Background Liquid biopsies have become an integral part of cancer management as minimally invasive options to detect molecular and genetic changes. However, current options show poor sensitivity in peritoneal carcinomatosis (PC). Novel exosome-based liquid biopsies may provide critical information on these challenging tumors. In this initial feasibility analysis, we identified an exosome gene signature of 445 genes (ExoSig445) from colon cancer patients, including those with PC, that is distinct from healthy controls. Methods Plasma exosomes from 42 patients with metastatic and non-metastatic colon cancer and 10 healthy controls were isolated and verified. RNAseq analysis of exosomal RNA was performed and differentially expressed genes (DEGs) were identified by the DESeq2 algorithm. The ability of RNA transcripts to discriminate control and cancer cases was assessed by principal component analysis (PCA) and Bayesian compound covariate predictor classification. An exosomal gene signature was compared with tumor expression profiles of The Cancer Genome Atlas. Results Unsupervised PCA using exosomal genes with greatest expression variance showed stark separation between controls and patient samples. Using separate training and test sets, gene classifiers were constructed capable of discriminating control and patient samples with 100% accuracy. Using a stringent statistical threshold, 445 DEGs fully delineated control from cancer samples. Furthermore, 58 of these exosomal DEGs were found to be overexpressed in colon tumors. Conclusions Plasma exosomal RNAs can robustly discriminate colon cancer patients, including patients with PC, from healthy controls. ExoSig445 can potentially be developed as a highly sensitive liquid biopsy test in colon cancer.

Purpose: Peritoneal carcinomatosis (PC), metastasized from colorectal cancer (CRC), remains a highly lethal disease. Outcomes of PC is significantly influenced by the amount of intra-abdominal tumor burden and therefore diagnostic tests that facilitate earlier diagnosis could improve PC treatment and patient outcomes.Experimental Design: Using mass-spectrometry-based proteomics, we characterized the protein features of circulating exosomes in the context of CRC PC, CRC with liver metastasis, and primary CRC limited to the colon. We profiled exosomes isolated from patient plasma to identify exosome-associated protein cargoes released by these cancer types.Results: Analysis of the resulting data identified metastasis-specific exosome protein signatures. Bioinformatic analyses confirmed enrichment of proteins annotated to vesicle-associated processes and intracellular compartments, as well as representation of cancer hallmark functions and processes. Conclusion and ClinicalRelevance: This research yielded distinct protein profiles for the CRC patient groups and suggests the utility of plasma exosome proteomic analysis for a better understanding of PC development and metastasis.

Recent military conflicts in Iraq and Afghanistan have resulted in the significant increase in blast-related traumatic brain injury (TBI), leading to increased Department of Defense interest in its potential long-term effects ranging from the mildest head injuries termed subconcussive trauma to the most debilitating termed chronic traumatic encephalopathy (CTE). Most patients with mild TBI will recover quickly while others report persistent symptoms called postconcussive syndrome. Repeated concussive and subconcussive head injuries result in neurodegenerative conditions that may hinder the injured for years. Fundamental questions about the nature of these injuries and recovery remain unanswered. Clinically, patients with CTE present with either affective changes or cognitive impairment. Genetically, there have been no clear risk factors identified. The discovery that microglia of the cerebral cortex discharged small extracellular vesicles in the injured and adjacent regions to a TBI may soon shed light on the immediate impact injury mechanisms. The combination of neuroimaging and advanced research techniques may, one day, fill critical knowledge gaps and lead to significant TBI research and treatment advancements.

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Tumor recurrence and metastasis contribute to the poor prognosis of ovarian cancer (OC) and glioblastoma multiforme (GBM). The aggressiveness of GBM and OC during recurrence creates a need to identify potential therapeutic targets. The cancer stem cell (CSC)-like phenotype and epithelial mesenchymal transition (EMT) have been shown to contribute to cancer aggressiveness, which makes CSCs and EMT viable targets for therapeutic intervention. EMT is a normal process during development and wound healing, but its reactivation in cancer cells contributes to metastasis. To detect CSC phenotypes in live cells without the use of antibodies, we are analyzing core stem cell transcription factors in which a SOX2/OCT4 response element, designated as SORE6, regulates a GFP reporter system. Also, to detect EMT we are using a GFP based reporter regulated by Zeb1 3’ UTR, a common EMT marker. Our aim is to detect changes in stemness and EMT in response to proton and photon ionizing radiation (IR). We hypothesize that proton and photon IR have similar effects on aggressiveness in OC and GBM. To investigate this, 2 GBM cancer cell lines, 1 breast cancer cell line, 1 OC cell line, and two patient derived samples were exposed to 0, 1, 2, 4 and 8Gy of 250 MeV proton and 6 MeV photon beams. 72 hours post IR exposure, flow cytometry analysis quantified reporter activity. We found an increase in stemness as radiation dosage increased in most cell lines. Results suggest that proton radiation increases stemness more than photon exposure. Citation Format: Aaron Keniston, Alondra Enciso, Yeonkyu Jung, Ashley Antonissen, Ryan Fuller, Ann Morcos, Antonella Bertucci, Marcello Vazquez, Juli Unternaehrer. Detection of stemness and EMT in response to radiation in glioblastoma and ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6396.

Background Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease with poor prognosis, as it is difficult to predict or circumvent, and it develops chemoresistance quickly. One cellular mechanism associated with chemoresistance is alternative splicing dysfunction, a process through which nascent mRNA is spliced into different isoforms. Survivin (Baculoviral IAP Repeat-Containing Protein 5 (BIRC5)), a member of the inhibitor of apoptosis (IAP) protein family and a cell cycle-associated oncoprotein, is overexpressed in most cancers and undergoes alternative splicing (AS) to generate six different splicing isoforms. Methods To determine if survivin splice variants (SSV) could be involved in PDAC chemoresistance, a Gemcitabine (Gem) resistant (GR) cell line, MIA PaCa-2 GR, was created and assessed for its SSV levels and their potential association with GR. Cross-resistance was assessed in MIA-PaCa-2 GR cells to FIRINOX (5-fluorouracil (5-FU), irinotecan, and oxaliplatin). Once chemoresistance was confirmed, RT-qPCR was used to assess the expression of survivin splice variants (SSVs) in PDAC cell lines. To confirm the effect of SSVs on chemoresistance, we used siRNA to knockdown all SSVs or SSV 2β. Results The MIA PaCa-2 GR cell line was 40 times more resistant to Gem and revealed increased resistance to FIRINOX (5-fluorouracil (5-FU), irinotecan, and oxaliplatin); when compared to the parental MIA-PaCa-2 cells. RT-qPCR studies revealed an 8-fold relative expression increase in SSV 2β and a 2- to 8-fold increase in the other five SSVs in the GR cells. Knockdown of all SSV or SSV 2β only, using small inhibitory RNA (siRNA), sensitized the GR cells to Gem, indicating that these SSVs play a role in PDAC chemoresistance. Conclusion These findings provide evidence for the potential role of SSV 2β and other SSVs in innate and acquired PDAC chemoresistance. We also show that the expression of SSVs is not affected by the type of chemoresistance, therefore targeting survivin splice variants in combination with chemotherapy could benefit a wide range of patients.