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Supplementary Table #1

Early detection of pancreatic ductal adenocarcinoma (PDAC) is key to improving patient outcomes; however, PDAC is usually diagnosed late. Therefore, blood-based minimally invasive biomarker assays for limited volume clinical samples are urgently needed. A novel miRNA profiling platform (Abcam Fireplex-Oncology Panel) was used to investigate the feasibility of developing early detection miRNA biomarkers with 20 μL plasma from a training set (58 stage II PDAC cases and 30 controls) and two validation sets (34 stage II PDAC cases and 25 controls; 44 stage II PDAC cases and 18 controls). miR-34a-5p [AUC = 0.77; 95% confidence interval (CI), 0.66–0.87], miR-130a-3p (AUC = 0.74; 95% CI, 0.63–0.84), and miR-222-3p (AUC = 0.70; 95% CI, 0.58–0.81) were identified as significantly differentially abundant in plasma from stage II PDAC versus controls. Although none of the miRNAs individually outperformed the currently used serologic biomarker for PDAC, carbohydrate antigen 19-9 (CA19-9), combining the miRNAs with CA 19-9 improved AUCs from 0.89 (95% CI, 0.81–0.95) for CA 19-9 alone to 0.92 (95% CI, 0.86–0.97), 0.94 (95% CI, 0.89–0.98), and 0.92 (95% CI, 0.87–0.97), respectively. Gene set enrichment analyses of transcripts correlated with high and low expression of the three miRNAs in The Cancer Genome Atlas PDAC sample set. These miRNA biomarkers, assayed in limited volume plasma together with CA19-9, discriminate stage II PDAC from controls with good sensitivity and specificity. Unbiased profiling of larger cohorts should help develop an informative early detection biomarker assay for diagnostic settings.Prevention Relevance:Development of minimally invasive biomarker assays for detection of premalignant disease and early-stage pancreatic cancer is key to improving patient survival. This study describes a limited volume plasma miRNA biomarker assay that can detect early-stage resectable pancreatic cancer in clinical samples necessary for effective prevention and clinical intervention.

Supplemental Figure #1

PURPOSE With the development of poly (ADP-ribose) polymerase inhibitors for treatment of patients with cancer with an altered BRCA1 or BRCA2 gene, there is an urgent need to ensure that there are appropriate strategies for identifying mutation carriers while balancing the increased demand for and cost of cancer genetics services. To date, the majority of mutation prediction tools have been developed in women of European descent where the age and cancer-subtype distributions are different from that in Asian women. METHODS In this study, we built a new model (Asian Risk Calculator) for estimating the likelihood of carrying a pathogenic variant in BRCA1 or BRCA2 gene, using germline BRCA genetic testing results in a cross-sectional population-based study of 8,162 Asian patients with breast cancer. We compared the model performance to existing mutation prediction models. The models were evaluated for discrimination and calibration. RESULTS Asian Risk Calculator included age of diagnosis, ethnicity, bilateral breast cancer, tumor biomarkers, and family history of breast cancer or ovarian cancer as predictors. The inclusion of tumor grade improved significantly the model performance. The full model was calibrated (Hosmer-Lemeshow P value = .614) and discriminated well between BRCA and non- BRCA pathogenic variant carriers (area under receiver operating curve, 0.80; 95% CI, 0.75 to 0.84). Addition of grade to the existing clinical genetic testing criteria targeting patients with breast cancer age younger than 45 years reduced the proportion of patients referred for genetic counseling and testing from 37% to 33% ( P value = .003), thereby improving the overall efficacy. CONCLUSION Population-specific customization of mutation prediction models and clinical genetic testing criteria improved the accuracy of BRCA mutation prediction in Asian patients.

Purpose Non-European populations are under-represented in genetics studies, hindering clinical implementation of breast cancer polygenic risk scores (PRSs). We aimed to develop PRSs using the largest available studies of Asian ancestry and to assess the transferability of PRS across ethnic subgroups. Methods The development data set comprised 138,309 women from 17 case-control studies. PRSs were generated using a clumping and thresholding method, lasso penalized regression, an Empirical Bayes approach, a Bayesian polygenic prediction approach, or linear combinations of multiple PRSs. These PRSs were evaluated in 89,898 women from 3 prospective studies (1592 incident cases). Results The best performing PRS (genome-wide set of single-nucleotide variations [formerly single-nucleotide polymorphism]) had a hazard ratio per unit SD of 1.62 (95% CI = 1.46-1.80) and an area under the receiver operating curve of 0.635 (95% CI = 0.622-0.649). Combined Asian and European PRSs (333 single-nucleotide variations) had a hazard ratio per SD of 1.53 (95% CI = 1.37-1.71) and an area under the receiver operating curve of 0.621 (95% CI = 0.608-0.635). The distribution of the latter PRS was different across ethnic subgroups, confirming the importance of population-specific calibration for valid estimation of breast cancer risk. Conclusion PRSs developed in this study, from association data from multiple ancestries, can enhance risk stratification for women of Asian ancestry.

Background Approximately 20% of lung adenocarcinoma (LUAD) is negative for the lineage-specific oncogene Thyroid transcription factor 1 (TTF-1) and exhibits worse clinical outcome with a low frequency of actionable genomic alterations. To identify molecular features associated with TTF-1–negative LUAD, we compared the transcriptomic and proteomic profiles of LUAD cell lines. SRGN , a chondroitin sulfate proteoglycan Serglycin, was identified as a markedly overexpressed gene in TTF-1–negative LUAD. We therefore investigated the roles and regulation of SRGN in TTF-1–negative LUAD. Methods Proteomic and metabolomic analyses of 41 LUAD cell lines were done using mass spectrometry. The function of SRGN was investigated in 3 TTF-1–negative and 4 TTF-1–positive LUAD cell lines and in a syngeneic mouse model (n = 5 to 8 mice per group). Expression of SRGN was evaluated in 94 and 105 surgically resected LUAD tumor specimens using immunohistochemistry. All statistical tests were 2-sided. Results SRGN was markedly overexpressed at mRNA and protein levels in TTF-1–negative LUAD cell lines (P < .001 for both mRNA and protein levels). Expression of SRGN in LUAD tumor tissue was associated with poor outcome (hazard ratio = 4.22, 95% confidence interval = 1.12 to 15.86, likelihood ratio test, P = .03), and with higher expression of Programmed cell death 1 ligand 1 (PD-L1) in tumor cells and higher infiltration of Programmed cell death protein 1–positive lymphocytes. SRGN regulated expression of PD-L1 as well as proinflammatory cytokines, including Interleukin-6, Interleukin-8, and C-X-C motif chemokine 1 in LUAD cell lines; increased migratory and invasive properties of LUAD cells and fibroblasts; and enhanced angiogenesis. SRGN was induced by DNA demethylation resulting from Nicotinamide N-methyltransferase–mediated impairment of methionine metabolism. Conclusions Our findings suggest that SRGN plays a pivotal role in tumor–stromal interaction and reprogramming into an aggressive and immunosuppressive tumor microenvironment in TTF-1–negative LUAD.

Background: Early-stage lung adenocarcinoma (LUAD) patients have substantial risk for recurrence and disease-related death. Cisplatin-based adjuvant chemotherapy remains the standard for care of LUAD patients who have undergone surgical resection with a high risk of recurrence. However, adjuvant chemotherapy is associated with increased risk of toxicity including chemotherapy-related death, with only a modest survival benefit. Therefore, there is an unmet need of biomarkers for assessment and identification of those in an early stage who would likely benefit from adjuvant chemotherapy. Circulating miRNAs are stably present in blood and potentially reflect different expressions in cancerous and non-cancerous tissues, making them attractive biomarkers. The purpose of this study was to identify circulating miRNAs useful for predicting recurrence in early-stage LUAD. Materials and Methods: miRNA microarray analysis was performed with pooled pretreatment plasma samples from stage I LUAD patients who developed recurrence within two years after curative surgery or remained recurrence free over a six-year follow-up period, as well as from healthy controls. miRNA biomarker candidates were assayed in two independent plasma sample sets from 85 stage I LUAD (validation set) and from 57 stage I and II LUAD (test set) patients. Results: Based on miRNA microarray data and previous reports, predictive performance of miR-23a-3p, miR-23b-3p, miR-191-5p, miR-185-5p, miR-151a-3p, miR-320c, miR-21-5p, miR-125b-5p, miR-30d-5p, and miR-197-3p was evaluated in the validation set. Plasma levels of miR-23a-3p, miR-185-5p, miR-320c, miR-21-5p, miR-125b-5p, miR-30d-5p, and miR-197-3p were significantly higher in those with recurrence as compared to those without. A miRNA panel comprised of miR-23a-3p, miR-320c, and miR-125b-5p was developed based on a logistic regression, with yielding an AUC of 0.776 (95% confidence interval [CI] = 0.660 to 0.893). The three-miRNA panel with fixed coefficients yielded an AUC of 0.804 (95% CI = 0.688 to 0.920) with a sensitivity of 45.8% at 95% specificity in the test set. The miRNA panel score was a significant and independent factor for predicting disease-free survival (DFS; P < 0.001, HR = 1.64, 95% CI = 1.51-4.22) and overall survival (OS; P = 0.001, HR = 1.51, 95% CI = 1.17-1.94). Conclusion: This circulating miRNA panel may serve as a noninvasive blood test for predicting DFS and OS in early-stage LUAD patients. Our findings provide rationale for further investigation to stratify early-stage LUAD patients using blood-based biomarkers to increase the ability to provide more personalized care. Citation Format: Mei-Chee Tai, Leonidas E. Bantis, Gargy Parhy, Taketo Kato, Ichidai Tanaka, Chi-Wan Chow, Junya Fujimoto, Carmen Behrens, Tetsunari Hase, Koji Kawaguchi, Johannes F. Fahrmann, Edwin Ostrin, Kohei Yokoi, Toyofumi F. Chen-Yoshikawa, Yoshinori Hasegawa, Samir M. Hanash, Ignacio I. Wistuba, Ayumu Taguchi. A circulating microRNA panel predicts recurrence and survival in early-stage lung 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 3744.

Purpose Conventional chemotherapy and enucleation usually fail to cure advanced retinoblastoma. We investigated the retinoblastoma immune microenvironment and the efficacy of the combination of dinutuximab and CD16-expressing NK-92MI (NK-92MIhCD16-GFP) cells on retinoblastoma cells in this study. Patients and methods Immunohistochemistry and flow cytometry (FC) were performed to assess the expression level of GD2 in retinoblastoma tissues and cells. Gene set enrichment analysis (GSEA), immunohistochemisrztry and immunocytochemistry were conducted to assess the retinoblastoma immune microenvironment and the integrity of the blood-retinal barrier (BRB). After overexpressing CD16 in NK-92MI cells, fluorescence-activated cell sorting (FACS) was applied to select the positive subpopulation. LDH assays and FC were used to detect LDH release and apoptosis in retinoblastoma cells subjected to a combination of dinutuximab and NK-92MIhCD16-GFP cells. Finally, the release of perforin-granzyme B and the expression of CD107a in NK-92MIhCD16-GFP stimulated by retinoblastoma cells were assessed via enzyme-linked immunosorbent assays (ELISAs) and FC in the presence of dinutuximab or an isotype control. Results GD2 was heterogeneously expressed in retinoblastoma tissues and cell lines and positively correlated with proliferation and staging. GSEA revealed the immunosuppressive status of retinoblastoma microenvironment. The immune cell profile of retinoblastoma tissues and vitreous bodies suggested BRB destruction. LDH release and apoptosis in retinoblastoma cells caused by NK-92MIhCD16-GFP cells were significantly enhanced by dinutuximab. Finally, the release of perforin-granzyme B and the expression of CD107a in NK-92MIhCD16-GFP cells stimulated by retinoblastoma cells were obviously increased by dinutuximab. Conclusion This study indicates that retinoblastoma impairs the integrity of the BRB and contributes to dysregulated immune cell infiltrates. GD2 is a specific target for natural killer (NK) cell-based immunotherapy and that the combination of dinutuximab and NK-92MIhCD16-GFP cells exerts potent antitumor effects through antibody-dependent cell-mediated cytotoxicity.

Background Family history, and genetic and non-genetic risk factors can stratify women according to their individual risk of developing breast cancer. The extent of overlap between these risk predictors is not clear. Methods In this case-only analysis involving 7600 Asian breast cancer patients diagnosed between age 30 and 75 years, we examined identification of high-risk patients based on positive family history, the Gail model 5-year absolute risk [5yAR] above 1.3%, breast cancer predisposition genes (protein-truncating variants [PTV] in ATM, BRCA1, BRCA2, CHEK2, PALB2, BARD1, RAD51C, RAD51D, or TP53), and polygenic risk score (PRS) 5yAR above 1.3%. Results Correlation between 5yAR (at age of diagnosis) predicted by PRS and the Gail model was low (r=0.27). Fifty-three percent of breast cancer patients (n=4041) were considered high risk by one or more classification criteria. Positive family history, PTV carriership, PRS, or the Gail model identified 1247 (16%), 385 (5%), 2774 (36%), and 1592 (21%) patients who were considered at high risk, respectively. In a subset of 3227 women aged below 50 years, the four models studied identified 470 (15%), 213 (7%), 769 (24%), and 325 (10%) unique patients who were considered at high risk, respectively. For younger women, PRS and PTVs together identified 745 (59% of 1276) high-risk individuals who were not identified by the Gail model or family history. Conclusions Family history and genetic and non-genetic risk stratification tools have the potential to complement one another to identify women at high risk.

Background Mutations in certain genes are known to increase breast cancer risk. We study the relevance of rare protein-truncating variants (PTVs) that may result in loss-of-function in breast cancer susceptibility genes on tumor characteristics and survival in 8852 breast cancer patients of Asian descent. Methods Gene panel sequencing was performed for 34 known or suspected breast cancer predisposition genes, of which nine genes (ATM, BRCA1, BRCA2, CHEK2, PALB2, BARD1, RAD51C, RAD51D, and TP53) were associated with breast cancer risk. Associations between PTV carriership in one or more genes and tumor characteristics were examined using multinomial logistic regression. Ten-year overall survival was estimated using Cox regression models in 6477 breast cancer patients after excluding older patients (≥75years) and stage 0 and IV disease. Results PTV9genes carriership (n = 690) was significantly associated (p < 0.001) with more aggressive tumor characteristics including high grade (poorly vs well-differentiated, odds ratio [95% confidence interval] 3.48 [2.35–5.17], moderately vs well-differentiated 2.33 [1.56–3.49]), as well as luminal B [HER−] and triple-negative subtypes (vs luminal A 2.15 [1.58–2.92] and 2.85 [2.17–3.73], respectively), adjusted for age at diagnosis, study, and ethnicity. Associations with grade and luminal B [HER2−] subtype remained significant after excluding BRCA1/2 carriers. PTV25genes carriership (n = 289, excluding carriers of the nine genes associated with breast cancer) was not associated with tumor characteristics. However, PTV25genes carriership, but not PTV9genes carriership, was suggested to be associated with worse 10-year overall survival (hazard ratio [CI] 1.63 [1.16–2.28]). Conclusions PTV9genes carriership is associated with more aggressive tumors. Variants in other genes might be associated with the survival of breast cancer patients. The finding that PTV carriership is not just associated with higher breast cancer risk, but also more severe and fatal forms of the disease, suggests that genetic testing has the potential to provide additional health information and help healthy individuals make screening decisions.

Polygenic risk scores (PRS) have been shown to predict breast cancer risk in European women, but their utility in Asian women is unclear. Here we evaluate the best performing PRSs for European-ancestry women using data from 17,262 breast cancer cases and 17,695 controls of Asian ancestry from 13 case-control studies, and 10,255 Chinese women from a prospective cohort (413 incident breast cancers). Compared to women in the middle quintile of the risk distribution, women in the highest 1% of PRS distribution have a ~2.7-fold risk and women in the lowest 1% of PRS distribution has ~0.4-fold risk of developing breast cancer. There is no evidence of heterogeneity in PRS performance in Chinese, Malay and Indian women. A PRS developed for European-ancestry women is also predictive of breast cancer risk in Asian women and can help in developing risk-stratified screening programmes in Asia., Polygenic risk scores predict the likelihood that an individual will develop a certain cancer, however these are often specific for a given population. Here, the authors show that a risk score developed to assess the risk of breast cancer in European women can also predict risk in Asian populations.

Early detection of pancreatic ductal adenocarcinoma (PDAC) is key to improving patient outcomes; however, PDAC is usually diagnosed late. Therefore, blood-based minimally invasive biomarker assays for limited volume clinical samples are urgently needed. A novel miRNA profiling platform (Abcam Fireplex-Oncology Panel) was used to investigate the feasibility of developing early detection miRNA biomarkers with 20 μL plasma from a training set (58 stage II PDAC cases and 30 controls) and two validation sets (34 stage II PDAC cases and 25 controls; 44 stage II PDAC cases and 18 controls). miR-34a-5p [AUC = 0.77; 95% confidence interval (CI), 0.66–0.87], miR-130a-3p (AUC = 0.74; 95% CI, 0.63–0.84), and miR-222-3p (AUC = 0.70; 95% CI, 0.58–0.81) were identified as significantly differentially abundant in plasma from stage II PDAC versus controls. Although none of the miRNAs individually outperformed the currently used serologic biomarker for PDAC, carbohydrate antigen 19-9 (CA19-9), combining the miRNAs with CA 19-9 improved AUCs from 0.89 (95% CI, 0.81–0.95) for CA 19-9 alone to 0.92 (95% CI, 0.86–0.97), 0.94 (95% CI, 0.89–0.98), and 0.92 (95% CI, 0.87–0.97), respectively. Gene set enrichment analyses of transcripts correlated with high and low expression of the three miRNAs in The Cancer Genome Atlas PDAC sample set. These miRNA biomarkers, assayed in limited volume plasma together with CA19-9, discriminate stage II PDAC from controls with good sensitivity and specificity. Unbiased profiling of larger cohorts should help develop an informative early detection biomarker assay for diagnostic settings. Prevention Relevance: Development of minimally invasive biomarker assays for detection of premalignant disease and early-stage pancreatic cancer is key to improving patient survival. This study describes a limited volume plasma miRNA biomarker assay that can detect early-stage resectable pancreatic cancer in clinical samples necessary for effective prevention and clinical intervention.

Sphingolipids constitute a class of bio‐reactive molecules that transmit signals and exhibit a variety of physical properties in various cell types, though their functions in cancer pathogenesis have yet to be elucidated. Analyses of gene expression profiles of clinical specimens and a panel of cell lines revealed that the ceramide synthase gene CERS6 was overexpressed in non–small‐cell lung cancer (NSCLC) tissues, while elevated expression was shown to be associated with poor prognosis and lymph node metastasis. NSCLC profile and in vitro luciferase analysis results suggested that CERS6 overexpression is promoted, at least in part, by reduced miR‐101 expression. Under a reduced CERS6 expression condition, the ceramide profile became altered, which was determined to be associated with decreased cell migration and invasion activities in vitro. Furthermore, CERS6 knockdown suppressed RAC1‐positive lamellipodia/ruffling formation and attenuated lung metastasis efficiency in mice, while forced expression of CERS6 resulted in an opposite phenotype in examined cell lines. Based on these findings, we consider that ceramide synthesis by CERS6 has important roles in lung cancer migration and metastasis.

Sphingolipids make up a family of molecules associated with an array of biological functions, including cell death and migration. Sphingolipids are often altered in cancer, though how these alterations lead to tumor formation and progression is largely unknown. Here, we analyzed non-small-cell lung cancer (NSCLC) specimens and cell lines and determined that ceramide synthase 6 (CERS6) is markedly overexpressed compared with controls. Elevated CERS6 expression was due in part to reduction of microRNA-101 (miR-101) and was associated with increased invasion and poor prognosis. CERS6 knockdown in NSCLC cells altered the ceramide profile, resulting in decreased cell migration and invasion in vitro, and decreased the frequency of RAC1-positive lamellipodia formation while CERS6 overexpression promoted it. In murine models, CERS6 knockdown in transplanted NSCLC cells attenuated lung metastasis. Furthermore, combined treatment with l-α-dimyristoylphosphatidylcholine liposome and the glucosylceramide synthase inhibitor D-PDMP induced cell death in association with ceramide accumulation and promoted cancer cell apoptosis and tumor regression in murine models. Together, these results indicate that CERS6-dependent ceramide synthesis and maintenance of ceramide in the cellular membrane are essential for lamellipodia formation and metastasis. Moreover, these results suggest that targeting this homeostasis has potential as a therapeutic strategy for CERS6-overexpressing NSCLC.

Thyroid transcription factor‐1 (TTF‐1), also known as NKX2‐1, plays a role as a lineage‐survival oncogene in lung adenocarcinoma that possesses double‐edged sword characteristics. Although evidence from previous studies has steadily accumulated regarding the roles of TTF‐1 in transcriptional regulation of protein‐coding genes, little is known about its regulatory relationship with microRNAs. Here, we utilized an integrative approach designed to extract maximal information from expression profiles of both patient tumors in vivo and TTF‐1‐inducible cell lines in vitro, which identified microRNA (miR)‐532‐5p as a novel transcriptional target of TTF‐1. We found that miR‐532‐5p is directly regulated by TTF‐1 through its binding to a genomic region located 8 kb upstream of miR‐532‐5p, which appears to impose transcriptional regulation independent of that of CLCN5, a protein‐coding gene harboring miR‐532‐5p in its intron 3. Furthermore, our results identified KRAS and MKL2 as novel direct targets of miR‐532‐5p. Introduction of miR‐532‐5p mimics markedly induced apoptosis in KRAS‐mutant as well as KRAS wild‐type lung adenocarcinoma cell lines. Interestingly, miR‐532‐5p showed effects on MEK‐ERK pathway signaling, specifically in cell lines sensitive to siKRAS treatment, whereas those miR‐532‐5p‐mediated effects were clearly rendered as phenocopies by repressing expression or inhibiting the function of MKL2 regardless of KRAS mutation status. In summary, our findings show that miR‐532‐5p is a novel transcriptional target of TTF‐1 that plays a tumor suppressive role by targeting KRAS and MKL2 in lung adenocarcinoma.

In preclinical models of glioblastoma, antigen escape variants can lead to tumor recurrence after treatment with CAR T cells that are redirected to single tumor antigens. Given the heterogeneous expression of antigens on glioblastomas, we hypothesized that a bispecific CAR molecule would mitigate antigen escape and improve the antitumor activity of T cells. Here, we created a CAR that joins a HER2-binding scFv and an IL13Rα2-binding IL-13 mutein to make a tandem CAR exodomain (TanCAR) and a CD28.ζ endodomain. We determined that patient TanCAR T cells showed distinct binding to HER2 or IL13Rα2 and had the capability to lyse autologous glioblastoma. TanCAR T cells exhibited activation dynamics that were comparable to those of single CAR T cells upon encounter of HER2 or IL13Rα2. We observed that TanCARs engaged HER2 and IL13Rα2 simultaneously by inducing HER2-IL13Rα2 heterodimers, which promoted superadditive T cell activation when both antigens were encountered concurrently. TanCAR T cell activity was more sustained but not more exhaustible than that of T cells that coexpressed a HER2 CAR and an IL13Rα2 CAR, T cells with a unispecific CAR, or a pooled product. In a murine glioblastoma model, TanCAR T cells mitigated antigen escape, displayed enhanced antitumor efficacy, and improved animal survival. Thus, TanCAR T cells show therapeutic potential to improve glioblastoma control by coengaging HER2 and IL13Rα2 in an augmented, bivalent immune synapse that enhances T cell functionality and reduces antigen escape.

Large consortia efforts and genome-wide association studies (GWASs) have linked a number of genetic variants within the 6p21 chromosomal region to non-Hodgkin lymphoma (NHL). Complementing these efforts, we genotyped previously reported SNPs in the human leukocyte antigen (HLA) class I (rs6457327) and class II (rs9271100, rs2647012 and rs10484561) regions in a total of 1,145 subjects (567 NHL cases and 578 healthy controls) from two major ethnic groups in Malaysia, the Malays and the Chinese. We identified a NHL-associated (PNHL_add = 0.0008; ORNHL_add = 0.54; 95% CI = 0.37–0.77) and B-cell associated (PBcell_add = 0.0007; ORBcell_add = 0.51; 95% CI = 0.35–0.76) SNP rs2647012 in the Malaysian Malays. In silico cis-eQTL analysis of rs2647012 suggests potential regulatory function of nearby HLA class II molecules. Minor allele rs2647012-T is linked to higher expression of HLA-DQB1, rendering a protective effect to NHL risk. Our findings suggest that the HLA class II region plays an important role in NHL etiology.

The standard of care in locally advanced rectal cancer is neoadjuvant chemoradiation (nCRT) followed by radical surgery. Response to nCRT varies among patients and pathological complete response is associated with better outcome. However, there is a lack of effective methods to select rectal cancer patients who would or would not have a benefit from nCRT. The utility of clinicopathological and radiological features are limited due to lack of adequate sensitivity and specificity. Molecular biomarkers have the potential to predict response to nCRT at an early time point, but none have currently reached the clinic. Integration of diverse types of biomarkers including clinicopathological and imaging features, identification of mechanistic link to tumor biology, and rigorous validation using samples which represent disease heterogeneity, will allow to develop a sensitive and cost-effective molecular biomarker panel for precision medicine in rectal cancer. Here, we aim to review the recent advance in tissue- and blood-based molecular biomarker research and illustrate their potential in predicting nCRT response in rectal cancer.

Background: The current standard treatment for patients with newly diagnosed diffuse large B cell lymphoma (DLBCL) is rituximab combined with cyclophosphamide, doxorubicin, vincristine and prednisolone (CHOP). A significant number of patients were not treated with recommended dose of rituximab due to limited financial resources in Malaysia. This study evaluates the efficacy of R-CHOP like chemotherapy in Malaysian patients with DLBCL. Materials and Methods: The study comprised a retrospective analysis of patients with DLBCL treated at a single centre. The outcome was compared with patients who were treated with R-CHOP like and CHOP like chemotherapy. Patients who were treated with lower dose of rituximab was subanalysed for outcome. Results: A total of 86 patients who had CHOP-like chemotherapy were included. Only 39 (45%) patients had rituximab and only 12 (29%) patients had the recommended dose. The overall response (OR) and complete response (CR) rates were 88% and 81% respectively. There was no significant difference in OR and CR in patients who had rituximab and those without rituxmab. Those with International Prognostic Index (IPI) score of ≤2 had significant higher CR rate, progression free survival (PFS) and overall survival (p

Accumulated evidence indicates that various types of miRNA are aberrantly expressed in lung cancer and secreted into the bloodstream. For this study, we constructed a serum diagnostic classifier based on detailed bioinformatics analysis of miRNA profiles from a training cohort of 143 lung adenocarcinoma patients and 49 healthy subjects, resulting in a 20 miRNA-based classifier. Validation performed with an independent cohort of samples from lung adenocarcinoma patients (n = 110), healthy subjects (n = 52) and benign pulmonary disease patients (n = 47) showed a sensitivity of 89.1% and specificity of 94.9%, with an area under the curve value of 0.958. Notably, 90.8% of Stage I lung adenocarcinoma cases were correctly diagnosed. Interestingly, this classifier also detected squamous and large cell lung carcinoma cases at relatively high rates (70.4% and 70.0%, respectively), which appears to be consistent with organ site-dependent miRNA expression in cancer tissues. In contrast, we observed significantly lower rates (0–35%) using samples from 96 cases of cancer in other major organs, with breast cancer the lowest. These findings warrant a future study to realize its clinical application as a part of diagnostic procedures for lung cancers, for which early detection and surgical removal is presently the only hope for eventual cure.

Accumulating evidence indicates that altered miRNA expression is crucially involved in lung cancer development, though scant information is available regarding how MYC, an archetypical oncogene, is regulated by miRNAs, especially via a mechanism involving MYC cofactors. In this study, we attempted to identify miRNAs involved in regulation of MYC transcriptional activity in lung cancer. To this end, we utilized an integrative approach with combinatorial usage of miRNA and mRNA expression profile datasets of patient tumor tissues, as well as those of MYC-inducible cell lines in vitro. In addition to miRNAs previously reported to be directly regulated by MYC, including let-7 and miR-17-92, our strategy also helped to identify miR-342-3p as capable of indirectly regulating MYC activity via direct repression of E2F1, a MYC-cooperating molecule. Furthermore, miR-342-3p module activity, which we defined as a gene set reflecting the experimentally substantiated influence of miR-342-3p on mRNA expression, was found to be inversely correlated with MYC activity reflected by MYC module activity in three independent datasets of lung adenocarcinoma patients obtained from the Director's Challenge Consortium of the United States (P = 1.94 × 10(-73)), the National Cancer Center of Japan (P = 9.05 × 10(-34)) and the present study (P = 1.17 × 10(-19)). Our integrative approach appears to be useful to elucidate inter-regulatory relationships between miRNAs and protein coding genes of interest, even those present in patient tumor tissues, which remains a challenge to better understand the pathogenesis of this devastating disease.

Our group and others have previously identified TTF-1/NKX2-1 as a lineage survival oncogene in lung adenocarcinoma. Subsequent studies revealed double-edged sword characteristics of TTF-1 in the development of lung adenocarcinoma. TTF-1 elicits lineage-survival signaling by inducing transcriptional targets such as ROR1, while TTF-1 also inhibits tumor progression by decreasing cell motility through direct activation of genes such as MYBPH. It is thus clear that a more comprehensive picture of this still enigmatic lineage-survival oncogene needs to be elucidated. Whereas most of the previous studies on TTF-1 have focused on its regulation of protein-coding genes, very little is known about regulation of microRNAs by TTF-1. In the present study, we combined in vivo patient data with in vitro data from lung adenocarcinoma cell lines, aiming at identifying miRNAs, which are transcriptionally regulated by TTF-1. Our integrative approach identified miR-532-5p as a microRNA regulated by TTF-1. Expression of miR-532-5p decreased, when cell lines with a high endogenous level of TTF-1 were transfected with siRNA against TTF-1, while it was increased by overexpression of exogenous TTF-1 in TTF-1-negative cell lines. By ChIP and luciferase assays, TTF-1 was found to induce miR-532-5p expression through its binding to the MIR532 promoter region, identifying miR-532-5p as a direct target of TTF-1. We next employed microarray analysis and target prediction using TARGETSCAN to search for potential targets of miR-532-5p. We consequently identified KRAS and MKL2 as candidates and confirmed this by qRT-PCR, Western blot and luciferase analyses. Overexpression of miR-532-5p induced apoptosis and inhibited colony formation in lung adenocarcinoma cell lines, while depletion of MKL2 phenocopied the effects of miR-532-5p introduction, similarly exhibiting occurrence of marked apoptosis. In addition, miR-532-5p significantly decreased tumor formations in vivo in a mouse xenograft model. It was also of note that miR-532-5p inhibited lung adenocarcinoma cell lines regardless of the presence or absence of KRAS mutations. In summary, we have found that TTF-1 transactivates miR-532-5p, which potently induces apoptosis in lung adenocarcinoma cells. MiR-532-5p directly targets MKL2, which we have also uncovered to be crucially involved in lung adenocarcinoma survival. In addition, while previous reports have shown that TTF-1 inhibits KRAS-driven tumorigenesis, this study identifies KRAS as a target of miR-532-5p, providing their potential mechanistic link. Our results thus shed light on how TTF-1 plays a tumor-suppressive role in the process of lung carcinogenesis, which might ultimately lead to development of a novel therapeutic strategy for this devastating disease. Citation Format: Sebastian Griesing, Taisuke Kajino, Mei Chee Tai, Zhuoran Liu, Masahiro Nakatochi, Motoshi Suzuki, Takashi Takahashi. TTF-1/NKX2-1 induced miR-532-5p targets KRAS and MKL2 oncogenes and causes apoptosis in lung adenocarcinoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2529. doi:10.1158/1538-7445.AM2017-2529

We evaluated the association of two IL10 single nucleotide polymorphisms (SNPs) (rs1800896 and rs1800871) with non-Hodgkin lymphoma (NHL) risk in the three major races of the Malaysian population (Malay, Chinese and Indian; 317 cases and 330 controls). Our initial screening demonstrated that rs1800871 but not rs1800896 was significantly associated with increased NHL risk in Malays (pMalay-Rec = 0.007) and Chinese only (pChinese-Rec = 0.039). Subsequent combined analysis of the Malay and Chinese revealed significant association of rs1800871 with all (ALL) NHL subtypes (pMeta-ALL-NHL-Rec = 0.001), ALL B-cell subtypes (pMeta-ALL-B-cell-Rec = 0.003), diffuse large B-cell lymphoma (DLBCL) subtype (pMeta-DLBCL-Rec = 0.002) and ALL T-cell subtypes (pMeta-ALL-T-cell-Rec = 0.031). SNP rs1800896 showed increased risk only in follicular lymphoma (FL) (pMeta-FL-Dom = 0.0004). We also detected a male-specific association of rs1800871 with increased NHL risk (pMeta-Male-ALL-NHL-Rec = 0.006) in the combined analysis. To our knowledge, this is the first report on the association of IL10 promoter SNPs with NHL susceptibility in the three major races of Malaysia.