Untreated Colorectal Cancer Patients Research Articles

GTF2H5 Identified as a crucial synthetic lethal target to counteract chemoresistance in colorectal cancer

BackgroundSynthetic lethality (SL) emerges as a novel concept being explored to combat cancer progression and resistance to conventional therapy. Despite the efficacy of chemotherapy in select cases of colorectal cancer (CRC), a substantial proportion of patients encounter challenges, leading to an adverse prognosis of CRC patients. CRC-related SL genes offer a potential avenue for identifying therapeutic targets. MethodsCRC-related SL genes were obtained from the SynLethDB database. The bulk RNA sequencing data, mutation data, and clinical information for treated and untreated CRC patients were enrolled from the UCSC and GEO databases. The Tumor Immunology Single Cell Center database served as the repository for collecting and analyzing single-cell RNA sequencing data. The synergistic killing effect of SL genes and chemotherapeutic drugs on resistant cells was experimentally verified. ResultsIn the present study, pivotal SL genes associated with chemoresistance identified by using WGCNA and CRC patients categorized into two groups based on these genes. Variations between the groups were most pronounced in pathways associated with extracellular matrix remodeling. Further by integrating mutation data, five potential SL genes were discerned, which were highly expressed in the presence of TP53 or KRAS mutations, leading to a severely poor prognosis. Subsequent time series analysis revealed that the expression of GTF2H5 was gradually elevated at different stages of the transition from sensitive to resistant in CRC cells. Finally, it was preliminarily verified by experiments that GTF2H5 may play a key role in driving the drug-resistant transition within CRC cells. ConclusionsThe identification of SL genes that collaboratively interact with chemotherapeutic agents could provide new insights into solving the issue of chemotherapy resistance in CRC patients. And GTF2H5 wields a fundamental influence in inducing chemoresistance in CRC, which provided a potential therapeutic target for CRC.

Hypomethylation of Sonic hedgehog in colorectal cancer

Background: Past research has demonstrated that a changed Sonic Hedgehog (SHH) pattern contributes to the development of colorectal cancer. This study’s objective was to determine whether the SHH gene’s promoter hypomethylation might be used to predict the likelihood of developing colorectal cancer.
 Methods: For the current investigation, 50 newly diagnosed and untreated colorectal cancer patients’ tumor samples and surrounding non-tumor tissues were gathered. SHH methylation was determined by methylation-specific PCR (MSP) and results correlated with several studied clinicopathological parameters.
 Results: Our results showed that SHH methylation levels are significantly present in normal tissues as compared to tumor tissues.
 Conclusion: We conclude SHH promoter is hypomethylated in colorectal cancers which may have an effect driving normal colon and rectum cells to cancer.
 Bangladesh Journal of Medical Science Vol. 23 No. 01 January’24 Page : 200-206

P-398 Early chemo- and immuno-related toxicities in previously untreated colorectal cancer patients according to age: A retrospective single-centre study

P-398 Early chemo- and immuno-related toxicities in previously untreated colorectal cancer patients according to age: A retrospective single-centre study

Selective vulnerability of ARID1A deficient colon cancer cells to combined radiation and ATR-inhibitor therapy.

ARID1A is frequently mutated in colorectal cancer (CRC) cells. Loss of ARID1A function compromises DNA damage repair and increases the reliance of tumor cells on ATR-dependent DNA repair pathways. Here, we investigated the effect of ionizing radiation (IR), in combination with ATR inhibitors (ATRi) in CRC cell lines with proficient and deficient ARID1A. The concept of selective vulnerability of ARID1A deficient CRC cells to ATRi was further tested in an ex vivo system by using the ATP-tumor chemosensitivity assay (ATP-TCA) in cells from untreated CRC patients, with and without ARID1A expression. We found selective sensitization upon ATRi treatment as well as after combined treatment with IR (P<0.001), especially in ARID1A deficient CRC cells (P <0.01). Knock-down of ARID1B further increased the selective radiosensitivity effect of ATRi in ARID1A negative cells (P<0.01). Mechanistically, ATRi abrogates the G2 checkpoint (P<0.01) and homologous recombination repair (P<0.01) in ARID1A deficient cells. Most importantly, ex-vivo experiments showed that ATRi had the highest radiosensitizing effect in ARID1A negative cells from CRC patients. Collectively, our results generate pre-clinical and clinical mechanistic rationale for assessing ARID1A defects as a biomarker for ATR inhibitor response as a single agent, or in a synthetic lethal approach in combination with IR.

Abstract B43: Genetic analysis of circulating tumor cells of colorectal cancer patients captured by multiantibodies technique

Abstract Background: Accurate genetic analysis is essential for molecular therapy. Conventionally, DNA derived from tumor tissue has been the source of the gold standard of DNA information in solid tumors including colorectal cancer (CRC). However, it is well known that molecular therapy induces emerging mutations that are not found in primary tumor; thus, real-time monitoring of gene information is ideal. Circulating tumor cells (CTCs) are circulating malignant cells of solid tumor origin that are found in the bloodstream and can be a powerful candidate to play an important role. Capturing a large amount of CTCs improves the accuracy of genetic analysis of DNA derived from CTC (ctcDNA). We used 3 (EpCAM, Her2, Trop2) or 4 (EpCAM, Her2, Trop2, EGFR) antibodies to capture CTCs, and analyzed ctcDNA using next-generation sequencing (NGS). Methods: Cohort 1: Untreated CRC patients were enrolled. Ten mL of whole blood was collected from each patient. The blood was processed using 3 antibodies (EpCAM, Her2, Trop2) and CTCs were collected. Tumor tissue was also collected from each patient. Tumor tissue DNA and ctcDNA were extracted and analyzed using NGS. Cohort 2: CRC patients, both treated and untreated, were enrolled. Twenty mL of whole blood was collected from each patient. Ten mL of the blood was processed using 3 antibodies (EpCAM, Her2, Trop2), and the remaining 10 mL was processed using 4 antibodies (EpCAM, Her2, Trop2, EGFR). The numbers of collected CTCs were counted and compared. Results: Cohort 1: We enrolled 34 patients (stage II: n=4, stage III: n=7, stage IV: n=23). Median number of extracted CTC was 34 cells. From tumor tissue DNA, 53 mutations were detected. The most frequent mutation was within TP53 (n=18), followed by mutations in APC (n=13) and KRAS (n=12). From ctcDNA, 16 mutations, including 5 mutations which were not found in tissue DNA, were detected. The most frequent mutation was within TP53 (n=5), followed by mutations in KRAS and APC (n=4 each). Cohort 2: We enrolled 10 patients (stage II: n=1, stage III: n=1, stage IV: n=8). Using 3 antibodies, the median number of collected CTCs was 27 cells (range, 2–112). Using 4 antibodies, the median number of collected CTCs was 33 cells (range, 7–260). There were no statistically significant differences between the 2 groups (p=0.40). Conclusions: Mutations not detected in primary tumors can be identified in ctcDNA, indicating the potential of CTCs in complementing gene analysis. The technique to capture CTCs using 3 antibodies appears to increase the detection rate and yield of CTCs. However, the present study did not show advantages of the 4-antibodies method, and future studies should investigate the best combination of antibodies to extract more CTCs with higher specificity. Citation Format: Kohki Takeda, Takeshi Yamada, Michihiro Koizumi, Seiichi Shinji, Akihisa Matsuda, Ryo Ohta, Yasuyuki Yokoyama, Goro Takahashi, Masahiro Hotta, Takuma Iwai, Keisuke Hara, Koji Ueda, Sho Kuriyama, Hiroshi Yoshida. Genetic analysis of circulating tumor cells of colorectal cancer patients captured by multiantibodies technique [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B43.

Neural mechanisms of impaired executive function in untreated colorectal cancer patients.

e24170 Background: Cancer-related cognitive impairment (CRCI) affects 45% of colorectal cancer (CRC) patients prior to treatment. This study aimed to explore the neural mechanisms of CRCI in this severely understudied population by using a combination of neuropsychological and cognitive neurophysiology assessments. Methods: Newly diagnosed Stage I-IV CRC patients were recruited at a Midwestern tertiary medical center. Enrollment criteria included normal or corrected to normal vision and no prior cancer within 3 years. Healthy comparisons were recruited and demographically-matched to Stage III/IV patients scheduled to receive chemotherapy. Participants completed neuropsychological measures of processing speed, psychomotor speed, executive function, and verbal memory at study baseline. We recorded scalp electroencephalography while participants performed a computer-based cognitive task designed to measure control over spatial attention. Two event-related potentials were measured following presentation of a task-irrelevant spatial cue: (1) N2, a neural correlate of stimulus selection emerging approximately 160 milliseconds after stimulus presentation; and (2) N2pc, a neural correlate of spatial attention allocation emerging approximately 200 milliseconds after stimulus presentation. Results: CRC patients were enrolled within 4-8 weeks following surgery for Stage I (11%), II (31%), and III (35%) CRC; Stage IV patients (23%) started chemotherapy without surgery. Patients (n = 26) and controls (n = 14) were matched on age and gender; patients were trending toward lower education than controls (p = .06). Patients performed worse than controls on executive function assessments (Trials B: p = .049; Stroop: p = .025). N2 amplitudes were smaller in patients than controls (p = 0.032. N2pc amplitudes were larger in patients than controls (p = .092). N2 amplitudes were associated with Stroop interference scores (r = 0.50, p = .068). Conclusions: CRC patients demonstrated impaired executive function performance at baseline. Neurophysiological data suggest impairments may emerge from limited neural resources available for stimulus selection and poor control over spatial attention. Correlations between neurophysiological data and executive function performance suggest participants better able to overcome task-irrelevant interference had relatively more neural resources available for stimulus selection. Overall, these results suggest patients may be limited in their ability to focus their attention on caregiver and health care team instructions. Clinical trial information: NCT03683004 .

Abstract 4594: Detection of colorectal cancer related gene mutations from CTC and ctDNA

Abstract [Background] It is well known that emerging mutations which is not found in the primary tumor exist in metastatic tumor and molecular therapy induce emerging mutation. Liquid biopsy, which includes circulating tumor cell (CTC) and circulating tumor DNA (ctDNA), may help detecting this spatial and temporal heterogeneity. We have reported that emerging KRAS mutation can be detected by using ctDNA (Yamada et al, Cancer Science 2016). However, mutation detection by using CTC has been difficult because enough amount of DNA cannot be extracted from CTC. Currently we have been able to collect more CTC than before, by using a new device which uses 3 antibodies (EpCAM, Her2, Trop2). In this study, we evaluated the potential to detect colorectal cancer (CRC) related gene mutations from CTC, and compared it with ctDNA. [Methods] Cohort 1: This cohort included untreated CRC patients. Tumor tissue was collected from each patient, either by primary surgery or by colonoscopic biopsy. DNA was extracted from tumor tissue and was analyzed using Next Generation Sequencing (NGS). Ten mL of whole blood was also collected from the same patient. CTC, serum and white blood cell (WBC) was collected by using the CTC recovery machine (Ion Torrent Liquid Biopsy Instrument®). Cytokeratin positive, DAPI positive, CD45 negative cells were defined as CTC. DNA was extracted from each sample (CTC-DNA, ctDNA, WBC-DNA) and was analyzed using NGS. Cohort 2: This cohort included unresectable CRC patients with KRAS mutation in their primary tumor. All patients in this cohort were under treatment or after completion of chemotherapy. CTC and ctDNA was collected in the same method as cohort 1. KRAS mutations of CTC and ctDNA were detected by using digital PCR (dPCR). [Results] Cohort 1: We enrolled 16 CRC patients (stage II: n=2, stage III: n=2, stage IV: n=12). A total of 30 somatic, hotspot mutations were detected from tumor tissue DNA. The median number of the detected mutation for each patient was 2 (0-4). The most frequent gene mutation was APC, followed by KRAS and TP53. In all patients, CTC was successfully collected. The median number of the CTC was 34 cells (5-94). However, only 6 somatic, mutations were detected from CTC-DNA. Conversely, 16 somatic mutations were detected from ctDNA. Cohort 2: We enrolled 14 stage IV CRC patients with KRAS mutation in their primary tumor. CTC was collected from 9 patients but not from 5 patients. In the 9 patients CTC was collected, the median number of the collected CTC was 26 cells (5-121). By dPCR, KRAS mutation was detected in 2 patients (2/14) from CTC-DNA, and 4 patients (4/14) from ctDNA. [Conclusions] The new CTC capturing technology using 3 antibodies can improve detection rate and yield of CTC. However, in patients undergoing chemotherapy, the amount of CTC and ctDNA drastically reduces. At present, ctDNA is superior to CTC in potential to detect mutations, and dPCR is more sensitive than NGS to detect mutations. Citation Format: Kohki Takeda, Takeshi Yamada, Michihiro Koizumi, Seiichi Shinji, Yasuyuki Yokoyama, Goro Takahashi, Masahiro Hotta, Takuma Iwai, Keisuke Hara, Hiroyasu Furuki, Eiji Uchida. Detection of colorectal cancer related gene mutations from CTC and ctDNA [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4594.

Measurement of DNA damage in peripheral blood by the γ-H2AX assay as predictor of colorectal cancer risk

The detection of γ-H2AX focus is one of the most sensitive ways to monitor DNA double-strand breaks (DSBs). Although changes in γ-H2AX activity have been studied in tumor cells in colorectal cancer (CRC), changes in peripheral blood lymphocytes (PBLs) have not been examined previously. We hypothesize that higher levels of irradiation-induced γ-H2AX in PBLs may be associated with an elevated risk of colorectal cancer (CRC). In a case-control study, the baseline and ionizing radiation (IR)-induced γ-H2AX levels in PBLs from frequency-matched 320 untreated CRC patients and 320 controls were detected by a laser scanning cytometer-based immunocytochemical method. We used unconditional multivariable logistic regression to evaluate CRC risk by using the ratio of IR-induced γ-H2AX to the baseline levels with adjustment of age, sex and smoking status. We found CRC cases had significantly higher γ-H2AX ratio (1.5 vs. 1.41, P<0.0001) compared with controls. When using the median γ-H2AX ratio of controls as a cutoff point, we found higher γ-H2AX ratio was significantly associated with an increased risk of CRC (OR=6.72, 95% CI=4.54–9.94). Quartile analyses also showed significant dose–response relationship between higher γ-H2AX ratio and increased risk of CRC (P for trend<0.0001). Age, sex, BMI and smoking status also influenced the association of γ-H2AX ratio with CRC risk; however, no interactions with γ-H2AX ratio were observed. These results support the premise that DSBs in peripheral blood as measured by γ-H2AX level might represent an intermediate phenotype to assess the risk of CRC. Future prospective studies are necessary to confirm our findings in independent populations.

Influence of thymidylate synthase expression on survival in patients with colorectal cancer

Background: Thymidylate synthase (TS) plays a critical role in nucleotide metabolism and is an important target for 5-fluorouracil (5-FU), the standard chemotherapeutic drug for treatment of colorectal cancer (CRC). Aims and Methods: The present study aimed to evaluate TS variable number tandem repeat sequences (VNTR) polymorphism by polymerase chain reaction and TS protein expression by immunohistochemistry and its association with clinicopathological parameters in untreated CRC patients (n = 100). Further, the prognostic and predictive role of TS has been evaluated. Results: For TS VNTR polymorphism, the observed frequencies of 2R/2R, 2R/3R, and 3R/3R genotypes were 22%, 51%, and 27%, respectively. When immunohistochemical localization was studied, cytoplasmic staining for TS was observed in 70% of patients. A significant inverse correlation was noted between TS protein expression and tumor, node, metastasis staging (P = 0.027), Dukes' staging (P = 0.039), and lymph node status (P = 0.012) of CRC patients. However, there was no significant correlation between TS VNTR polymorphism and TS protein expression. On survival analysis, a significantly shorter overall survival (OS) was seen in CRC patients with negative protein expression (P = 0.031). Moreover, the subgroup of CRC patients treated only with surgery also showed a trend of poor OS in patients with negative TS protein expression (P = 0.058). However, neither TS polymorphism nor its protein expression was able to predict relapse-free survival. Conclusion: Negative TS protein expression may be related to unfavorable clinical outcome in CRC patients. However, further studies in a larger set of patients are necessary to better assess TS as a prognostic and predictive marker for 5-FU response in CRC patients.

Feedback mechanisms between M2 macrophages and Th17 cells in colorectal cancer patients.

IL-17 and IL-22 are linked to the development of intestinal inflammation and colorectal cancer (CRC). However, the maintenance of IL-17 and IL-22 production, as well as the cell type (Th17) that mediates these cytokines in CRC patients, remains unknown. To examine this, untreated CRC patients and healthy controls were recruited in this study. We first observed that CRC patients contained significantly elevated levels of IL-17- and IL-22-producing CD4+ T cells. The vast majority of IL-22-expressing CD4+ T cells also expressed IL-17. We then found that the production of both IL-17 and IL-22 required support from autologous monocytes, since the depletion of monocytes significantly downregulated IL-17 and IL-22 secretion. Naive T cells from CRC patients did not secrete IL-17 or IL-22 initially, but long-term coculture with autologous monocytes significantly upregulated IL-17 and IL-22 production in an IL-6-dependent manner. Blockade of IL-6 significantly reduced the levels of both IL-17 and IL-22. We then observed that CD163+ M2 macrophages were the main contributor of IL-6. Interestingly, incubation of monocytes with CCR4+CCR6+ Th17 cells resulted in significantly higher levels of CD163+ macrophages as well as higher IL-6 secretion, than incubation with non-Th17 CD4+ T cells. Together, our study discovered a positive feedback mechanism between Th17 and M2 macrophages in CRC patients.

Nicotinamide Mononucleotide Adenylyl Transferase 2: A Promising Diagnostic and Therapeutic Target for Colorectal Cancer.

Colorectal cancer (CRC) is one of the most common cancers all over the world. It is essential to search for more effective diagnostic and therapeutic methods for CRC. Abnormal nicotinamide adenine dinucleotide (NAD) metabolism has been considered as a characteristic of cancer cells. In this study, nicotinamide mononucleotide adenylyl transferases (NMNATs) as well as p53-mediated cancer signaling pathways were investigated in patients with colorectal cancer. The CRC tissues and adjacent normal tissues were obtained from 95 untreated colorectal cancer patients and were stained for expression of nicotinamide mononucleotide adenylyl transferase 2 (NMNAT2) and p53. The survival rate was analyzed by the Kaplan-Meier method and the log-rank test. The multivariate Cox proportional hazard regression analysis was conducted as well. Our data demonstrated that expression of NMNAT2 and p53 was significantly higher in CRC tissues, while NMNAT2 expression is in correlation with the invasive depth of tumors and TNM stage. Significant positive correlation was found between the expression of NMNAT2 and the expression of p53. However, NMNAT2 expression was not a statistically significant prognostic factor for overall survival. In conclusion, our results indicated that NMNAT2 might participate in tumorigenesis of CRC in a p53-dependent manner and NMNAT2 expression might be a potential therapeutic target for CRC.

Abstract 4760: Stromal contribution to the colorectal cancer transcriptome

Abstract Recent studies have identified a stem/serrated/mesenchymal (SSM) molecular subtype of colorectal cancer (CRC) that is associated with poor prognosis. We noted that genes upregulated in this subtype are also prominently expressed by stromal cells. This led us to hypothesize that the SSM transcripts could derive from the tumor microenvironment, rather than being intrinsic to cancer cells. To test this hypothesis, we analyzed microarray and RNAseq expression data from patient-derived xenografts (PDXs) of CRC, where human cancer cells are supported by murine stroma. Species-specific expression analysis revealed that mRNA levels of SSM genes are mostly due to stromal expression. Considering only genes exclusively expressed by stromal cells in PDXs, We then built three expression signatures specifically reporting the abundance of cancer-associated fibroblasts (CAFs), leucocytes or endothelial cells. In human CRC samples, all stromal signatures were strongly associated with the SSM subtype. A high CAF signature was associated with poor prognosis in untreated CRC patients, while in rectal cancer high stromal signatures jointly predicted radioresistance. These data show that the distinctive transcriptional and clinical features of the SSM subtype can be ascribed to its particularly abundant stroma. Citation Format: Claudio Isella, Andrea Terrasi, Sara E. Bellomo, Consalvo Petti, Andrea Muratore, Alfredo Mellano, Mark De Ridder, Paola Cassoni, Guy Storme, Andrea Bertotti, Enzo Medico. Stromal contribution to the colorectal cancer transcriptome. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4760. doi:10.1158/1538-7445.AM2015-4760

Peripheral blood monocytes as biomarkers for colorectal cancer.

3588 Background: Peripheral blood monocytes (PBM) represent a reservoir of inflammatory cells that contribute to cancer progression. When recruited into tumors, monocytes give rise to either macrophages or dendritic cells. Tumor-derived soluble factors can influence the differentiation of tumor-associated macrophages (TAMs) and drive their phenotype to either tumoricidal (M1) or pro-tumorigenic macrophages (M2). We hypothesized that PBM might change their phenotype already when in circulation and can be used as potential markers for early diagnosis of colorectal cancer (CRC) and its progression to metastatic disease. Methods: Monocytes were isolated by using magnetic microbeads conjugated to antibodies against CD14. In a monocentric training set we included 55 (27 non metastatic (NM), 38 metastatic (M)) untreated CRC patients. The third group included age and gender matched healthy volunteers (HV). Following RNA extraction, gene expression profiles of PBM were investigated by high throughput screening using Illumina. Statistical analysis was done on the microarray expression data to delineate a disease specific signature. A cut off was set to select the genes showing a minimum fold change of 1,5. A validation study was performed in three additional major oncological centers throughout Europe. Results: There was a broad overlap in the gene signature of PBM from NM and M patients, indicating that this signature already exists in an early stage. Then we assessed the performance of the gene signature of cancer versus HV as diagnostic tool, in particular the sensitivity and specificity, which were remarkably high in the ROC analysis. Out of 40 differentially expressed genes, 21 genes were confirmed by qRT-PCR analysis on samples processed independently. In the validation study the number of genes was downsized to a 12 gene signature without loss of specificity and sensitivity. Conclusions: This first biomarker study of PBM in CRC suggests an ‘imprinting’ of PBM by the tumor in CRC patients. Since the imprinting profile might also be reversed, the PBM signature might not only be useful in CRC diagnosis, but also for the follow-up of treated CRC patients. Overall, our data offer new opportunities to develop a non-invasive test for CRC detection and monitoring.

Baseline lymphopenia as a prognostic marker for colorectal carcinoma.

e14107 Background: Baseline lymphopenia has been proved to be a marker for poor prognosis, chemotherapy-induced toxicity and increased risk of febrile neutropenia, trombocytopenia and anemia in advanced solid neoplasms. This study aims to evaluate the effect of pretreatment lymphopenia on prognosis and hematological toxicity in colorectal cancer patients who received first line systemic chemotherapy. Methods: Lymphocyte count was evaluated in 386 pretreated colorectal cancer patients who do not have a seconder malignancy, HIV infection, bone involvement and primary G-CSF prophylaxis. Overall survival, progression free survival and disease free survival were calculated from date of diagnosis to date of relapse, progression and death. Kaplan-Meier, chi-square and Student-t test were used. Results: Mean follow-up was 30 months (range 1-180 months). Mean age was 57.4±12.5 years. Of all patients, 160 (41 %) were women. Rectum ( 26.2 %) and transvers colon (4.7 %) were the most and the least common anatomic locations, respectively. Mean lymphocyte count before treatment was 1964/µl (170-7000/µl). There were no relationship between lymphopenia and age, sex, performans status, presence of initial metastasis, adjuvant or palliative chemotherapy and hematological toxicity (p&gt;0.05). Grade 3-4 hematological toxicity was found in 40 patients and was significantly higher in patients receiving bi- or tri-chemotherapy regimen (p:0.017). Among 208 patients with relapse or progression, 40 patients had lymphopenia (19.2 %). 1, 3 and 5-year OS were significantly lower in lymphopenic patients (p:0.033). DFS was longer in non-lymphopenic patients but this data didn’t have statistical significance (p&gt;0.05). Conclusions: This study support that lymphocyte number prior to chemotherapy may be a simple but useful prognostic and predictive marker in untreated colorectal cancer patients. Patients with lower pretreatment lymphopenia have lower OS when compared to others (p&lt;0.05). This study has the highest colorectal cancer population in the literature.

Septin 9 methylated DNA is a sensitive and specific blood test for colorectal cancer.

BackgroundAbout half of Americans 50 to 75 years old do not follow recommended colorectal cancer (CRC) screening guidelines, leaving 40 million individuals unscreened. A simple blood test would increase screening compliance, promoting early detection and better patient outcomes. The objective of this study is to demonstrate the performance of an improved sensitivity blood-based Septin 9 (SEPT9) methylated DNA test for colorectal cancer. Study variables include clinical stage, tumor location and histologic grade.MethodsPlasma samples were collected from 50 untreated CRC patients at 3 institutions; 94 control samples were collected at 4 US institutions; samples were collected from 300 colonoscopy patients at 1 US clinic prior to endoscopy. SEPT9 methylated DNA concentration was tested in analytical specimens, plasma of known CRC cases, healthy control subjects, and plasma collected from colonoscopy patients.ResultsThe improved SEPT9 methylated DNA test was more sensitive than previously described methods; the test had an overall sensitivity for CRC of 90% (95% CI, 77.4% to 96.3%) and specificity of 88% (95% CI, 79.6% to 93.7%), detecting CRC in patients of all stages. For early stage cancer (I and II) the test was 87% (95% CI, 71.1% to 95.1%) sensitive. The test identified CRC from all regions, including proximal colon (for example, the cecum) and had a 12% false-positive rate. In a small prospective study, the SEPT9 test detected 12% of adenomas with a false-positive rate of 3%.ConclusionsA sensitive blood-based CRC screening test using the SEPT9 biomarker specifically detects a majority of CRCs of all stages and colorectal locations. The test could be offered to individuals of average risk for CRC who are unwilling or unable to undergo colonscopy.

Phase II trial of imatinib, bevacizumab and cetuximab plus modified FOLFOX-6 in advanced untreated colorectal cancer

e15030 Background: Imatinib inhibits PDGFR interfering with pericytes, the structural support to newly formed tumor blood vessels. It may thus synergize with bevacizumab. Microenvironment and tumor targeted agents along with chemotherapy could be a promising add-on approach. Methods: cetuximab 500 mg/m2, bevacizumab 5 mg/kg and modified FOLFOX-6 were given i.v. on day 1 and repeated every 2 weeks. Imatinib 400 mg/day per os was given continuously. Due to the cost and potential toxicity of the combination, the endpoint for this phase II study was very ambitious: at least 25% of complete response (medically or surgically achieved), lasting a minimum of 12 months in advanced untreated colorectal cancer patients with clearly unresectable disease. Results: Of 26 patients (16 with 1 site of disease), 17 completed the first 4 months of treatment according to the protocol, while 9 had to discontinue one biologic drug due to side effects (5 cetuximab, 3 imatinib and 1 bevacizumab). Grade 3–4 toxicity: diarrhea 12%, neutropenia 24%, skin rash 24%, hypersensitivity reactions 16%, asthenia 8%, neuro 8%. All patients were evaluable for response. Eleven responses ( 1 CR and 10 PR), 13 SD and 2 PD were observed, corresponding to 42% RR ( 95% CI = 23–61). The minimum follow up is 12 months; median PFS is 10 months. One patient among responders underwent radiofrequency ablation and 17 patients underwent surgery: 8 R-0, 3 R-1, 5 R-2 and one exploratory laparotomy. Major post surgical complications occurred in 5/17 patients. No evidence of macroscopic disease after the entire treatment plan was obtained in 13/26 patients: 12 surgical and 1 medical CR. Seven/13 were disease free at 6 months, but only 3 were still disease free at 12 months. ERCC1, ERCC2/XDP, GSTP1,TS,EGF, COX2, CYCLIN D, FCgR polymorphisms and K-RAS mutations were evaluated on all 26 patients, but no correlations were found with clinical outcome. Conclusions: The triple combination of biologics with modified FOLFOX-6 is feasible and tolerable as initial aggressive treatment. The primary endpoint of the study was not met . In fact the activity, 42% RR, was not outstanding and the high resectability rate, 69%, is misleading in the light of the short duration of the surgically induced CR. [Table: see text]

Clinical Significance of Serum Cytokine Measurements in Untreated Colorectal Cancer Patients: Soluble Tumor Necrosis Factor Receptor Type I – An Independent Prognostic Factor

The aim of this study was to exploit the potential clinical use of circulating cytokine measurements in colorectal cancer (CRC) patients. The levels of cytokines and cytokine receptors were assessed by ELISA in the sera of 50 healthy volunteers and 157 patients with previously untreated CRC and then related to clinicopathological features and prognosis. All tumors were verified histologically as colorectal adenocarcinomas and staged according to TNM classification. The levels of circulating interleukin (IL)-6, IL-8, macrophage colony-stimulating factor (M-CSF) and interleukin 1 receptor antagonist (IL-1ra) significantly increased with the clinical stage of CRC, and the levels of IL-6, soluble tumor necrosis factor (sTNF) receptor type I (RI), soluble interleukin 2 receptor α and TNFα with tumor grade, while IL-6, IL-8, M-CSF, IL-1ra and sTNF RI levels significantly rose with bowel wall invasion. None of the cytokine or soluble cytokine receptor levels were influenced by age, gender and colon versus rectum localization. sTNF RI, IL-8, IL-6 and vascular endothelial growth factor measurements demonstrated the highest diagnostic sensitivity. sTNF RI was found elevated in the greatest percentage of all CRC patients, in the greatest proportion of stage I patients and presented the best diagnostic sensitivity. In addition, the sTNF RI level strongly correlated with tumor grade and invasion and proved to be an independent prognostic factor.

Chronomodulated delivery schedule of oxaliplatin, 5 fluorouracil and folinic acid (FFL4/10) as first line treatment in metastatic colorectal cancer: Preliminary data of a phase II study

3738 Background: Oxaliplatin(OHP) and 5Fluorouracil(5FU) efficacy and toxicity strictly depend on circadian rhythms in cell cycle regulation, reduced glutathione content, dehydropyrimidine and thymidilate synthase activities, topoisomerase I activity, membrane permeability as well as liver and renal blood flow. Methods: From August 2002 to December 2003 (Tab.1) 27 untreated colorectal cancer patients(pt) were treated with OHP based chronomodulated delivery schedule using Melodie infusion system: OHP 25 mg / m2 /d1–4q14 (sinusoidal 12 hour infusion with flow rate peak at 4:00pm); 5FU 900 mg/m2 and L-Folinic Acid (LFA) 150 mg/m2 d1–4,q14 (sinusoidal 12 hour infusion with flow rate peak at 4:00am. Results: 147 cycles (6.11 average per pt.) were performed: average dose intensity was 48,21 mg/sqm/w, 319,28 mg/sqm/w 1630,41mg/sqm/w for OHP, LFA and 5FU respectively. Average dose according to body surface was 501mg/sqm, 3184,17mg/sqm, 16705,74mg/sqm respectively. No cycle was delayed due to toxicity. Haematological and hepatic toxicity weren't observed, neither was any gr. 4(WHO) toxicity. Radiological Objective Response evaluation has been performed in 23 pt (Tab.2). ORR was 65,22%. Median survival reached 13 months (only 3 pt died). Conclusions: These data seem confirm that FFL4/10 schedule has very interesting activity in metastatic colorectal cancer. Moreover, chronomodulated infusion is considerably less toxic than conventional flat or bolo infusion. No significant financial relationships to disclose.

Chronomodulated delivery schedule of oxaliplatin, 5 fluorouracil and folinic acid (FFL4/10) as first line treatment in metastatic colorectal cancer: Preliminary data of a phase II study

3738 Background: Oxaliplatin(OHP) and 5Fluorouracil(5FU) efficacy and toxicity strictly depend on circadian rhythms in cell cycle regulation, reduced glutathione content, dehydropyrimidine and thymidilate synthase activities, topoisomerase I activity, membrane permeability as well as liver and renal blood flow. Methods: From August 2002 to December 2003 (Tab.1) 27 untreated colorectal cancer patients(pt) were treated with OHP based chronomodulated delivery schedule using Melodie infusion system: OHP 25 mg / m2 /d1–4q14 (sinusoidal 12 hour infusion with flow rate peak at 4:00pm); 5FU 900 mg/m2 and L-Folinic Acid (LFA) 150 mg/m2 d1–4,q14 (sinusoidal 12 hour infusion with flow rate peak at 4:00am. Results: 147 cycles (6.11 average per pt.) were performed: average dose intensity was 48,21 mg/sqm/w, 319,28 mg/sqm/w 1630,41mg/sqm/w for OHP, LFA and 5FU respectively. Average dose according to body surface was 501mg/sqm, 3184,17mg/sqm, 16705,74mg/sqm respectively. No cycle was delayed due to toxicity. Haematological and hepatic toxicity weren't observed, neither was any gr. 4(WHO) toxicity. Radiological Objective Response evaluation has been performed in 23 pt (Tab.2). ORR was 65,22%. Median survival reached 13 months (only 3 pt died). Conclusions: These data seem confirm that FFL4/10 schedule has very interesting activity in metastatic colorectal cancer. Moreover, chronomodulated infusion is considerably less toxic than conventional flat or bolo infusion. No significant financial relationships to disclose.

Relationship among red blood cell polyamine content, phagocytic activities and plasma endotoxins in untreated colorectal cancer patients.

Polyamines are actively involved in immune processes and it is known that patients with cancer often exhibit immune deficits. Twenty-two patients with colorectal cancer were enrolled into this study, before starting conventional treatments. The relationship among the content of polyamines in red blood cells and phagocytosis and killing of monocytes and polymorphonuclear cells, and endotoxemia was investigated. The data show a negative correlation among levels of total polyamines and spermine and monocyte phagocytosis. Higher levels of spermine were present in patients with detectable circulating endotoxins. Our findings suggest a down-modulating effect of polyamines on the monocyte phagocytosis in untreated colorectal cancer patients; this effect could explain the presence of circulating endotoxins in cancer bearing patients.