Outcome in Patients With Advanced Esophageal Squamous Cell Carcinoma.

The major roles of keratin 17 (K17) as a prognostic biomarker have been highlighted in a range of human malignancies. However, its relevance to esophageal squamous cell carcinoma (ESCC) remains unexplored. In this study, the relationship between K17 expression and clinicopathologic parameters and survival were determined by RNA sequencing (RNA-Seq) in 90 ESCCs and by immunohistochemistry (IHC) in 68 ESCCs. K17 expression was significantly higher in ESCC than in paired normal tissues at both the messenger RNA and protein levels. K17 messenger RNA and staining by IHC were significantly correlated with aggressive characteristics, including advanced clinical stage, invasion depth, and lymph node metastases; and were predictive of poor prognosis in advanced disease patients. Furthermore, K17 expression was detected by IHC in high-grade premalignant lesions of the esophageal mucosa, suggesting that K17 could also be a biomarker of dysplasia of the esophageal mucosa. Overall, this study established that K17 is a negative prognostic biomarker for the most common subtype of esophageal cancer.

The Utility of MYB Immunohistochemistry (IHC) in Fine Needle Aspiration (FNA) Diagnosis of Adenoid Cystic Carcinoma (AdCC).

Differentiating adenoid cystic carcinoma (AdCC) from other basaloid neoplasm in a fine needle aspiration (FNA) sample can be challenging. Activation of MYB in AdCC by the fusion transcript MYB-NFIB has been recently demonstrated in salivary gland and other organs. The aim of this study is to evaluate the utility of MYB immunohistochemistry (IHC) in distinguishing AdCCs and other basaloid neoplasm in cytology specimens. Eighteen FNA cases, from salivary gland and other sites, and their subsequent surgical resection specimens were included in the study. Eight cases were confirmed AdCC on resection. MYB IHC was performed on slides made from cytology cell block and surgical resection paraffin blocks. Percentage and intensity of nuclear staining in tumor cells was scored as 0 to 3. The staining results were concordant between cytology specimens and their corresponding surgical resection tumors. Strong diffuse nuclear staining (score 3, N = 5) was exclusively observed in AdCC, both in cytology and surgical specimens. Only one pleomorphic adenoma and one poorly differentiated basaloid carcinoma were positive for MYB staining (score 1 to 2). Any degree of nuclear MYB labeling was seen in 100% AdCC cases (N = 8/8) compared with of 20% (N = 2/10) of all other non-AdCC cases (P = < 0.001). The sensitivity and specificity of any degree MYB positivity for AdCC in cytology specimen is 100% and 78%. The sensitivity and specificity of strong diffuse MYB labeling (score 2 to 3) for AdCC is 83% and 100% in cytology specimen. Strong diffuse nuclear staining of MYB is valuable in supporting a cytologic diagnosis of AdCC. However, weak and focal labeling of MYB should be interpreted with caution as it can be seen in benign and other malignant basaloid lesions.

Keratin 17 is a negative prognostic biomarker in high-grade endometrial carcinomas.

Keratin 17 (K17) has been established as a negative prognostic biomarker in cervical and ovarian cancers but has not previously been evaluated as a prognostic biomarker in endometrial adenocarcinoma. The association of K17 with decreased patient survival may be explained in part by the discovery that K17 drives tumor aggression by serving as a nuclear shuttle of p27, leading to cell cycle progression and tumor growth. The current study tests the hypothesis that K17 mRNA and protein levels correlate with decreased survival of patients with high-grade endometrial cancer. Gene expression data (mRNA) from The Cancer Genome Atlas were analyzed for 271 high-grade endometrial carcinomas and K17 immunohistochemistry (IHC) was performed on a separate cohort of 119 high-grade endometrial cancer cases from two academic medical centers. Survival analyses were determined by Cox proportional hazards regression. High K17 mRNA and IHC correlated with decreased overall survival (HR: 1.8, P = .0101, HR: 1.8, P = .0488, respectively). K17 was positive in malignant glandular cells of the endometrium but not in other tissues, including endometrial stroma, myometrium and uterine sarcoma. These results support the conclusion that K17 is a negative prognostic biomarker in high-grade endometrial carcinoma and that K17 IHC test results could be used to inform decisions related to therapeutic intervention.

Insulin Receptor Substrate-1 (IRS-1) and IRS-2 expression levels are associated with prognosis in non-small cell lung cancer (NSCLC).

The insulin-like growth factor-1 (IGF-1) signaling pathway has been implicated in non-small cell lung cancer (NSCLC) outcomes and resistance to targeted therapies. However, little is known regarding the molecular mechanisms by which this pathway contributes to the biology of NSCLC. The insulin receptor substrate (IRS) proteins are cytoplasmic adaptor proteins that signal downstream of the IGF-1R and determine the functional outcomes of this signaling pathway. In this study, we assessed the expression patterns of IRS-1 and IRS-2 in NSCLC to identify associations between IRS-1 and IRS-2 expression levels and survival outcomes in the two major histological subtypes of NSCLC, adenocarcinoma (ADC) and squamous cell carcinoma (SCC). High IRS-2 expression was significantly associated with decreased overall survival in adenocarcinoma (ADC) patients, whereas low IRS-1 cytoplasmic expression showed a trend toward association with decreased overall survival in squamous cell carcinoma (SCC) patients. Tumors with low IRS-1 and high IRS-2 expression were found to be associated with poor outcomes in ADC and SCC, indicating a potential role for IRS-2 in the aggressive behavior of NSCLC. Our results suggest distinct contributions of IRS-1 and IRS-2 to the biology of ADC and SCC that impact disease progression.

Keratin 17 identifies the most lethal molecular subtype of pancreatic cancer.

Although the overall five-year survival of patients with pancreatic ductal adenocarcinoma (PDAC) is dismal, there are survival differences between cases with clinically and pathologically indistinguishable characteristics, suggesting that there are uncharacterized properties that drive tumor progression. Recent mRNA sequencing studies reported gene-expression signatures that define PDAC molecular subtypes that correlate with differences in survival. We previously identified Keratin 17 (K17) as a negative prognostic biomarker in other cancer types. Here, we set out to determine if K17 is as accurate as molecular subtyping of PDAC to identify patients with the shortest survival. K17 mRNA was analyzed in two independent PDAC cohorts for discovery (n = 124) and validation (n = 145). Immunohistochemical localization and scoring of K17 immunohistochemistry (IHC) was performed in a third independent cohort (n = 74). Kaplan-Meier and Cox proportional-hazard regression models were analyzed to determine cancer specific survival differences in low vs. high mRNA K17 expressing cases. We established that K17 expression in PDACs defines the most aggressive form of the disease. By using Cox proportional hazard ratio, we found that increased expression of K17 at the IHC level is also associated with decreased survival of PDAC patients. Additionally, within PDACs of advanced stage and negative surgical margins, K17 at both mRNA and IHC level is sufficient to identify the subgroup with the shortest survival. These results identify K17 as a novel negative prognostic biomarker that could inform patient management decisions.

A 54-Year-Old Man Presenting With Progressive Dyspnea and Interstitial Lung Abnormalities.

CASE PRESENTATION: A 54-year-old man presented with a 6-month history of progressive dyspnea occurring at rest and with minimal exertion. His medical history was notable for hypertension, non-insulin-dependent diabetes, coronary artery disease, and factor V Leiden heterozygosity. His social history was notable for a 43-pack-year smoking history but was otherwise unremarkable.

B7-H4 is Inversely Correlated With T-Cell Infiltration in Clear Cell but Not Serous or Endometrioid Ovarian Cancer.

B7-H4, a tumor-associated cell surface protein, is expressed in endometrioid (EM), serous (SE), and clear cell (CC) ovarian carcinomas. Prior in vitro studies from other groups indicated that elevated B7-H4 expression by tumor cells blocks T-cell activation; therefore, it had been postulated to play a role in shielding cancer cells from immune surveillance and averting apoptotic programs. To test the validity of these hypotheses, the present study was designed to compare the immunohistochemical staining intensity of B7-H4 in tumor cells of ovarian cancers with the number of tumor-infiltrating T cells and macrophages and with the levels of caspase-3 staining in apoptotic debris. Serial tissue sections from EM, SE, and CC carcinomas were analyzed across representative cross-sections of tumor resection specimens, demonstrating different levels of B7-H4 expression, highest in CC cancers. B7-H4 staining in CC tissue sections was significantly correlated with the number of CD3, CD4, and CD8 tumor-infiltrating T cells and with the number of CD14 tumor-infiltrating macrophages, but was not significantly related to caspase-3 staining. These results support the concept that high levels of B7-H4 expression are inversely correlated with tumor T-cell infiltration and with CD14-labeled macrophages but not caspase-3 expression in CC carcinomas. We did not, however, find clear evidence of a relationship between the lower levels of B7-H4 seen in EM and SE carcinomas and T cell or macrophage infiltration. Thus, high levels of B7-H4, as seen in CC carcinomas, is associated with decreased tumor infiltration by T cells and macrophages but the lower levels of expression, as observed in EM and SE carcinomas, appear less likely to play an effective role in protection from immune surveillance. Furthermore, we found no evidence of a correlation between B7-H4 expression and apoptosis. These findings highlight the importance of further investigation of B7-H4 as an immunomodulatory protein, to support the development of novel therapeutic interventions for improved efficacy of treatments for CC carcinoma.

Resolving Interobserver Discrepancies in Lung Cancer Diagnoses by Spectral Histopathology.

This paper reports the results of a collaborative lung cancer study between City of Hope Cancer Center (Duarte, California) and CIRECA, LLC (Cambridge, Massachusetts), comprising 328 samples from 249 patients, that used an optical technique known as spectral histopathology (SHP) for tissue classification. Because SHP is based on a physical measurement, it renders diagnoses on a more objective and reproducible basis than methods based on assessing cell morphology and tissue architecture. This report demonstrates that SHP provides distinction of adenocarcinomas from squamous cell carcinomas of the lung with an accuracy comparable to that of immunohistochemistry and highly reliable classification of adenosquamous carcinoma. Furthermore, this report shows that SHP can be used to resolve interobserver differences in lung pathology. Spectral histopathology is based on the detection of changes in biochemical composition, rather than morphologic features, and is therefore more akin to methods such as matrix-assisted laser desorption ionization time-of-flight mass spectrometry imaging. Both matrix-assisted laser desorption ionization time-of-flight mass spectrometry and SHP imaging modalities demonstrate that changes in tissue morphologic features observed in classical pathology are accompanied by, and may be correlated to, changes in the biochemical composition at the cellular level. Thus, these imaging methods provide novel insight into biochemical changes due to disease.