Search

Samples from patients with colorectal cancer (CRC) are valuable tools for understanding the development, progression, and treatment of the disease. However, to date, the integrity of long-term preserved human specimens in biobanks has not been well understood. In this study, we investigated the RNA quality of 12-year-stored specimens, including frozen and formalin-fixed, paraffin-embedded (FFPE) tumor tissues from CRC patients at the Siriraj Biobank. We assessed the RNA quality of 12-year and 2-year storage samples using three technologies: next-generation sequencing (NGS), Nanostring nCounter® platform, and GeoMx® digital spatial profiling (DSP). We found that the RNA quality of 12-year storage frozen tissues was consistent with the criteria for RNA sequencing. Although RNA in long-term storage FFPE tissues was degraded, the normalized counts of RNA from the 12-year-stored FFPE samples were comparable to those from the 2-year-stored FFPE samples in the Nanostring nCounter® gene expression assay. For histological staining, clear tissue microanatomy was observed in the FFPE blocks stored for 12 years. In GeoMx® DSP, there was no statistically significant difference in the normalized counts of RNA from the 12- and 2-year stored FFPE samples. Our results suggest the potential utilization of long-term storage biobank specimens from patients with CRC for NGS, Nanostring nCounter® gene expression analysis, and GeoMx® DSP.

Objective: The aim of this study was to evaluate the efficacy and cost-effectiveness of a ready-to-use applicator containing iodine povacrylex and isopropyl alcohol (IPIA) for the prevention of surgical site infection (SSI) following colorectal surgery. Materials and Methods: The IPIA was randomly used in patients who underwent colorectal surgical procedures. The control group for comparison was a group of patients who underwent colorectal surgical procedures using conventional skin scrubbing and painting with antiseptic solutions without IPIA. In total, 100 patients were included in the study, randomized into 2 groups: IPIA was applied in study group and convention skin preparation was applied in control group.The outcome measurements included ease-of-use as assessed by a questionnaire, preparation time comparison, estimated skin preparation expense, adverse reactions, and rate of SSI. All the patients were visited daily up to 7 days postoperation or until discharge, and then 14 and 30 days postoperatively for monitoring the occurrence of SSI. Results: Of the 100 patients undergoing elective colorectal surgery enrolled in the study, 51 were males and 49 females, with the mean age of 63.5 ± 11.3 years. The majority of the patients had colorectal cancer undergone colectomies or rectal resections. There was no mortality. Seven patients (7%) had postoperative SSI (4 patients in the control group and 3 patients in the IPIA group, 8% vs. 6%, p = 0.45). The bacterial cultures revealed Gram negative-bacilli in all of the patients with SSI. The preparation time for the skin preparation was 5.48 ± 2.49 min in the control group and 2.65 ± 1.55 min in the IPIA group (p = 0.002), without statistical significance of expenses. Conclusion: IPIA was demonstrated to be as safe and effective as conventional antiseptic solutions as a skin preparation to prevent SSI following colorectal surgery. With good ease of use, IPIA proved more convenient than a scrubbing preparation as well as offered better cost effectiveness by significantly reducing the time and cost of the skin preparation

Introduction: Transmembrane proteins and their overexpression have been highly involved in malignancies. In particular, Tetraspanins (TSPANs)—a family of four-span transmembrane proteins—have been proposed as “master organizers” of multi-molecular membrane complexes by associating with specific partners and thus contributed to the progression of many cancer types. To date, transcriptomic analysis of all human TSPANs in colorectal cancer (CRC) has remained unclear. Here, we focused on utilizing next-generation RNA sequencing (RNA-Seq) to explore the expression level of TSPANs in primary tumor tissues of CRC, which is a complex disease and the second cause of cancer mortality worldwide. Experimental Procedures: Resected primary tumor tissues of CRC were collected from 100 patients along with the written informed consents. Later, total RNA was extracted from all frozen patient tissues. RNA-Seq of those RNA samples was next done before the transcriptomic analysis of TSPANs in CRC was performed. Immunofluorescence (IF) and Immunohistochemistry (IHC) stains were also employed to confirm the target protein expression in formalin-fixed paraffin-embedded tissue (FFPE) slides from CRC patients. Results: As a result of transcriptomic analysis of TSPANs in CRC tissues, among 33 human TSPANs, the high expression levels (transcript per million above the average of all TSPANs) of TSPAN1, TSPAN8, TSPAN24, TSPAN29 and TSPAN30 were observed in the patient-derived CRC tissues. Notably, the expression level of TSPAN8 was comparable with that of EPCAM, which is a well-known tumor marker. Moreover, the high expression of TSPAN8 protein could be clearly observed in both IF and IHC-stained FFPE slides from CRC patients. Our result highlights the possible relevance of these overexpressed TSPANs in CRC development. Conclusions: This study is the first RNA-Seq-based transcriptomic analysis of all human TSPANs in CRC. Our finding suggests the significant overexpression of particular TSPANs, especially TSPAN8, in CRC. Further analyses will be beneficial for translational research in CRC. The overexpression of TSPANs may suggest a framework to further explore diagnostic, prognostic, and predictive surface biomarkers, therapeutic targets, and other applications for CRC. Citation Format: Thanawat Suwatthanarak, Pariyada Tanjak, Amphun Chaiboonchoe, Onchira Acharayothin, Kullanist Thanormjit, Tharathorn Suwatthanarak, Watsaphon Tangkullayanone, Manop Pithukpakorn, Vitoon Chinswangwatanakul. Transcriptomic analysis of tetraspanins in colorectal cancer [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 2241.

Background: KRAS mutations (KRASmut) drive tumorigenesis through various signaling pathways that are associated with worse survival in colorectal cancer patients (CRC). Immune evasion is a hallmark of KRAS-driven cancer, but the role of KRASmut in the regulation of the immune response is unclear, particularly in the tumor microenvironment (TME), which often assists cancer cells to thrive and successfully escape immune surveillance. Here, we explore the immunobiological impacts of KRASmut in CRC patients. Methods: A total of 97 fresh frozen tumor samples were collected from each CRC patient with stage I-IV who underwent surgical treatment at the Department of Surgery, Faculty of Medicine Siriraj Hospital between October 2010 and March 2011. First, all patients were identified KRAS mutational status and compared overall survival (OS) with wild-type KRAS (KRASwt). Next, we analyzed the gene expression profiling of KRASmut which was performed using the Nanostring platform. Finally, we performed the biological network analysis focusing on immunological pathways associated with KRASmut using the Ingenuity Pathway Analysis (IPA). Results: We observed KRASmut in 41.24% (n=40) of the total cases. Mutations occurred primarily in exon 2 (total n=36; codon 12, n=28; codon 13, n=8) and rarely in exon 3 (n=4). Patients with KRASmut CRC in exon 2 and exon 3, showed significantly worse OS compared to KRASwt (HR 1.77, 95% CI 1.07-2.93; P Conclusions: Our results suggested that KRASmut activated TGFβ and interrupted pro-inflammatory cytokine to suppress tumor immune microenvironment in CRC. Future studies and clinical trials in patients with CRC with KRASmut should take these transcriptional profiles into account. Citation Format: Pariyada Tanjak, Amphun Chaiboonchoe, Tharathorn Suwatthanarak, Onchira Acharayothin, Kullanist Thanormjit, Jantappapa Chanthercrob, Thanawat Suwatthanarak, Bundit Wannasuphaphol, Kemmapon Chumchuen, Bhoom Suktitipat, Somponnat Sampattavanich, Krittiya Korphaisarn, Ananya Pongpaibul, Naravat Poungvarin, Harald Grove, Woramin Riansuwan, Atthaphorn Trakarnsanga, Asada Methasate, Manop Pithukpakorn, Vitoon Chinswangwatanakul. KRAS mutations suppress the tumor immune microenvironment in colorectal cancer [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 2242.

Colorectal cancers (CRC) with KRAS mutations (KRASmut) are frequently included in consensus molecular subtype 3 (CMS3) with profound metabolic deregulation. We explored the transcriptomic impact of KRASmut, focusing on the tumor microenvironment (TME) and pathways beyond metabolic deregulation. The status of KRASmut in patients with CRC was investigated and overall survival (OS) was compared with wild-type KRAS (KRASwt). Next, we identified CMS, and further investigated differentially expressed genes (DEG) of KRASmut and distinctive pathways. Lastly, we used spatially resolved gene expression profiling to define the effect of KRASmut in the TME regions of CMS3-classified CRC tissues. CRC patients with KRASmut were mainly enriched in CMS3. Their specific enrichments of immune gene signatures in immunosuppressive TME were associated with worse OS. Activation of TGFβ signaling by KRASmut was related to reduced pro-inflammatory and cytokine gene signatures, leading to suppression of immune infiltration. Digital spatial profiling in TME regions of KRASmut CMS3-classified tissues suggested up-regulated genes, CD40, CTLA4, ARG1, STAT3, IDO, and CD274, that could be characteristic of immune suppression in TME. This study may help to depict the complex transcriptomic profile of KRASmut in immunosuppressive TME. Future studies and clinical trials in CRC patients with KRASmut should consider these transcriptional landscapes.

Background Multiple primary cancer refers to the synchronous or metachronous appearances in the same individual. The accurate approach, which distinguishes between multiple primary cancer and metastasis, is important to make an appropriate management. This study aims to systematically classify the synchronous and metachronous multiple primary cancers, and to identify multiple primary cancer from cancer metastasis. Methods Cancer registry was compiled at the Faculty of Medicine Siriraj Hospital, Mahidol University based on guidelines from the Surveillance Epidemiology and End Results (SEER) project. Data was validated through clinical records, coded, and verified for multiple primary cancer according to International Classification of Diseases for Oncology 3rdEdition (ICD-O-3). The cancer registry for the year 1991-2015 was queried for numbers of the multiple primary cancer. Results A total of 1,913 patients presented multiple primary cancer, accounting for 1.48% of all patients (129,216). Median age at diagnosis of the first cancer was 60. The most multiple primary cancer in female is synchronous breast cancers, diagnosed two or more histologically distinct cancer within two months. In male, the most combination is prostate, urinary tract, and gastrointestinal cancer. In addition to understanding of the connections between primary and subsequent cancer, we plan to perform panel sequencing in each primary site of multiple gastrointestinal primary cancer to access for the causal driver mutation. Conclusion Cancer registry is a valuable approach to punctually identify and record multiple primary cancer. Further study in genetic profiles of each primary cancer may improve our understanding of multiple primary cancer for enhancing cancer management. Citation Format: Pariyada Tanjak, Benjarat Thiengtrong, Kanokporn Thamapala, Tippawan Gerdsuriwong, Cholticha Songjang, Aem-on Rattanakumnerd, Onchira Acharayothin, Supatee Karakate, Nongnard Thamnongdee, Suwanit Therasakvichya, Somsri Laiteerapong, Vitoon Chinswangwatanakul. Multiple primary cancer from a retrospective population-based cancer registry, 1991 through 2015 [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5767.

Cholangiocarcinoma (CCA) is a malignant tumor of the biliary epithelium associated with Opisthorchis viverrini, primary sclerosing cholangitis and hepatitis viral infection. In the global population, men have higher incidence rates for CCA than women; thus, a gender disparity in the progression of chronic inflammation of the biliary duct leading to malignancy may involve the effects of estrogen (E2). Genistein (GE), a prominent phytoestrogen found in soy products, is an estrogen receptor β (ERβ) agonist and a tyrosine kinase inhibitor. The present study investigated the effects of GE on the growth of CCA cells by cell viability assay. The effects on signaling proteins were detected by western blot analysis and ELISA. Gene expression was examined by RT-qPCR. Two human intrahepatic CCA cell lines, HuCCA‑1 and RMCCA‑1, were utilized. GE (50‑200 µM) reduced the viability of the two cell lines, and also inhibited the activation of epidermal growth factor receptor (EGFR) and AKT, as evidenced by decreasing protein levels of phosphorylated (p)-EGFR (Tyr1173) and p‑AKT (Ser473), respectively. GE altered the mitogen‑activated protein kinase signaling cascade by mediating decreased protein levels of p‑p38 and increased protein levels of p‑ERK1/2. GE significantly decreased the levels of interleukin 6 (IL6) and induced the expression of inducible nitric oxide synthase (iNOS). GE also downregulated the expression of p‑ERα (Ser118) protein and ERα mRNA levels. Finally, GE induced the downregulation of the protein levels of ERβ. Of note, E2 deprivation potentiated the GE-induced reduction of p‑EGFR (Tyr1173) and total AKT proteins and production of IL6, and mediated the downregulation of GE-induced iNOS protein. In conclusion, GE inhibited the growth of human CCA cell lines by reducing the activation of EGFR and AKT, and by attenuating the production of IL6. E2 and ER were also involved in the growth-inhibitory effect of GE in CCA cells.

Background Patient-derived organoids (PDOs) are versatile experimental models for cancer research. At the Faculty of medicine Siriraj Hospital, we have recently established live biobank of over seventy PDO models from stage I-IV Thai colorectal cancer patients. Our biobank included at least 10 patients who underwent neoadjuvant chemotherapy. We aimed to apply this resource for N-of-1 co-clinical trials, for the discovery of novel therapeutic agents and for biomarker development. We report here our application of the developed PDO models for predicting clinical outcome of colorectal cancer patients. Methods Surgically resected colorectal specimens from both normal and cancerous tissue were digested and embedded in appropriate matrices to form PDO models. Subsequently, all successful PDO models were collected in Siriraj live biobank together with their clinical data. Herein, we characterized our derived PDO models by pathological and molecular techniques. Some models were profiled with panel sequencing to assess for the causal driver mutations. Drug sensitivity of each PDO model to standard-of-care drugs were generated and compared with clinical outcome in each patient. Results We successfully established PDO models from different colorectal sites, of different pathological grades and staging. We identified heterogeneous morphologies of PDO models which appear to be corresponding well with the pathological and genomic profiles of each specimen origin. Although the PDO models from different colorectal cancer patients exhibited variable response to 5-fluorouracil (5-Fu), Leucovorin and Oxaliplatin, the response patterns are consistent with the clinical outcome of the individual patients as determined by tumor regression grade or radiologic imaging for neoadjuvant cases. We are examining the use of drug sensitivity profiling of PDO models for prioritizing anti-cancer treatment choices in a prospective clinical study. Conclusions The Siriraj live biobank of PDO from colorectal cancer patients can serve as a useful resource for N-of-1 co-clinical trials, and for comparison of drug response among patients from different ethnic backgrounds. Legal entity responsible for the study Faculty of Medicine Siriraj Hospital, Mahidol University, Thailand. Funding The National Research Council of Thailand and Siriraj foundation, Siriraj Hospital. Disclosure All authors have declared no conflicts of interest.

Since many studies reveal that the biological properties of various biopolymers depend on their molecular weight. Therefore, it was considered of importance to investigate how molecular weight affected on the interleukin-8 (IL-8) stimulating activity of the β-glucan from Ophiocordyceps dipterigena BCC2073. γ-Irradiation of the glucan with various doses (0–100 kGy) was chosen as a clean method to produce different low molecular weight samples. The result showed that average molecular weight (MW) of irradiated samples significantly decreased as the irradiation dose increased whereas the functional groups of before and after irradiated glucans detected by FTIR and 13C-NMR were identical. However, difference in intensity of an absorption peak at 270 nm was found in the UV/vis spectra. The biological properties such as cytotoxicity, proliferation and IL-8 secretion of normal human dermal fibroblast contacted with various MW glucans were also tested. It was found that the glucan with MW of approximately 5 kDa exhibited the highest ability to induce IL-8 production. Apart from the MW effect, chain conformation seems to be involved. Thus, differences in solution conformation of various MW glucans via Congo red analysis were evaluated.

Optimization of exopolysaccharides (EPS) produced by three strains of entomopathogenic fungi (Beauveria bassiana BCC 2692, Ophiocordyceps dipterigena BCC 2073, and Paecilomyces tenuipes BCC 2656) was carried out together with analyses of their prebiotic properties. B. bassiana BCC 2692 produced 6.27±0.22 g/L EPS on optimal medium using two-level fractional factorial design and 4.7 g/L EPS in bioreactor. EPS productions of O. dipterigena BCC 2073 were 13.2 g/L and 41.2 g/L in shake flask and bioreactor, respectively. For P. tenuipes BCC 2656, 1.47±0.21 g/L EPS in shake flask and 28.1 g/L EPS in bioreactor were obtained. These EPS were previously characterized as -glucan with differences in molecular weights and degree of branching. They were resistant to hydrolysis by both hydrochloric acid and porcine pancreatic α-amylase. Furthermore, when used as the sole carbon source, all three types of EPS supported growth in vitro of two different probiotic bacteria (Lactobacillus acidoplilus BCC 13839 and Bifidobacterium animalis ATCC 25527). A constant viability of L. acidophilus BCC 13839 was maintained throughout the cultivation period (48 hours) on all three entomopatogenic fungal EPS. All EPS also supported better growth and maintained longer growth period of B. animalis ATCC 25527 than glucose or inulin. Thus these entomopathogenic fungi EPS are promising candidates in prebiotic industry, expanding the pool of current commercial prebiotics.