Pulmonary Adenocarcinoma In Situ and Minimally Invasive Adenocarcinomas in European Patients Have Less KRAS and More EGFR Mutations Compared to Advanced Adenocarcinomas.

Pulmonary adenocarcinoma (ADC) is a very diverse disease, both genetically and histologically, which displays extensive intratumor heterogeneity with numerous acquired mutations. ADC is the most common type of lung cancer and is believed to arise from adenocarcinoma in situ (AIS) which then progresses to minimally invasive adenocarcinoma (MIA). In patients of European ethnicity, we analyzed genetic mutations in AIS (n = 10) and MIA (n = 18) and compared the number of genetic mutations with advanced ADC (n = 2419). Using next-generation sequencing, the number of different mutations detected in both AIS (87.5%) and MIA (94.5%) were higher (p < 0.001) than in advanced ADC (53.7%). In contrast to the high number of mutations in Kirsten rat sarcoma virus gene (KRAS) in advanced ADC (34.6%), there was only one case of AIS with KRAS G12C mutation (3.5%; p < 0.001) and no cases of MIA with KRAS mutation (p < 0.001). In contrast to the modest prevalence of epidermal growth factor receptor (EGFR) mutations in advanced ADC (15.0%), the fraction of EGFR mutant cases was higher in both in AIS (22.2%) and MIA (59.5%; p < 0.001). The EGFR exon 19 deletion mutation was more common in both MIA (50%; n = 6/12) and ADC (41%; n = 149/363), whereas p.L858R was more prevalent in AIS (75%; n = 3/4). In contrast to pulmonary advanced ADC, KRAS driver mutations are less common, whereas mutations in EGFR are more common, in detectable AIS and MIA.

Clinical Outcomes of Resected Pure Ground-Glass, Heterogeneous Ground-Glass, and Part-Solid Pulmonary Nodules.

BACKGROUND. Increased (but not definitively solid) attenuation within pure ground-glass nodules (pGGNs) may indicate invasive adenocarcinoma and the need for resection rather than surveillance. OBJECTIVE. The purpose of this study was to compare the clinical outcomes among resected pGGNs, heterogeneous ground-glass nodules (GGNs), and part-solid nodules (PSNs). METHODS. This retrospective study included 469 patients (335 female patients and 134 male patients; median age, 68 years [IQR, 62.5-73.5 years]) who, between January 2012 and December 2020, underwent resection of lung adenocarcinoma that appeared as a subsolid nodule on CT. Two radiologists, using lung windows, independently classified each nodule as a pGGN, a heterogeneous GGN, or a PSN, resolving discrepancies through discussion. A heterogeneous GGN was defined as a GGN with internal increased attenuation not quite as dense as that of pulmonary vessels, and a PSN was defined as having an internal solid component with the same attenuation as that of the pulmonary vessels. Outcomes included pathologic diagnosis of invasive adenocarcinoma, 5-year recurrence rates (locoregional or distant), and recurrence-free survival (RFS) and overall survival (OS) over 7 years, as analyzed by Kaplan-Meier and Cox proportional hazards regression analyses, with censoring of patients with incomplete follow-up. RESULTS. Interobserver agreement for nodule type, expressed as a kappa coefficient, was 0.69. Using consensus assessments, 59 nodules were pGGNs, 109 were heterogeneous GGNs, and 301 were PSNs. The frequency of invasive adenocarcinoma was 39.0% in pGGNs, 67.9% in heterogeneous GGNs, and 75.7% in PSNs (for pGGNs vs heterogeneous GGNs, p < .001; for pGGNs vs PSNs, p < .001; and for heterogeneous GGNs vs PSNs, p = .28). The 5-year recurrence rate was 0.0% in patients with pGGNs, 6.3% in those with heterogeneous GGNs, and 10.8% in those with PSNs (for pGGNs vs heterogeneous GGNs, p = .06; for pGGNs vs PSNs, p = .02; and for heterogeneous GGNs vs PSNs, p = .18). At 7 years, RFS was 97.7% in patients with pGGNs, 82.0% in those with heterogeneous GGNs, and 79.4% in those with PSNs (for pGGNs vs heterogeneous GGNs, p = .02; for pGGNs vs PSNs, p = .006; and for heterogeneous GGNs vs PSNs, p = .40); OS was 98.0% in patients with pGGNs, 84.6% in those with heterogeneous GGNs, and 82.9% in those with PSNs (for pGGNs vs heterogeneous GGNs, p = .04; for pGGNs vs PSNs, p = .01; and for heterogeneous GGNs vs PSNs, p = .50). CONCLUSION. Resected pGGNs had excellent clinical outcomes. Heterogeneous GGNs had relatively worse outcomes, more closely resembling outcomes for PSNs. CLINICAL IMPACT. The findings support surveillance for truly homogeneous pGGNs versus resection for GGNs showing internal increased attenuation even if not having a true solid component.

CXCL13 Positive Cells Localization Predict Response to Anti-PD-1/PD-L1 in Pulmonary Non-Small Cell Carcinoma.

Background: Immune checkpoint inhibitors (ICIs) have revolutionized non-small cell lung cancers (NSCLCs) treatment, but only 20-30% of patients benefit from these treatments. Currently, PD-L1 expression in tumor cells is the only clinically approved predictor of ICI response in lung cancer, but concerns arise due to its low negative and positive predictive value. Recent studies suggest that CXCL13+ T cells in the tumor microenvironment (TME) may be a good predictor of response. We aimed to assess if CXCL13+ cell localization within the TME can predict ICI response in advanced NSCLC patients. Methods: This retrospective study included 65 advanced NSCLC patients treated with Nivolumab/Pembrolizumab at IUCPQ or CHUM and for whom a pretreatment surgical specimen was available. Good responders were defined as having a complete radiologic response at 1 year, and bad responders were defined as showing cancer progression at 1 year. IHC staining for CXCL13 was carried out on a representative slide from a resection specimen, and CXCL13+ cell density was evaluated in tumor (T), invasive margin (IM), non-tumor (NT), and tertiary lymphoid structure (TLS) compartments. Cox models were used to analyze progression-free survival (PFS) and overall survival (OS) probability, while the Mann-Whitney test was used to compare CXCL13+ cell density between responders and non-responders. Results: We showed that CXCL13+ cell density localization within the TME is associated with ICI efficacy. An increased density of CXCL13+ cells across all compartments was associated with a poorer prognostic (OS; HR = 1.22; 95%CI = 1.04-1.42; p = 0.01, PFS; HR = 1.16; p = 0.02), or a better prognostic when colocalized within TLSs (PFS; HR = 0.84, p = 0.03). Conclusion: Our results support the role of CXCL13+ cells in advanced NSCLC patients, with favorable prognosis when localized within TLSs and unfavorable prognosis when present elsewhere. The concomitant proximity of CXCL13+ and CD20+ cells within TLSs may favor antigen presentation to T cells, thus enhancing the effect of PD-1/PD-L1 axis inhibition. Further validation is warranted to confirm the potential relevance of this biomarker in a clinical setting.

Amplification of Wild-Type RET Represents a Novel Molecular Subtype of Several Cancer Types With Clinical Response to Selpercatinib.

PURPOSE: RET rearrangements and RET activating point mutations represent targetable genomic alterations in advanced solid tumors. However, the frequency and clinicopathologic characteristics of wild-type RET amplification in cancer and its potential role as a targetable oncogenic driver are not well-characterized. METHODS: In two institutional cohorts of patients with solid cancers from the Dana-Farber Cancer Institute (DFCI) and Memorial Sloan Kettering Cancer Center (MSKCC) whose tumors underwent next-generation sequencing (NGS), the frequency and clinicopathologic features of wild-type RET amplification in the absence of RET rearrangements or activating mutations was assessed. The findings were validated using merged data from The Cancer Genome Atlas (TCGA), Genomics Evidence Neoplasia Information Exchange (GENIE), and China Pan-Cancer data sets. RESULTS: The frequency of wild-type RET amplification across all solid cancers was 0.08% (26 of 32,505) in the DFCI cohort, 0.05% (26 of 53,152) in the MSKCC cohort, and 0.25% (71 of 28,623) in the cohort from TCGA, GENIE, and China Pan-Cancer. Cancer types with RET amplification included non-small-cell lung cancer (NSCLC), hepatobiliary cancer, prostate cancer, breast cancer, and others. The median RET copy number in RET-amplified cases was 7.5 (range, 6-36) in the DFCI cohort and 5.7 (range, 4-27.7) in the MSKCC cohort. Among 11 RET-amplified NSCLCs, eight had no other concurrent driver mutations. Finally, we report on a 69-year-old man with recurrent NSCLC harboring high-level wild-type RET amplification (22-28 copies) as the only identified putative genomic driver who experienced both a systemic and intracranial confirmed response to the RET inhibitor selpercatinib. CONCLUSION: Amplification of wild-type RET represents a novel, targetable molecular subset of cancer.

Generalization optimizing machine learning to improve CT scan radiomics and assess immune checkpoint inhibitors' response in non-small cell lung cancer: a multicenter cohort study.

Background: Recent developments in artificial intelligence suggest that radiomics may represent a promising non-invasive biomarker to predict response to immune checkpoint inhibitors (ICIs). Nevertheless, validation of radiomics algorithms in independent cohorts remains a challenge due to variations in image acquisition and reconstruction. Using radiomics, we investigated the importance of scan normalization as part of a broader machine learning framework to enable model external generalizability to predict ICI response in non-small cell lung cancer (NSCLC) patients across different centers. Methods: Radiomics features were extracted and compared from 642 advanced NSCLC patients on pre-ICI scans using established open-source PyRadiomics and a proprietary DeepRadiomics deep learning technology. The population was separated into two groups: a discovery cohort of 512 NSCLC patients from three academic centers and a validation cohort that included 130 NSCLC patients from a fourth center. We harmonized images to account for variations in reconstruction kernel, slice thicknesses, and device manufacturers. Multivariable models, evaluated using cross-validation, were used to estimate the predictive value of clinical variables, PD-L1 expression, and PyRadiomics or DeepRadiomics for progression-free survival at 6 months (PFS-6). Results: The best prognostic factor for PFS-6, excluding radiomics features, was obtained with the combination of Clinical + PD-L1 expression (AUC = 0.66 in the discovery and 0.62 in the validation cohort). Without image harmonization, combining Clinical + PyRadiomics or DeepRadiomics delivered an AUC = 0.69 and 0.69, respectively, in the discovery cohort, but dropped to 0.57 and 0.52, in the validation cohort. This lack of generalizability was consistent with observations in principal component analysis clustered by CT scan parameters. Subsequently, image harmonization eliminated these clusters. The combination of Clinical + DeepRadiomics reached an AUC = 0.67 and 0.63 in the discovery and validation cohort, respectively. Conversely, the combination of Clinical + PyRadiomics failed generalizability validations, with AUC = 0.66 and 0.59. Conclusion: We demonstrated that a risk prediction model combining Clinical + DeepRadiomics was generalizable following CT scan harmonization and machine learning generalization methods. These results had similar performances to routine oncology practice using Clinical + PD-L1. This study supports the strong potential of radiomics as a future non-invasive strategy to predict ICI response in advanced NSCLC.

A Simplified Version of the IASLC Grading System for Invasive Pulmonary Adenocarcinomas With Improved Prognosis Discrimination.

Tumor grading enables better management of patients and treatment options. The International Association for the Study of Lung Cancer (IASLC) Pathology Committee has recently released a 3-tier grading system for invasive pulmonary adenocarcinoma consisting of predominant histologic patterns plus a cutoff of 20% of high-grade components including solid, micropapillary, and complex glandular patterns. The goal of this study was to validate the prognostic value of the new IASLC grading system and to compare its discriminatory performance to the predominant pattern-based grading system and a simplified version of the IASLC grading system without complex glandular patterns. This was a single-site retrospective study based on a 20-year data collection of patients that underwent lung cancer surgery. All invasive pulmonary adenocarcinomas confirmed by the histologic review were evaluated in a discovery cohort (n=676) and a validation cohort (n=717). The median duration of follow-up in the combined dataset (n=1393) was 7.5 years. The primary outcome was overall survival after surgery. The 3 grading systems had strong and relatively similar predictive performance, but the best parsimonious model was the simplified IASLC grading system (log-rank P =1.39E-13). The latter was strongly associated with survival in the validation set ( P =1.1E-18) and the combined set ( P =5.01E-35). We observed a large proportion of patients upgraded to the poor prognosis group using the IASLC grading system, which was attenuated when using the simplified IASLC grading system. In conclusion, we identified a histologic simpler classification for invasive pulmonary adenocarcinomas that outperformed the recently proposed IASLC grading system. A simplified grading system is clinically convenient and will facilitate widespread implementation.

Single-cell spatial landscape of immunotherapy response reveals mechanisms of CXCL13 enhanced antitumor immunity.

BACKGROUND: Immunotherapy has revolutionized clinical outcomes for patients suffering from lung cancer, yet relatively few patients sustain long-term durable responses. Recent studies have demonstrated that the tumor immune microenvironment fosters tumorous heterogeneity and mediates both disease progression and response to immune checkpoint inhibitors (ICI). As such, there is an unmet need to elucidate the spatially defined single-cell landscape of the lung cancer microenvironment to understand the mechanisms of disease progression and identify biomarkers of response to ICI. METHODS: Here, in this study, we applied imaging mass cytometry to characterize the tumor and immunological landscape of immunotherapy response in non-small cell lung cancer by describing activated cell states, cellular interactions and neighborhoods associated with improved efficacy. We functionally validated our findings using preclinical mouse models of cancer treated with anti-programmed cell death protein-1 (PD-1) immune checkpoint blockade. RESULTS: We resolved 114,524 single cells in 27 patients treated with ICI, enabling spatial resolution of immune lineages and activation states with distinct clinical outcomes. We demonstrated that CXCL13 expression is associated with ICI efficacy in patients, and that recombinant CXCL13 potentiates anti-PD-1 response in vivo in association with increased antigen experienced T cell subsets and reduced CCR2+ monocytes. DISCUSSION: Our results provide a high-resolution molecular resource and illustrate the importance of major immune lineages as well as their functional substates in understanding the role of the tumor immune microenvironment in response to ICIs.

Single-cell spatial landscapes of the lung tumour immune microenvironment.

Single-cell technologies have revealed the complexity of the tumour immune microenvironment with unparalleled resolution1-9. Most clinical strategies rely on histopathological stratification of tumour subtypes, yet the spatial context of single-cell phenotypes within these stratified subgroups is poorly understood. Here we apply imaging mass cytometry to characterize the tumour and immunological landscape of samples from 416 patients with lung adenocarcinoma across five histological patterns. We resolve more than 1.6 million cells, enabling spatial analysis of immune lineages and activation states with distinct clinical correlates, including survival. Using deep learning, we can predict with high accuracy those patients who will progress after surgery using a single 1-mm2 tumour core, which could be informative for clinical management following surgical resection. Our dataset represents a valuable resource for the non-small cell lung cancer research community and exemplifies the utility of spatial resolution within single-cell analyses. This study also highlights how artificial intelligence can improve our understanding of microenvironmental features that underlie cancer progression and may influence future clinical practice.

Prognostic value of complex glandular patterns in invasive pulmonary adenocarcinomas.

Prognostic stratification of patients surgically resected with invasive pulmonary adenocarcinoma must be improved. Previous studies reported that complex glandular patterns (CGPs), cribriform and fused gland growth patterns, are associated with unfavorable prognosis. The goal of this study is to evaluate the prognostic value of CGPs in patients with resected stage I-IV lung adenocarcinoma. The presence of CGPs as a minor to predominant component was tested for association with overall survival (OS, n = 676) and relapse-free survival (RFS, n = 463) after surgery. CGPs were observed in 284 tumors (42.0%). Cribriform and fused gland were the predominant patterns in 35 and 37 cases, respectively. The presence of cribriform pattern was associated with worse RFS, but not OS. The fused gland pattern alone or grouped into CGPs with the cribriform pattern was not associated with OS and RFS. As a predominant pattern, cribriform was associated with the worse survival compared to the 5 recognized histologic patterns. Patients with fused gland-predominant tumors had 5-year survival that ranged between papillary- and micropapillary-predominant tumors. We conclude that cribriform-predominant, but not fused gland-predominant, is a subtype with poor prognosis similar to the solid and micropapillary subtypes. In contrast, the presence of a minor component of fused gland or CGPs (cribriform + fused gland) is not associated with survival. The cribriform pattern alone offers prognosis stratification improvement, but this effect is attenuated when combined into CGPs to define a subset of acinar-predominant tumors with poor prognosis. This argues against combining cribriform and fused gland into CGPs to summarize high-grade patterns.

Wait Times and Survival in Lung Cancer Patients across the Province of Quebec, Canada.

Lung cancer is the leading cause of cancer death worldwide, with a five-year survival of 22% in Canada. Guidelines recommend rapid evaluation of patients with suspected lung cancer, but the impact on survival remains unclear. We reviewed medical records of all patients with newly diagnosed lung cancer in four hospital networks across the province of Quebec, Canada, between 1 February and 30 April 2017. Patients were followed for 3 years. Wait times for diagnosis and treatment were collected, and survival analysis using a Cox regression model was conducted. We included 1309 patients, of whom 39% had stage IV non-small cell lung cancer (NSCLC). Median wait times were, in general, significantly shorter in patients with stage III-IV NSCLC or SCLC. Surgery was associated with delays compared to other types of treatments. Median survival was 12.9 (11.1-15.7) months. The multivariate survival model included age, female sex, performance status, histology and stage, treatment, and the time interval between diagnosis and treatment. Longer wait times had a slightly protective to neutral effect on survival, but this was not significant in the stage I-II NSCLC subgroup. Wait times for the diagnosis and treatment of lung cancer were generally within targets. The shorter wait times observed for advanced NSCLC and SCLC might indicate a tendency for clinicians to act quicker on sicker patients. This study did not demonstrate the detrimental effect of longer wait times on survival.

Gut microbiota and fermentation-derived branched chain hydroxy acids mediate health benefits of yogurt consumption in obese mice.

Meta-analyses suggest that yogurt consumption reduces type 2 diabetes incidence in humans, but the molecular basis of these observations remains unknown. Here we show that dietary yogurt intake preserves whole-body glucose homeostasis and prevents hepatic insulin resistance and liver steatosis in a dietary mouse model of obesity-linked type 2 diabetes. Fecal microbiota transplantation studies reveal that these effects are partly linked to the gut microbiota. We further show that yogurt intake impacts the hepatic metabolome, notably maintaining the levels of branched chain hydroxy acids (BCHA) which correlate with improved metabolic parameters. These metabolites are generated upon milk fermentation and concentrated in yogurt. Remarkably, diet-induced obesity reduces plasma and tissue BCHA levels, and this is partly prevented by dietary yogurt intake. We further show that BCHA improve insulin action on glucose metabolism in liver and muscle cells, identifying BCHA as cell-autonomous metabolic regulators and potential mediators of yogurt's health effects.

Profiles of teacher-child interaction quality in groups of 3-year-old children in Quebec and France.

Theory and studies support that educational quality may differ according to socio-political context even in states with similar cultures. Based on a secondary analysis of data, this study aims at identifying latent profiles of adult-child interaction quality in groups of three-year-old children in Quebec's (Canada) early childhood centers and France's kindergarten classrooms using the CLASS Pre-K. This study also aims to explore existing associations between identified profiles, socio-political contexts, and structural characteristics (staff qualifications, ages, group size). Latent profile analyses showed four interaction quality profiles, namely a high-quality profile (HQ), a medium-high-quality profile (MHQ), a medium quality profile (MQ), and a medium-low-quality profile (MLQ). The scores of the three CLASS Pre-K domains associated with identified profiles show a higher average interaction quality in Quebec compared with France, suggesting a more favorable sociocultural context for interaction quality in Quebec. As for characteristics of structural quality, analyses suggest that the group size variable is significantly associated with scores of interaction quality, with the HQ and the MHQ profiles showing a significantly lower group size than the MQ and MLQ profiles. Age is also significantly associated with profiles, exhibiting a general trend of younger participants found in higher quality profiles. Courses of action to enhance French policies are discussed.

ZNF768 links oncogenic RAS to cellular senescence.

RAS proteins are GTPases that lie upstream of a signaling network impacting cell fate determination. How cells integrate RAS activity to balance proliferation and cellular senescence is still incompletely characterized. Here, we identify ZNF768 as a phosphoprotein destabilized upon RAS activation. We report that ZNF768 depletion impairs proliferation and induces senescence by modulating the expression of key cell cycle effectors and established p53 targets. ZNF768 levels decrease in response to replicative-, stress- and oncogene-induced senescence. Interestingly, ZNF768 overexpression contributes to bypass RAS-induced senescence by repressing the p53 pathway. Furthermore, we show that ZNF768 interacts with and represses p53 phosphorylation and activity. Cancer genomics and immunohistochemical analyses reveal that ZNF768 is often amplified and/or overexpressed in tumors, suggesting that cells could use ZNF768 to bypass senescence, sustain proliferation and promote malignant transformation. Thus, we identify ZNF768 as a protein linking oncogenic signaling to the control of cell fate decision and proliferation.

ZNF768 Expression Associates with High Proliferative Clinicopathological Features in Lung Adenocarcinoma.

Lung adenocarcinoma (LUAD) is the most common type of lung cancer and a leading cause of cancer-related deaths worldwide. Despite important recent advances, the prognosis for LUAD patients is still unfavourable, with a 5 year-survival rate close to 15%. Improving the characterization of lung tumors is important to develop alternative options for the diagnosis and the treatment of this disease. Zinc-finger protein 768 (ZNF768) is a transcription factor that was recently shown to promote proliferation and repress senescence downstream of growth factor signaling. Although ZNF768 protein levels were found to be elevated in LUAD compared to normal lung tissue, it is currently unknown whether ZNF768 expression associates with clinicopathological features in LUAD. Here, using tissue microarrays of clinical LUAD surgical specimens collected from 364 patients, we observed that high levels of ZNF768 is a common characteristic of LUAD. We show that ZNF768 protein levels correlate with high proliferative features in LUAD, including the mitotic score and Ki-67 expression. Supporting a role for ZNF768 in promoting proliferation, we report that ZNF768 depletion severely impairs proliferation in several lung cancer cell lines in vitro. A marked decrease in the expression of key proliferative genes was observed in cancer cell lines depleted from ZNF768. Altogether, our findings support a role for ZNF768 in promoting proliferation of LUAD.

Venous thrombotic events in patients treated with immune checkpoint inhibitors for non-small cell lung cancer: A retrospective multicentric cohort study.

OBJECTIVES: Venous thrombotic events (VTEs) are a frequent complication of non-small cell lung cancer (NSCLC) and are associated with increased morbidity. Immune checkpoint inhibitors (ICIs) are revolutionizing the management of NSCLC, but little is known about their impact on thrombosis. This study aims to define the incidence and clinical relevance of VTEs in NSCLC patients receiving these treatments. METHODS: A retrospective multicentric cohort study including 593 patients from three centers in Canada and France was performed. The cumulative incidence of VTEs after ICIs was estimated using competing risk analysis, and the association of these events with survival and response to treatment was determined. Finally, univariate and multivariate tests were performed to identify VTE risk factors. RESULTS: The cumulative incidence of VTEs in the cohort was 14.8% (95% CI = 7.4-22.2%) for an incidence rate of 76.5 (95% CI = 59.9-97.8) thrombosis per 1000 person-years, with most thromboses occurring rapidly after treatment initiation. VTEs were not correlated with overall survival, progression-free survival, or objective response to ICIs. Age ˂ 65 years old (HR = 2.00; 95% CI = 1.11-3.59) and tumors with PD-L1 1-49% (HR = 3.36; 95% CI = 1.19-9.50) or PD-L1 ≥ 50% (HR = 3.22; 95% CI = 1.21-8.57) were associated with more VTEs after 12 months of ICI initiation. Also, a delay of less than 12 months from diagnosis to the first ICI treatment (HR = 2.06; 95% CI = 1.09-3.89) and active smoking (HR = 2.00; 95% CI = 1.12-3.58) are probable risk factors of VTEs. CONCLUSION: This study suggests that the incidence of VTEs in NSCLC patients treated with ICIs is comparable to what is reported in other cohorts of patients treated with chemotherapy. In our cohort, VTEs were not associated with a decreased survival or response to therapy. Patient age < 65 and tumors with PD-L1 ≥ 1% were associated with a higher risk of VTEs under ICIs.

Unravelling actionable biology using transcriptomic data to integrate mitotic index and Ki-67 in the management of lung neuroendocrine tumors.

Pulmonary neuroendocrine tumors (NETs) are a heterogeneous family of malignancies whose classification relies on morphology and mitotic rate, unlike extrapulmonary neuroendocrine tumors that require both mitotic rate and Ki-67. As mitotic count is proportional to Ki-67, it is crucial to understand if Ki-67 can complement the existing diagnostic guidelines, as well as discover the benefit of these two markers to unravel the biological heterogeneity. In this study, we investigated the association of mitotic rate and Ki-67 at gene- and pathway-level using transcriptomic data in lung NET malignancies. Lung resection tumor specimens obtained from 28 patients diagnosed with NETs were selected. Mitotic rate, Ki-67 and transcriptomic data were obtained for all samples. The concordance between mitotic rate and Ki-67 was evaluated at gene-level and pathway-level using gene expression data. Our analysis revealed a strong association between mitotic rate and Ki-67 across all samples and cell cycle genes were found to be differentially ranked between them. Pathway analysis indicated that a greater number of pathways overlapped between these markers. Analyses based on lung NET subtypes revealed that mitotic rate in carcinoids and Ki-67 in large cell neuroendocrine carcinomas provided comprehensive characterization of pathways among these malignancies. Among the two subtypes, we found distinct leading-edge gene sets that drive the enrichment signal of commonly enriched pathways between mitotic index and Ki-67. Overall, our findings delineated the degree of benefit of the two proliferation markers, and offers new layer to predict the biological behavior and identify high-risk patients using a more comprehensive diagnostic workup.

Cross-reactivity between tumor MHC class I-restricted antigens and an enterococcal bacteriophage.

Intestinal microbiota have been proposed to induce commensal-specific memory T cells that cross-react with tumor-associated antigens. We identified major histocompatibility complex (MHC) class I-binding epitopes in the tail length tape measure protein (TMP) of a prophage found in the genome of the bacteriophage Enterococcus hirae Mice bearing E. hirae harboring this prophage mounted a TMP-specific H-2Kb-restricted CD8+ T lymphocyte response upon immunotherapy with cyclophosphamide or anti-PD-1 antibodies. Administration of bacterial strains engineered to express the TMP epitope improved immunotherapy in mice. In renal and lung cancer patients, the presence of the enterococcal prophage in stools and expression of a TMP-cross-reactive antigen by tumors correlated with long-term benefit of PD-1 blockade therapy. In melanoma patients, T cell clones recognizing naturally processed cancer antigens that are cross-reactive with microbial peptides were detected.

Comprehensive assessment of PD-L1 immunohistochemistry on paired tissue and cytology specimens from non-small cell lung cancer.

OBJECTIVES: PD-L1 staining assessed by immunohistochemistry (IHC) is a predictive biomarker used to select advanced stage non-small cell lung carcinoma (NSCLC) patients who are likely to respond to PD-1/PD-L1 inhibitors. Cytology specimens represent a significant percentage of the diagnostic samples and additional data are required to show that they provide reliable PD-L1 results when compared to tissue specimens. We aimed to compare PD-L1 staining obtained from patient-matched tissue and cytology specimens. We also want to assess the feasibility of PD-L1 testing on cell blocks with two assays by evaluating the intra- and inter-observer agreement and the level of difficulty for determining the percentage of stained tumor cells (TPS). MATERIALS AND METHODS: Forty-six patients with NSCLC were selected. Each patient provided a surgical specimen and a cytology sample (cell block) and/or a biopsy at diagnosis. PD-L1 staining using Agilent PD-L1 IHC 28-8 pharmDx and VENTANA PD-L1 (SP263) assays was evaluated by four pathologists using the TPS. Sixty slides were rescored to document intra-observer agreement. Pathologists were asked to score the level of difficulty for evaluating PD-L1 TPS for each slide. Fleiss's and Cohen's kappas (k) were used to assess the agreement between paired specimens as well as intra- and inter-observer agreement. RESULTS: The concordance in PD-L1 TPS between cell blocks and surgical specimens (k varying from 0.56 to 0.82) or biopsies (k from 0.43 to 0.81) was moderate to substantial, depending on the cut-off. On cell blocks, inter-observer agreement was substantial (k from 0.74 to 0.82) and intra-observer agreement was almost perfect (k from 0.85 to 0.93). The perceived difficulty of PD-L1 evaluation of cell blocks was not different from surgical specimens but more difficult than biopsy samples. CONCLUSION: PD-L1 TPS was concordant between cell blocks and tissue specimens, mainly at 10, 25 and 50 % cut-offs. PD-L1 evaluation on cell blocks was feasible and reproducible between different observers and assays.

Dietary sucrose induces metabolic inflammation and atherosclerotic cardiovascular diseases more than dietary fat in LDLr-/-ApoB100/100 mice.

BACKGROUND AND AIMS: Poor dietary habits contribute to the obesity pandemic and related cardiovascular diseases but the respective impact of high saturated fat versus added sugar consumption remains debated. Herein, we aimed to disentangle the individual role of dietary fat versus sugar in cardiometabolic disease progression. METHODS: We fed pro-atherogenic LDLr-/-ApoB100/100 mice either a low-fat/high-sucrose (LFHS) or a high-fat/low-sucrose (HFLS) diet for 24 weeks. Weekly body weight gain was registered. 16S rRNA gene-based gut microbial analysis was performed to investigate gut microbial modulations. Intraperitoneal insulin (ipITT) and oral glucose tolerance test (oGTT) were conducted to assess glucose homeostasis and insulin sensitivity. Cytokines were assessed in fasted plasma, epididymal white adipose tissue and liver lysates. Heart function was evaluated by echocardiography. Aortic atheroma lesions were quantified according to the en face technique. RESULTS: HFLS feeding increased obesity, insulin resistance and dyslipidemia compared to LFHS feeding. Conversely, high sucrose consumption decreased gut microbial diversity while augmenting inflammation and the adaptative immune defense against metabolic endotoxemia and reduced macrophage cholesterol efflux capacity. This led to more severe cardiovascular complications as revealed by remarkably high level of atherosclerotic lesions and the early development of cardiac dysfunction in LFHS vs HFLS fed mice. CONCLUSIONS: We uncoupled obesity-associated insulin resistance from cardiovascular diseases and provided novel evidence that dietary sucrose, not fat, is the main driver of metabolic inflammation accelerating severe atherosclerosis in hyperlipidemic mice.