Prognostic Marker For Overall Survival Research Articles

Abstract LB274: Nuclear transporter HIKESHI is a novel independent poor overall survival for pancreatic cancer; targeting can inhibit pancreatic cancer growth via HSP70 suppression

Abstract HIKESHI, under heat stress conditions, has been reported to transport molecular chaperone HSP70 into the nucleus to prevent cancer cell death from heat stress. The tumor-supporting roles of molecular chaperones in various solid tumors are explored extensively. Yet, the sole targeting of the molecular chaperones could not significantly impact anticancer treatment. Thus, we hypothesized that the specific transporter protein HIKESHI might improve this outcome. However, understanding of the clinical relevance of HIKESHI in pancreatic cancer remained unclear, and few studies addressed the HIKESHI role in cancer and whether targeting can inhibit pancreatic cancer cell viability and the other HSPs. Here, we first investigated the expression pattern of the HIKESHI protein in surgically resected pancreatic cancer samples (n=142) and analyzed the clinical relevance of the HIKESHI. We found that the HIKESHI protein was overexpressed in cancer compared to the adjacent normal tissue, and positive expression of cancer HIKESHI was related to poor overall survival. In addition, HIKESHI-positive expression in cancer was identified as an independent prognostic marker for overall survival. Next, using shRNA, we established the stable HIKESHI-suppressed pancreatic cancer cell line (PANC-1). HIKESHI suppression inhibited cell growth and colony formation ability in vitro. The expression and nuclear translocation of HSP70 and HSP110/105 were suppressed in HIKESHI-suppressed cells compared to the control shRNA cells despite no heat stress conditions. Furthermore, we found that the tumor growth, HIKESHI, and Ki67 expression in HIKESHI-suppressed tumors were inhibited compared to the control shRNA tumors. Next, we performed Cap Analysis of Gene Expression and IP-MS analysis to discover the potential downstream and novel interactive partner of the HIKESHI protein. The extensive mechanism of these analyses is under investigation. Our findings suggest that targeting HIKESHI may be a promising therapeutic strategy against refractory pancreatic cancer. Citation Format: Bilguun Erkhem-Ochir, Takehiko Yokobori, Gendensuren Dorjkhorloo, Haruka Okami, Takaomi Seki, Norifumi Harimoto, Ryo Muranushi, Kouki Hoshino, Kei Hagiwara, Norihiro Ishii, Mariko Tsukagoshi, Kenichiro Araki, Hiroshi Saeki, Ken Shirabe. Nuclear transporter HIKESHI is a novel independent poor overall survival for pancreatic cancer; targeting can inhibit pancreatic cancer growth via HSP70 suppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB274.

Abstract 5298: BSI-093, a best-in-class anti-BTN3A monoclonal antibody for cancer immunotherapy by activation of Vg9Vd2 T cells

Abstract Background: Gamma delta T (γδ T) cells are potent anti-cancer effectors with the potential to target tumors broadly, independent of patient-specific neoantigens or human leukocyte antigen background. Among tumor-infiltrating lymphocytes, the presence of γδ T cells is the most favorable prognostic marker for overall survival of patients with various solid and hematologic tumors. Vγ9Vδ2 T cells are the predominant γδ T cells in peripheral circulation. Butyrophilin 3A (BTN3A) members, widely expressed in tumors, have emerged as novel molecules modulating the function of Vγ9Vδ2 T cells. Targeting BTN3A for activation of Vγ9Vδ2 T cells provides an alternative strategy for cancer immunotherapy. Experimental procedures: BALB/c mice were immunized with recombinant BTN3A1-ECD-Fc. The Biosion proprietary H3 (High-throughput, High-content and High-efficiency) antibody discovery platform was used to identify an anti-BTN3A monoclonal antibody lead candidate - BSI-093. The target binding specificity, binding activity, and affinity of BSI-093 were evaluated by protein-based ELISA, cell-based FACS and Biacore-based SPR. Ex vivo assays were used to evaluate the cellular bioactivity of BSI-093 on activating Vγ9Vδ2 T cells for killing activity of various tumor cells. B-NDG mice bearing Jurkat-GFP-Luc tumors were used to evaluate the tumor inhibitory activity of the anti-BTN3A antibody. Summary: BSI-093 is a humanized monoclonal antibody with the following critical properties: (1) binds to all BTN3A members (BTN3A1, BTN3A2 and BTN3A3) with high affinity; (2) has potentially longer half-life and silenced Fc-effector function by Fc-engineering; (3) exhibits cross-reactivity to rhesus BTN3A members; (4) demonstrates strong activity on activating Vγ9Vδ2 T cells to kill a variety of natural cancer cells in ex vivo assays; and (5) shows significant anti-tumor efficacy in a CDX model. Conclusion: BSI-093 demonstrates potential best-in-class biophysical properties and functional characteristics, supporting the initiation of development activities including manufacturing and IND-enabling studies. Citation Format: Wenwen Dai, Jun Li, Jinyu Liu, Hongyan Li, Xiaodong F. Liu, Shukai Xia, Qun Lyu, Hugh M. Davis, Mingjiu Chen, Zeyu Peng. BSI-093, a best-in-class anti-BTN3A monoclonal antibody for cancer immunotherapy by activation of Vg9Vd2 T cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5298.

Abstract 5525: Tissue-resident cytotoxic T cell infiltration predicts metachronous colorectal cancer metastasis

Abstract Background: Identifying patients at high risk for metachronous colorectal cancer metastasis is essential for improving prognosis. The balance between host immune regulation and tumor immune evasion ultimately permits or prevents cancer progression. Lymphocytic infiltration in colorectal cancer can predict long-term clinical outcomes. However, these lymphocytes' origin, subtype and function have yet to be adequately explored. We investigated the association between the development of metastasis and CD8+T cells expressing the integrin protein CD103, a marker of tissue-resident memory T cells. Methods: In this retrospective, case-control matched study, we conducted multiplex immunofluorescence staining of tumor samples from 124 patients with colorectal cancer. Tissue microarrays containing representative cores of the central tumor, invasive margin, and adjacent normal tissue were immunostained for CD3, CD8, CD103, pSMAD3, and cytokeratin, using tyramide signal amplification. Cells were quantified using StrataQuest digital image analysis software, with intratumoral and stromal regions analyzed separately. Kruskal-Wallis one-way analysis of variance with subsequent Dunn’s test for pair-wise comparisons was performed using the Bonferroni method of multiple-comparison correction. Results: Central tumor and invasive margin CD3+ densities were associated with synchronous metastasis. Compared to non-metastatic patients, those who went on to develop metachronous metastasis (n=42) had a low proportion of intra-epithelial CD8+CD103+ T cells at the central tumor (non-metastatic: 32.8%, IQR 15.0-42.7%; metachronous metastasis: 12.9%, IQR 2.08-28.4%; P = 0.013). Similarly, patients with metachronous metastasis had a low proportion of stromal CD8+CD103+ T cells at the invasive margin (non-metastatic: 3.51%, IQR 1.83-6.55%; metachronous metastasis: 1.75%, IQR 0.62-3.72%; P = 0.019). On multivariate Cox regression of histological features, amongst patients with metastasis, significant overall survival prognostic markers included a low proportional CD8+CD103+ T cell infiltration (HR 2.41, 95%CI 1.16-5.01, P = 0.019), mucinous features (HR 4.28, 95%CI 1.98-9.24, P < 0.001), and the presence of perineural invasion (HR 2.75, 95%CI 1.28-5.89, P < 0.001). Conclusion: CD8+CD103+ lymphocytes may protect from the development of colorectal metastasis. Proportional CD8+CD103+ T cell infiltration is a promising prognostic marker in colorectal adenocarcinoma for the development of metachronous metastasis, and, amongst patients with metastasis, it is a prognostic marker of overall survival. Citation Format: Ryan Cohen, Tracey Lee-Pullen, Timothy Miller, Shadi Pirasteh, Cameron Platell, Katie Meehan, Kathy Fuller, Melanie McCoy. Tissue-resident cytotoxic T cell infiltration predicts metachronous colorectal cancer metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5525.

Three- and Seven-month Prostate-specific Antigen Levels as Prognostic Markers for Overall Survival in Metastatic Hormone-sensitive Prostate Cancer: Results from SWOG S1216, a Phase 3 Randomized Trial of Androgen Deprivation Plus Orteronel or Bicalutamide

BackgroundA robust decrease in prostate-specific antigen (PSA) in response to androgen deprivation therapy (ADT) has been evaluated as a prognostic factor in patients with metastatic hormone-sensitive prostate cancer (mHSPC) since 2006, but the treatment of mHSPC has since evolved to include intensified therapy. ObjectiveWe assessed the association of PSA levels at 3 (PSA-3mo) and 7 (PSA-7mo) mo with overall survival (OS) in patients with mHSPC treated with ADT combined with either bicalutamide or orteronel in the S1216 phase 3 clinical trial. Design, setting, and participantsPSA responses to treatment of patients in the S1216 trial were categorized as: complete response (CR) if PSA was ≤0.2 ng/ml, partial response if PSA was >0.2 and ≤4 ng/ml, and no response (NR) if PSA was >4 ng/ml. Outcome measurements and statistical analysisA Cox analysis (adjusted for treatment arm and three stratification factors: performance status, severity of disease, and early vs late induction) was used for OS association. While PSA-7mo association was a prespecified objective, PSA-3mo association was also evaluated. Results and limitationsA total of 1251 and 1231 patients from the S1216 study were evaluable for PSA-3mo and PSA-7mo, respectively. A PSA-7mo CR was associated with improved OS compared with NR (HR: 0.20; p < 0.0001). A PSA-3mo CR showed a similar association to NR (HR: 0.34; p < 0.0001). The association of a PSA response with survival did not differ by treatment arm at either time point. ConclusionsThe PSA-3mo and PSA-7mo responses were strongly associated with OS; taken with other emerging prognostic biomarkers, these markers may allow for early identification of patients at the highest risk of death, aid with counseling in clinical practice, and permit design of future clinical trials targeting these patients. Patient summaryA low prostate-specific antigen level at 3 or 7 mo after starting treatment for metastatic hormone-sensitive prostate cancer predicts longer survival regardless of the first treatment given with androgen deprivation therapy.

Development of statistical auto-segmentation method for diffusion restriction gray matter lesions in patients with newly diagnosed sporadic Creutzfeldt–Jakob disease

Quantification of diffusion restriction lesions in sporadic Creutzfeldt-Jakob disease (sCJD) may provide information of the disease burden. We aim to develop an automatic segmentation model for sCJD and to evaluate the volume of disease extent as a prognostic marker for overall survival. Fifty-six patients (mean age ± SD, 61.2 ± 9.9 years) were included from February 2000 to July 2020. A threshold-based segmentation was used to obtain abnormal signal intensity masks. Segmented volumes were compared with the visual grade. The Dice similarity coefficient was calculated to measure the similarity between the automatic vs. manual segmentation. Cox proportional hazards regression analysis was performed to evaluate the volume of disease extent as a prognostic marker. The automatic segmentation showed good correlation with the visual grading. The cortical lesion volumes significantly increased as the visual grade aggravated (extensive: 112.9 ± 73.2; moderate: 45.4 ± 30.4; minimal involvement: 29.6 ± 18.1 mm3) (P < 0.001). The deep gray matter lesion volumes were significantly higher for positive than for negative involvement of the deep gray matter (5.6 ± 4.6 mm3 vs. 1.0 ± 1.3 mm3, P < 0.001). The mean Dice similarity coefficients were 0.90 and 0.94 for cortical and deep gray matter lesions, respectively. However, the volume of disease extent was not associated with worse overall survival (cortical extent: P = 0.07; deep gray matter extent: P = 0.12).

Advanced lung cancer inflammation index predicts overall survival of hepatocellular carcinoma after hepatectomy.

Limited data are available regarding ALI's clinical relevance and prognostic value in patients with hepatocellular carcinoma (HCC) after hepatectomy. HCC patients who received hepatectomy at the Meizhou People's Hospital from May 2011 to February 2022 were enrolled in the study cohort. The ALI was calculated as follows: ALI = BMI (kg/m2) × ALB (g/dL)/(absolute neutrophil count/absolute lymphocyte count). The primary outcome was overall survival (OS). The secondary outcome was cancer-specific survival (CSS). Univariate and multivariate Cox regression analyses were performed, followed by nomogram construction and decision curve analysis (DCA). 425 HCC patients were enrolled for analyses. Lower preoperative ALI was significantly correlated with incomplete tumor capsule and advanced tumor stage. Lower preoperative ALI was an adverse independent prognostic factor for OS (HR: 1.512, 95% CI: 1.122-2.039, P 0.007) and CSS (HR: 1.754, 95% CI: 1.262-2.438, P <0.001) in HCC patients. The nomogram plot was built based on three (including age, TNM stage, and ALI) and two (including TNM stage and ALI) independent prognostic factors for OS and CSS, respectively. Further analyses indicated that the nomogram had better predictive value and some net benefit than the traditional TNM stage alone, especially in long-term OS. Our study further indicated that ALI could be a prognostic marker for OS and CSS in HCC patients after hepatectomy, especially in long-term OS.

Serum CEA as a Prognostic Marker for Overall Survival in Patients with Localized Pancreatic Adenocarcinoma and Non-Elevated CA19-9 Levels Treated with FOLFIRINOX as Initial Treatment: A TAPS Consortium Study.

About 25% of patients with localized pancreatic adenocarcinoma have non-elevated serum carbohydrate antigen (CA) 19-9 levels at baseline, hampering evaluation of response to preoperative treatment. Serum carcinoembryonic antigen (CEA) is a potential alternative. This retrospective cohort study from five referral centers included consecutive patients with localized pancreatic adenocarcinoma (2012-2019), treated with one or more cycles of (m)FOLFIRINOX, and non-elevated CA19-9 levels (i.e., < 37U/mL) at baseline. Cox regression analyses were performed to assess prognostic factors for overall survival (OS), including CEA level at baseline, restaging, and dynamics. Overall, 277 patients were included in this study. CEA at baseline was elevated (≥5ng/mL) in 53 patients (33%) and normalized following preoperative therapy in 14 patients (26%). In patients with elevated CEA at baseline, median OS in patients with CEA normalization following preoperative therapy was 33 months versus 19 months in patients without CEA normalization (p = 0.088). At time of baseline, only elevated CEA was independently associated with (worse) OS (hazard ratio [HR] 1.44, 95% confidence interval [CI] 1.04-1.98). At time of restaging, elevated CEA at baseline was still the only independent predictor for (worse) OS (HR 1.44, 95% CI 1.04-1.98), whereas elevated CEA at restaging (HR 1.16, 95% CI 0.77-1.77) was not. Serum CEA was elevated in one-third of patients with localized pancreatic adenocarcinoma having non-elevated CA19-9 at baseline. At both time of baseline and time of restaging, elevated serum CEA measured at baseline was the only predictor for (worse) OS. Therefore, serum CEA may be a useful tool for decision making at both initial staging and time of restaging in patients with non-elevated CA19-9.

ASO Visual Abstract: Serum CEA as Prognostic Marker for Overall Survival in Patients with Localized Pancreatic Adenocarcinoma and Nonelevated CA19-9 Levels Treated with FOLFIRINOX as Initial Treatment-A TAPS Consortium Study.

ASO Visual Abstract: Serum CEA as Prognostic Marker for Overall Survival in Patients with Localized Pancreatic Adenocarcinoma and Nonelevated CA19-9 Levels Treated with FOLFIRINOX as Initial Treatment-A TAPS Consortium Study.

Tumor infiltrating lymphocytes (TILs) are a prognosis biomarker in Colombian patients with triple negative breast cancer

Triple negative breast cancer (TNBC) is highly immunogenic and high levels of tumor infiltrating lymphocytes (TILs) have been associated with a better prognosis and higher probability to achieve pathological complete response. Here, we explore the potential role of stromal TILs level and composition as a prognostic and predictive biomarker in TNBC. 195 Tumor biospecimens from patients diagnosed with TNBC were included. Stromal TILs (sTILs), positive CD4/CD8 cells were evaluated. Differences in clinic-pathological characteristics according to immune infiltration were assessed. The predictive and prognostic value of immune infiltration was analyzed by multivariate models. Higher immune infiltration was observed in patients with favorable clinical–pathological features. Survival analysis showed that longer overall survival times were observed in patients with a higher infiltration of sTILs (p = 0.00043), CD4 + (p = 0.0074) and CD8 + (p = 0.008). In the multivariate analysis, low levels of sTILs were found to be associated with a higher mortality hazard (HR: 1.59, 95% CI 1.01–2.48). CD4 and CD8 immune infiltration were associated with higher odds for pathological complete response (OR: 1.20, 95% CI 1.00–1.46, OR: 1.28, 1.02–1.65, respectively). Our results suggest that immune infiltration could be used as a prognostic marker for overall survival in TNBC patients.

NLRP12 is a prognostic biomarker and correlated with immune infiltrates in epithelial ovarian cancer.

NLRP12 is a member of the intracellular Nod-like receptor (NLR) family, suggesting it is an innate immune receptor for the initiation and progression of several cancers. However, its role on prognosis and immune infiltrates in epithelial ovarian cancer (EOC) is still unknown. The present study aimed to evaluate its prognostic value and its association with immune infiltrates in EOC. The mRNA expression of NLRP12 of EOC from The Cancer Genome Atlas (TCGA) was analyzed. The association between NLRP12 and clinicopathological characters was evaluated with logistic regression. The association between NLRP12 expression and survival was analyzed by Cox regression and Kaplan-Meier analyses. A nomogram was used to predict the impact of NLRP12 on prognosis. Gene Ontology term analysis and gene set enrichment analysis (GSEA) were performed to identify the signaling pathways related to NLRP12 expression. Immune cells infiltration for NLRP12 was analyzed using single-sample GSEA. The relationship between NLRP12 and tumor-infiltrating immune cells (TICs) was investigated by a Wilcoxon rank sum test. The expression of NLRP12 were also further verified in EOC tissues and cell lines. Additionally, we confirmed the biological function of NLRP12 in vitro. NLRP12 was highly expressed in patients with EOC from TCGA. High NLRP12 expression correlated with poor disease-specific survival (p < 0.001) and overall survival (p < 0.001). Multivariate analysis revealed that NLRP12 expression was an independent prognostic marker for overall survival (p = 0.042). The C-indexes and calibration plots of the nomogram based on multivariate analysis indicated an effective predictive performance for EOC patients. GSEA showed enrichment of cell adhesion, tumorigenesis and immune response in the NLRP12 high expression group. Increased NLRP12 expression correlated positively with several TICs, including macrophages, neutrophils, T effector memory cells and immature dendritic cells (p < 0.001). In addition, NLRP12 silencing inhibited cell proliferation and migration in EOC cells. In conclusion, increased NLRP12 expression correlated significantly with poor survival and immune infiltration in EOC.

LRP1B—a prognostic marker in tubo-ovarian high-grade serous carcinoma

Low-density lipoprotein (LDL) receptor-related protein 1B (LRP1B) is a member of the LDLreceptor family and has often been discussed as a tumor suppressor gene, as its down-regulation is correlated with a poor prognosis in multiple carcinoma entities. Due to the high metastasis rate into the fatty peritoneal cavity and current research findings showing a dysregulation of lipid metabolism in tubo-ovarian high-grade serous carcinoma (HGSC), we questioned the prognostic impact of the LRP1B protein expression. We examined a well-characterized large cohort of 571 patients with primary HGSC and analyzed the LRP1B protein expression via immunohistochemical staining (both in tumor and stroma cells separately), performed precise bioimage analysis with QuPath, and calculated the prognostic impact using SPSS. Our results demonstrate that LRP1B functions as a significant prognostic marker for overall survival (OS) and progression-free survival (PFS) in HGSC on the protein level. High cytoplasmic expression of LRP1B in tumor, stroma, and combined tumor and stroma cells has a significantly positive association with a mean prolongation of the OS by 42 months (P=.005), 29 months (P=.005), and 25 months (P=.001), respectively. Additionally, the mean PFS was 18 months longer in tumor (P=.002), 19 months in stroma (P=.004), and 19 months in both cell types combined (P=.01). Our results remained significant in multivariate analysis. We envision LRP1B as a potential prognostic tool that could help us understand the functional role of lipid metabolism in advanced HGSC, especially regarding liposomal medications.

LRRC59 serves as a novel biomarker for predicting the progression and prognosis of bladder cancer.

Leucine-rich repeat-containing protein 59 (LRRC59) is an endoplasmic reticulum membrane protein involved in various cancers, but its role in bladder cancer (BC) has not been reported. The aim of the present study was to investigate the role of LRRC59 protein in BC progression and prognosis. The expression profile and clinical significance were retrieved from BC patients in the Cancer Genome Atlas database. The methylation status of LRRC59 was analyzed by UALCAN and MethSurv databases. Potential signaling pathways and biological functions were explored by functional enrichment analysis. Immunocyte infiltration was evaluated by CIBERSORT analysis. The prognostic value of LRRC59 was evaluated by Kaplan-Meier and Cox regression analyses. Overall survival (OS) was predicted by the nomogram plot established in this study. LRRC59 expression in 10 pairs BC and adjacent noncancerous tissues were analyzed by immunohistochemistry (IHC). Cell proliferation, migration, and invasion were detected by CCK8, colony formation assay, transwell assay, and cell scratch assay, respectively. Proteins related to epithelial-mesenchymal transition and apoptosis were detected by western blot. LRRC59 overexpression significantly decreased OS, disease-specific survival, and progress-free interval of BC patients. LRRC59 was a prognostic marker for OS and its hypomethylation status signified a poor prognosis. LRRC59 overexpression was correlated with infiltration of resting memory CD4 T cells, memory activated CD4 T cells, resting NK cells, macrophages M0, M1, M2, and neutrophils. IHC showed that the LRRC59 expression in BC tissue was significantly higher than that in adjacent noncancerous tissue. Knockdown of LRRC59 expression inhibited the proliferation of BC cells and reduced their migratory ability. Western blot showed that Snail and vimentin protein expressions decreased, while E-cadherin expressions increased. LRRC59 expression can predict the outcome of BC independently and serve as a new biomarker for diagnosis.

Increased expression of IDO1 is associated with improved survival and increased number of TILs in patients with high-grade serous ovarian cancer

BackgroundThe enzyme indoleamine 2,3-dioxygenase 1 (IDO1) plays a crucial role in regulating the immune system's response to tumors, but its exact role in cancer, especially in high-grade serous ovarian cancer (HGSOC), remains controversial. We aimed to investigate the prognostic impact of IDO1 expression and its correlation with tumor-infiltrating lymphocytes (TILs) in HGSOC. MethodsImmunohistochemical (IHC) staining and bioimage analysis using the QuPath software were employed to assess IDO1 protein expression in a well-characterized cohort of 507 patients with primary HGSOC. Statistical evaluation was performed using SPSS, and in silico validation considering IDO1 mRNA expression in bulk and single-cell gene expression datasets was conducted. Additionally, IDO1 expression in interferon-gamma (IFNG) stimulated HGSOC cell lines was analyzed. ResultsOur findings revealed that IDO1 protein and mRNA expression serve as positive prognostic markers for overall survival (OS) and progression-free survival (PFS) in HGSOC. High IDO1 expression was associated with a significant improvement in OS by 21 months (p < 0.001) and PFS by 6 months (p = 0.016). Notably, elevated IDO1 expression correlated with an increased number of CD3+ (p < 0.001), CD4+ (p < 0.001), and CD8+ TILs (p < 0.001). Furthermore, high IDO1 mRNA expression and protein level were found to be associated with enhanced responsiveness to pro-inflammatory cytokines, particularly IFNG. ConclusionsOur study provides evidence that IDO1 expression serves as a positive prognostic marker in HGSOC and is associated with an increased number of CD3+, CD4+ and CD8+ TILs. Understanding the intricate relationship between IDO1, TILs, and the tumor microenvironment may hold the key to improving outcomes in HGSOC.

Higher Mixed lineage Kinase Domain-like protein (MLKL) is associated with worst overall survival in adult-type diffuse glioma patients.

Recently, the search for novel molecular markers in adult-type diffuse gliomas has grown substantially, yet with few novel breakthroughs. As the presence of a necrotic center is a differential diagnosis for more aggressive entities, we hypothesized that genes involved in necroptosis may play a role in tumor progression. Given that MLKL is the executioner of the necroptotic pathway, we evaluated whether this gene would help to predict prognosis of adult gliomas patients. We analyzed a publicly available retrospective cohort (n = 530) with Kaplan Meier survival analysis (p<0.0001) and both uni- and multivariate Cox regression models. We determined that MLKL is an independent predictive prognostic marker for overall survival in these patients (HR: 2.56, p<0.001), even when controlled by the CNS5 gold-standard markers, namely IDH mutation and 1p/19q Codeletion (HR: 1.68, p = 0.013). These findings were confirmed in a validation cohort (n = 325), using the same cutoff value. Interestingly, higher expression of MLKL is associated with worse clinical outcome for adult-type diffuse glioma patients, which is opposite to what was found in other cell cancer types, suggesting that necroptosis undertakes an atypical detrimental role in glioma progression.

5ʹ-Ectonucleotidase CD73/NT5E supports EGFR-mediated invasion of HPV-negative head and neck carcinoma cells

BackgroundEpithelial-to-mesenchymal transition (EMT) of malignant cells is a driving force of disease progression in human papillomavirus-negative (HPV-negative) head and neck squamous cell carcinomas (HNSCC). Sustained hyper-activation of epidermal growth factor receptor (EGFR) induces an invasion-promoting subtype of EMT (EGFR-EMT) characterized by a gene signature (“‘EGFR-EMT_Signature’”) comprising 5´-ectonucleotidase CD73. Generally, CD73 promotes immune evasion via adenosine (ADO) formation and associates with EMT and metastases. However, CD73 regulation through EGFR signaling remains under-explored and targeting options are amiss.MethodsCD73 functions in EGFR-mediated tumor cell dissemination were addressed in 2D and 3D cellular models of migration and invasion. The novel antagonizing antibody 22E6 and therapeutic antibody Cetuximab served as inhibitors of CD73 and EGFR, respectively, in combinatorial treatment. Specificity for CD73 and its role as effector or regulator of EGFR-EMT were assessed upon CD73 knock-down and over-expression. CD73 correlation to tumor budding was studied in an in-house primary HNSCC cohort. Expression correlations, and prognostic and predictive values were analyzed using machine learning-based algorithms and Kaplan–Meier survival curves in single cell and bulk RNA sequencing datasets.ResultsCD73/NT5E is induced by the EGF/EGFR-EMT-axis and blocked by Cetuximab and MEK inhibitor. Inhibition of CD73 with the novel antagonizing antibody 22E6 specifically repressed EGFR-dependent migration and invasion of HNSCC cells in 2D. Cetuximab and 22E6 alone reduced local invasion in a 3D-model. Interestingly, combining inefficient low-dose concentrations of Cetuximab and 22E6 revealed highly potent in invasion inhibition, substantially reducing the functional IC50 of Cetuximab regarding local invasion. A role for CD73 as an effector of EGFR-EMT in local invasion was further supported by knock-down and over-expression experiments in vitro and by high expression in malignant cells budding from primary tumors. CD73 expression correlated with EGFR pathway activity, EMT, and partial EMT (p-EMT) in malignant single HNSCC cells and in large patient cohorts. Contrary to published data, CD73 was not a prognostic marker of overall survival (OS) in the TCGA-HNSCC cohort when patients were stratified for HPV-status. However, CD73 prognosticated OS of oral cavity carcinomas. Furthermore, CD73 expression levels correlated with response to Cetuximab in HPV-negative advanced, metastasized HNSCC patients.ConclusionsIn sum, CD73 is an effector of EGF/EGFR-mediated local invasion and a potential therapeutic target and candidate predictive marker for advanced HPV-negative HNSCC.

Baseline circulating tumor cell (CTC) count as a prognostic marker of overall survival (OS) in metastatic hormone sensitive prostate cancer (mHSPC): Results from SWOG S1216, a phase III randomized trial of androgen deprivation plus orteronel (cyp17 inhibitor) or bicalutamide.

5080 Background: Biomarkers are needed to predict clinical outcomes in mHSPC. We previously reported that baseline CTC counts in S1216 were prognostic of PSA response and disease progression ( ASCO 2020; CCR 2021). However, the trial’s primary overall survival (OS) endpoint was not yet mature at that time. Here we present the final results of S1216 using all available samples to assess the prognostic value of CTC counts for response, progression, OS, and treatment arm interactions. Methods: In S1216, peripheral blood was drawn with informed consent at registration (baseline) and at progression to metastatic castrate resistant prostate cancer (mCRPC). CTCs were enumerated on the FDA-cleared CellSearch platform (Menarini). CTC counts were evaluated for associations with 3 pre-specified trial endpoints: 7-month PSA (7mPSA) ≤ 0.2 ng/ml vs. 0.2–4.0 vs. &gt; 4.0 using a generalized logit model; and progression-free survival (PFS) and OS using Cox regression. Results: From 2014 to 2021, 503 evaluable samples were collected at baseline and 93 at progression. Baseline CTC count was ≥5 in 11.9% of samples, 1-4 in 19.3%, and 0 in 66.8%. Comparing men with 0 vs. ≥5 CTCs at baseline, those with 0 CTCs were significantly more likely to attain a complete PSA response and had significantly lower risk of a PFS event and death after adjusting for clinical covariates (disease burden, bone metastases, age, race, alkaline phosphatase, hemoglobin, PSA, treatment arm). These associations were not modified by treatment arm. Progression CTCs &lt;5 vs. ≥5 also were prognostic of OS (logrank p=0.001), consistent with prior findings in mCRPC. Same-patient CTC counts in matched baseline-progression pairs (n=61) were highly correlated (Spearman p&lt;0.001). Conclusions: CTC count at the start of treatment for mHSPC was highly prognostic of PSA response, PFS, and OS; associations that held in both treatment arms. Moreover, baseline CTC count strongly correlated with progression count, suggesting that mild vs. aggressive disease phenotypes may persist from mHSPC to mCRPC. This evidence from a large prospective phase 3 trial suggests that baseline CTC count is a valuable prognostic marker in men initiating therapy for mHSPC, discriminating those likely to experience favorable vs. unfavorable response and OS. [Table: see text]

The prognostic value of degree of pre- and post-treatment anemia in patients with advanced gastroesophageal cancers.

e16012 Background: Anemia is common in cancer patients and cancer-related anemia has been associated with poorer clinical outcomes in various malignancies. The degree of anemia in patients receiving chemotherapy, as a prognostic factor in gastroesophageal cancers (GE), is not well understood. There have been studies looking at clinical prognostic scores and their efficacy of predicting outcomes, of which the Gustave Roussy Immune (GRIm-Score) has been seen to be most predictive of early death in GE cancers. This study aims to compare hemoglobin (g/L) values before treatment and at multiple longitudinal timepoints during treatment as a prognostic marker of overall survival (OS) and progression-free survival (PFS). It also aims to identify whether various laboratory measures in the GRIm-Score, which includes lactate dehydrogenase (LDH), neutrophil to lymphocyte ratio (NLR) and body mass index (BMI) may hold prognostic value. Methods: A retrospective analysis of 171 patients with advanced GE cancer receiving first-line palliative-intent systemic therapy at the Princess Margaret Cancer Centre in Toronto, Canada from 2011 to 2021 was performed. Laboratory data were longitudinally collected across four time points: pre-chemotherapy, at first restaging scan, at disease progression and at final blood draw at last known follow up. Overall survival (OS) and progression-free survival (PFS) were estimated using the Kaplan-Meier method. Cox proportional hazards regression models were used to assess the association between change in hemoglobin from baseline, adjusted for LDH, NLR and BMI. Results: Mean hemoglobin value decreased with chemotherapy initiation at the first staging time point compared to pre-chemotherapy values by 10.82 g/L (-10.82, CI: -13.57, -8.07, p &lt; 0.001). On univariate analysis, patients with a hemoglobin increase of 10 g/L at their final blood draw compared to pre-chemotherapy decreased their overall risk of death by 8.50% (HR 0.915, CI: 0.843-0.993, p = 0.033) as well as their PFS by 6.90% (HR 0.931, CI: 0.873-0.994, p = 0.031). On multivariable analysis, adjusting for relevant clinical factors including LDH, NLR and BMI, an increase in hemoglobin increase of 10 g/L from baseline to final was associated with better OS (HR 0.893, CI: 0.817-0.977, p = 0.01). On multivariate analysis, an increase in LDH at progression bloodwork compared to baseline levels was associated with poorer OS (HR 1.002, CI: 1.001-1.003, p = 0.026) and an increase in NLR at final blood draw compared to baseline was associated with poorer OS (HR 1.02, CI: 1.012-1.028, p &lt; 0.001). Conclusions: Patients with metastatic GE cancers can develop treatment-associated anemia and this prognosticates a poorer overall survival. Prognostic models incorporating change in hemoglobin, LDH and NLR ratios should be considered in future clinical evaluation of metastatic GE cancers.

Impact of Smoking, Body Weight, Diabetes, Hypertension and Kidney Dysfunction on Survival in Pancreatic Cancer Patients-A Single Center Analysis of 2323 Patients within the Last Decade.

In addition to being risk factors for pancreatic cancer, parameters such as smoking, diabetes, or obesity might also act as potential prognostic factors for the survival of patients initially diagnosed with pancreatic cancer. By implementing one of the largest retrospective study cohorts of 2323 pancreatic adenocarcinoma (PDAC) patients treated at a single high-volume center, potential prognostic factors for survival were evaluated on the basis of 863 cases. Since parameters such as smoking, obesity, diabetes, and hypertension can cause severe chronic kidney dysfunction, the glomerular filtration rate was also considered. In the univariate analyses, albumin (p < 0.001), active smoking (p = 0.024), BMI (p = 0.018), and GFR (p = 0.002) were identified as metabolic prognostic markers for overall survival. In multivariate analyses, albumin (p < 0.001) and chronic kidney disease stage 2 (GFR < 90 mL/min/1.37 m2; p = 0.042) were identified as independent metabolic prognostic markers for survival. Smoking presented a nearly statistically significant independent prognostic factor for survival with a p-value of 0.052. In summary, low BMI, status of active smoking, and reduced kidney function at the time of diagnosis were associated with lower overall survival. No prognostic association could be observed for presence of diabetes or hypertension.

CRISPR/Cas9-based genome-wide screening of the deubiquitinase subfamily identifies USP3 as a protein stabilizer of REST blocking neuronal differentiation and promotes neuroblastoma tumorigenesis

BackgroundThe repressor element-1 silencing transcription factor (REST), a master transcriptional repressor, is essential for maintenance, self-renewal, and differentiation in neuroblastoma. An elevated expression of REST is associated with impaired neuronal differentiation, which results in aggressive neuroblastoma formation. E3 ligases are known to regulate REST protein abundance through the 26 S proteasomal degradation pathway in neuroblastoma. However, deubiquitinating enzymes (DUBs), which counteract the function of E3 ligase-mediated REST protein degradation and their impact on neuroblastoma tumorigenesis have remained unexplored.MethodsWe employed a CRISPR/Cas9 system to perform a genome-wide knockout of ubiquitin-specific proteases (USPs) and used western blot analysis to screen for DUBs that regulate REST protein abundance. The interaction between USP3 and REST was confirmed by immunoprecipitation and Duolink in situ proximity assays. The deubiquitinating effect of USP3 on REST protein degradation, half-life, and neuronal differentiation was validated by immunoprecipitation, in vitro deubiquitination, protein-turnover, and immunostaining assays. The correlation between USP3 and REST expression was assessed using patient neuroblastoma datasets. The USP3 gene knockout in neuroblastoma cells was performed using CRISPR/Cas9, and the clinical relevance of USP3 regulating REST-mediated neuroblastoma tumorigenesis was confirmed by in vitro and in vivo oncogenic experiments.ResultsWe identified a deubiquitinase USP3 that interacts with, stabilizes, and increases the half-life of REST protein by counteracting its ubiquitination in neuroblastoma. An in silico analysis showed a correlation between USP3 and REST in multiple neuroblastoma cell lines and identified USP3 as a prognostic marker for overall survival in neuroblastoma patients. Silencing of USP3 led to a decreased self-renewal capacity and promoted retinoic acid-induced differentiation in neuroblastoma. A loss of USP3 led to attenuation of REST-mediated neuroblastoma tumorigenesis in a mouse xenograft model.ConclusionThe findings of this study indicate that USP3 is a critical factor that blocks neuronal differentiation, which can lead to neuroblastoma. We envision that targeting USP3 in neuroblastoma tumors might provide an effective therapeutic differentiation strategy for improved survival rates of neuroblastoma patients.

Prognostic significance of dysregulation of shelterin complex and its correlation with telomere length and cytogenetics in multiple myeloma

BackgroundMM (multiple myeloma) is a bone marrow disease with the accumulation of malignant plasma cells characterized by the neoplastic transformation of differentiated B cells.The onset and progression of cancer are greatly influenced by telomere dysfunction. We aimed to study the biomarker potential and prognostic significance of shelterin complex and hTERT. Telomere length and gene expression were measured using real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR), and these results were further correlated with clinical parameters. ResultsOur study showed increased expression of all genes in complex, hTERT, and TL in MM (n = 72) in comparison with controls (n = 31). TRF2 (P = 0.025) and hTERT (P = 0.0002) displayed significant association among cytogenetic analysis. The receiver operative curve showed POT1 and RAP1 with a greater area under the curve (AUC). RAP1 (P = 0.020) and hTERT (P = 0.037) displayed to be independent prognostic markers for overall survival. Clinical parameters and genes were observed to be significantly correlated. ConclusionOur study findings showed variation in telomere-associated genes and suggest the participation of these genes as prognostic markers in MM. These results all together highlight the evaluation and role of genes involved in telomeric alteration and TL, providing the opportunity to study new therapeutic approaches in patients with MM.