Folates in the treatment of colorectal cancer

ABSTRACT Folates in the Treatment of Colorectal Cancer Department of Surgery Institute of Clinical Sciences at Sahlgrenska Academy University of Gothenburg Helena Taflin Background: Colorectal cancer is one of the most common cancers in the world, and radical surgery with total removal of the tumour (RO-resection) is the single most important treatment. However, chemotherapy is recommended for patients with risk factors and patients with metastatic disease. 5-fluoruracil (5-FU) is the cornerstone of chemotherapy, used either as a single drug or in combination with other drugs. 5-FU it is almost always combined with the folate leucovorin (LV). The aim of this thesis was to examine the role of polymorphisms in genes involved in folate metabolism in relation to treatment and to examine the levels of various folate forms in the tumours, mucosa, and plasma of patients who received LV or Modufolin® which is the biological isomer of 5,10-methylenetetrahydrofolate. Methods: Polymorphisms in the methylenetetrahydrofolate reductase (MTHFR), methionine synthase (MTR), and thymidylate synthase (TYMS) genes were analysed using real-time PCR and TaqMan chemistry. The various folate forms were analysed in tumours, mucosa, and plasma using a sensitive liquid chromatography electrospray ionization tandem mass spectrometry technique. Results: There was interdependency between polymorphisms in the MTFHR and MTR genes, which was associated with risk of side effects and overall survival in patients with stage III colorectal cancer receiving adjuvant chemotherapy. Total folate levels, all well as tetrahydrofolate (THF) and 5,10-methyleneTHF levels were significantly higher in tumours than in mucosa tissue. The individual variation in folate levels in both tumours and mucosa was greater than the variation found when the patients were subgrouped by gene polymorphisms. Only half of the patients who received 60 mg/m2 LV had higher levels of 5,10-methyleneTHF in tumours than patients who received 0 mg/m2 LV. Patients with rectal cancer had significantly lower levels of 5,10-methyleneTHF compared with patients with colon cancer. 5,10-methyleneTHF and THF concentrations were significantly higher in mucosa (p<0.003, both dosages) and tumours (p<0.015) 200 mg/m2) after Modufolin® administration than after LV (Isovorin®) administration. Conclusions: Polymorphisms in folate-associated genes can affect the risk that patients with colorectal cancer suffer from side effects during treatment with 5-FU-based chemotherapy. There is wide interindividual variation in 5,10-methyleneTHF levels in tumour tissue and mucosa after administration of standardised doses of LV. The doses of LV used in Nordic FLV-treatment may result in suboptimal levels of 5,10-methyleneTHF, especially in patients with rectal cancer. Modufolin® administration resulted in significantly higher 5,10-methyleneTHF levels than the natural l-form of LV, Isovorin®, and may potentially increase the efficacy of 5-FU-based chemotherapy. Keywords: Folates, colorectal cancer, leucovorin, Modufolin®, polymorphisms, MTHFR; Methionine Synthase, TS, side effects, adjuvant chemotherapy LS-MS/M

Rapport från projektet Multimodala uppgifter och bedömning i matematik : Video och kollaborativa samtal som metod vid lärares bedömning av elevers matematiska kunskaper

This research will discuss some experiences from a four year school research study. It was conducted in cooperation with teachers from four municipalities in Dalarna. The aim of the research was to examine teachers´ professional development when they participated in collaborative discussions based on video recordings and video edited material from specific lessons in their own practice. The study had two foci one was to investigate methods and tools that teachers can use to develop their ability to assess their students while working on multimodal tasks. The other was to examine how video can be used by teachers wanting to obtain knowledge about assessing students. The study is based on several theories about when teachers collaborate to create new knowledge. The first is the design theoretical approach – where visual ethnography and a semiotic approach contribute to problematize the use and mixture of different modes. A basic assumption of the framework here is that meanings are made and communicated in mathematics through a wide range of semiotic modes. By using video as an essential tool in the research the framework theories concerning visual ethnography, video documentation and individuals as reflective practitioners were also needed. The findings can be divided into the following themes: the use of tasks for assessment, collaborative discussion, equipment, ethical dilemmas. Collaborative discussions were evaluated as a meaningful way of sharing knowledge. The use of video recordings in association with these discussions raised important ethical issues. Working with the assessment framework was of great interest to the teachers but it took a lot of time from their ordinary work. In this way the project highlighted more general aspects of school development. The research also concerns teachers´ use of collaborative discussions in assessment work, multimodal tasks in mathematics and video as a research tool in general.Finansierat av Skolforskningsfonden</p

Folate Levels and Polymorphisms in the Genes , and in Colorectal Cancer

Aim The aim of the study was to explore and describe the effect of polymorphisms in folate-associated genes regarding the levels of different folate forms and their distribution in tumors and mucosa in patients with colorectal cancer. Materials and Methods Tumor and mucosa tissues from 53 patients with colorectal cancer were analyzed. The concentrations of tetrahydrofolate (THF), 5-methylTHF, and 5,10-methyleneTHF were measured by liquid chromatography—mass spectrometry. Genotyping of polymorphisms in the folate-associated genes methylenetetrahydrofolate reductase ( MTHFR, C677T ), methionine synthase ( MTR, A2756G ), and thymidylate synthase ( TS , 5'-TSER 28 bp tandem repeat and 3'-TSUTR 6 bp deletion/insertion), were done by real-time polymerase chain reaction. Folate levels and distributions were determined in the total patient cohort and after subgrouping by genotypes. Results The total folate level, as well as the THF and 5,10-methyleneTHF levels, were significantly higher in the tumor compared with mucosa tissue ( P = 0.030, 0.031, and 0.015, respectively). The individual variation in folate levels in both tumor and mucosa were larger than the variation found when the patients were subgrouped by the gene polymorphisms. No significant differences in the mean concentration of any folate in the mucosa or tumor tissue were found in relation to the analyzed polymorphisms. The percentage level of 5,10-methyleneTHF in tumors was highest in patients with the MTHFR 677 CC genotype, and lowest in patients with the TT genotype ( P = 0.033). A significantly lower percentage level of the 5,10-methyleneTHF level was found in tumors of patients with the 5'-TSER 3R/3R genotype ( P = 0.0031). Conclusion A significant difference was found between the percentage level of 5,10-methyleneTHF in tumor tissues in relation to the MTHFR C677T and 5'-TSER 28 bp repeat polymorphisms. However, no differences were found in the actual tissue folate levels, or in their distribution, in relation to the polymorphisms in the MTHFR, MTR , or TS genes. These findings could be of importance for further research in the field by explaining some of the difficulties of obtaining reproducible and uniform results when using a few selected polymorphisms as predictive markers

Surgical treatment of breast cancer liver metastases : A nationwide registry-based case control study

Introduction: The benefit of liver resection or ablation for breast cancer liver metastases (BCLM) remains unclear. The aim of the study was to determine survival after isolated BCLM in nationwide cohorts and compare surgical versus systemic treatment regimens. Materials and methods: The Swedish register for cancer in the liver and the bile ducts (SweLiv) and the National register for breast cancer (NBCR) was studied to identify patients with 1-5 BCLM without extrahepatic spread diagnosed 2009-2016. Data from the registers were validated and completed by review of medical records. A Kaplan-Meier plot and log rank test were used to analyse survival. Prognostic and predictive factors were evaluated by Cox regression analysis. Results: A surgical cohort (n = 29) was identified and compared to a control cohort (n = 33) receiving systemic treatment only. There was no 90-day mortality after surgery. Median survival from BCLM diagnosis was 77 months (95% CI 41-113) in the surgical cohort and 28 months (95% CI 13-43) in the control cohort, (p = 0.004). There was a longer disease-free interval and more oestrogen receptor positive tumours in the surgical cohort. Surgery was a significant positive predictive factor in univariate analysis while a multivariable analysis resulted in HR 0.478 (CI 0.193-1.181, p = 0.110) for surgical treatment. Conclusion: Surgery for BCLM is safe and might provide a survival benefit in selected patients but prospective trials are warranted to avoid selection bias

Increasing Dosage of Leucovorin Results in Pharmacokinetic and Gene Expression Differences When Administered as Two-Hour Infusion or Bolus Injection to Patients with Colon Cancer

The combination of 5-fluorouracil (5-FU) and leucovorin (LV) forms the chemotherapy backbone for patients with colorectal cancer. However, the LV administration is often standardized and not based on robust scientific data. To address these issues, a randomized pharmacokinetics study was performed in patients with colon cancer. Thirty patients were enrolled, receiving 60, 200 or 500 mg/m2 LV as a single two-hour infusion. Blood, tumor, mucosa, and resection margin biopsies were collected. Folate concentrations were analyzed with LC-MS/MS and gene expression with qPCR. Data from a previous study where patients received LV as bolus injections were used as comparison. Saturation of methylenetetrahydrofolate (MeTHF) and tetrahydrofolate (THF) levels was seen after two-hour infusion and polyglutamated MeTHF + THF levels in tumors decreased with increasing LV dosage. The decrease was associated with decreased FPGS and increased GGH expression, which was not observed after LV bolus injection. In the bolus group, results indicate activation of a metabolic switch possibly promoting TYMS inhibition in response to 5-FU. Different metabolic mechanisms appear to be induced when LV is administered as infusion and bolus injection. Since maximal inhibition of TYMS by the 5-FU metabolite 5-fluoro-2&prime;-deoxyuridine 5&prime;-monophosphate (FdUMP) requires excess polyglutamated MeTHF, the results point in favor of the bolus regimen

Increasing Dosage of Leucovorin Results in Pharmacokinetic and Gene Expression Differences When Administered as Two-Hour Infusion or Bolus Injection to Patients with Colon Cancer

The combination of 5-fluorouracil (5-FU) and leucovorin (LV) forms the chemotherapy backbone for patients with colorectal cancer. However, the LV administration is often standardized and not based on robust scientific data. To address these issues, a randomized pharmacokinetics study was performed in patients with colon cancer. Thirty patients were enrolled, receiving 60, 200 or 500 mg/m2 LV as a single two-hour infusion. Blood, tumor, mucosa, and resection margin biopsies were collected. Folate concentrations were analyzed with LC-MS/MS and gene expression with qPCR. Data from a previous study where patients received LV as bolus injections were used as comparison. Saturation of methylenetetrahydrofolate (MeTHF) and tetrahydrofolate (THF) levels was seen after two-hour infusion and polyglutamated MeTHF + THF levels in tumors decreased with increasing LV dosage. The decrease was associated with decreased FPGS and increased GGH expression, which was not observed after LV bolus injection. In the bolus group, results indicate activation of a metabolic switch possibly promoting TYMS inhibition in response to 5-FU. Different metabolic mechanisms appear to be induced when LV is administered as infusion and bolus injection. Since maximal inhibition of TYMS by the 5-FU metabolite 5-fluoro-2′-deoxyuridine 5′-monophosphate (FdUMP) requires excess polyglutamated MeTHF, the results point in favor of the bolus regimen

Quality-of-life after bile duct injury repaired by hepaticojejunostomy : a national cohort study.

OBJECTIVES: Reports on quality-of-life (QoL) after bile duct injury (BDI) show conflicting results. The aim of this cohort study was to evaluate QoL stratified according to type of treatment. METHODS: QoL assessment using the SF-36 (36-item short form health survey) questionnaire. Patients with post-cholecystectomy BDI needing hepaticojejunostomy (HJ) were compared to all other treatments (BDI repair) and to patients without BDI at cholecystectomy (controls). RESULTS: Patients needing a HJ after BDI reported reduced long-term QoL irrespective of time for diagnosis and repair in both the physical (PCS; p &lt; .001) and mental (MCS; p &lt; .001) domain compared to both controls and patients with less severe BDI. QoL was comparable for BDI repair (n = 86) and controls (n = 192) in both PCS (p = .171) and MCS (p = .654). As a group, patients with BDI (n = 155) reported worse QoL than controls, in both the PCS (p &lt; .001) and MCS (p = .012). Patients with a BDI detected intraoperatively (n = 124) reported better QoL than patients with a postoperative diagnosis. Patients with an immediate intraoperative repair (n = 99), including HJ, reported a better long-term QoL compared to patients subjected to a later procedure (n = 54). CONCLUSIONS: Patients with postoperative diagnosis and patients with BDIs needing biliary reconstruction with HJ both reported reduced long-term QoL

Proteomic profiling of tumour tissue‐derived extracellular vesicles in colon cancer

Abstract Colon cancer is one of the most commonly occurring tumours among both women and men, and over the past decades the incidence has been on the rise. As such, the need for biomarker identification as well as an understanding of the underlying disease mechanism has never been greater. Extracellular vesicles are integral mediators of cell‐to‐cell communication and offer a unique opportunity to study the machinery that drives disease progression, and they also function as vectors for potential biomarkers. Tumour tissue and healthy mucosal tissue from the colons of ten patients were used to isolate tissue‐resident EVs that were subsequently subjected to global quantitative proteomic analysis through LC‐MS/MS. In total, more than 2000 proteins were identified, with most of the common EV markers being among them. Bioinformatics revealed a clear underrepresentation of proteins involved in energy production and cellular adhesion in tumour EVs, while proteins involved in protein biosynthesis were overrepresented. Additionally, 53 membrane proteins were found to be significantly upregulated in tumour EVs. Among them were several proteins with enzymatic functions that degrade the extracellular matrix, and three of these, Fibroblast activating factor (FAP), Cell surface hyaluronidase (CEMIP2), as well as Ephrin receptor B3 (EPHB3), were validated and found to be consistent with the global quantitative results. These stark differences in the proteomes between healthy and cancerous tissue emphasise the importance of the interstitial vesicle secretome as a major player of disease development

A Fraction of CD8+ T Cells from Colorectal Liver Metastases Preferentially Repopulate Autologous Patient-Derived Xenograft Tumors as Tissue-Resident Memory T Cells

The diversity of T cells in the human liver may reflect the composition of TILs in CRLM. Our ex vivo characterization of CRLM vs. adjacent liver tissue detected CD103+CD39+CD8+ TRM cells predominantly in CRLM, which prompted further assessments. These TRM cells responded to cognate antigens in vitro. As functional activities of autologous TILs are central to the implementation of personalized cancer treatments, we applied a patient-derived xenograft (PDX) model to monitor TILs’ capacity to control CRLM-derived tumors in vivo. We established PDX mice with CRLMs from two patients, and in vitro expansion of their respective TILs resulted in opposing CD4+ vs. CD8+ TIL ratios. These CRLMs also displayed mutated KRAS, which enabled trametinib-mediated inhibition of MEK. Regardless of the TIL subset ratio, persistent or transient control of CRLM-derived tumors of limited size by the transferred TILs was observed only after trametinib treatment. Of note, a portion of transferred TILs was observed as CD103+CD8+ TRM cells that strictly accumulated within the autologous CRLM-derived tumor rather than in the spleen or blood. Thus, the predominance of CD103+CD39+CD8+ TRM cells in CRLM relative to the adjacent liver and the propensity of CD103+CD8+ TRM cells to repopulate the autologous tumor may identify these TILs as strategic targets for therapies against advanced CRC