Tormod Kyrre Guren

Phase III Trial of Cetuximab With Continuous or Intermittent Fluorouracil, Leucovorin, and Oxaliplatin (Nordic FLOX) Versus FLOX Alone in First-Line Treatment of Metastatic Colorectal Cancer: The NORDIC-VII Study

PURPOSE The NORDIC-VII multicenter phase III trial investigated the efficacy of cetuximab when added to bolus fluorouracil/folinic acid and oxaliplatin (Nordic FLOX), administered continuously or intermittently, in previously untreated metastatic colorectal cancer (mCRC). The influence of KRAS mutation status on treatment outcome was also investigated. PATIENTS AND METHODS Patients were randomly assigned to receive either standard Nordic FLOX (arm A), cetuximab and FLOX (arm B), or cetuximab combined with intermittent FLOX (arm C). Primary end point was progression-free survival (PFS). Overall survival (OS), response rate, R0 resection rate, and safety were secondary end points. RESULTS Of the 571 patients randomly assigned, 566 were evaluable in intention-to-treat (ITT) analyses. KRAS and BRAF mutation analyses were obtained in 498 (88%) and 457 patients (81%), respectively. KRAS mutations were present in 39% of the tumors; 12% of tumors had BRAF mutations. The presence of BRAF mutations was a strong negative prognostic factor. In the ITT population, median PFS was 7.9, 8.3, and 7.3 months for the three arms, respectively (not significantly different). OS was almost identical for the three groups (20.4, 19.7, 20.3 months, respectively), and confirmed response rates were 41%, 49%, and 47%, respectively. In patients with KRAS wild-type tumors, cetuximab did not provide any additional benefit compared with FLOX alone. In patients with KRAS mutations, no significant difference was detected, although a trend toward improved PFS was observed in arm B. The regimens were well tolerated. CONCLUSION Cetuximab did not add significant benefit to the Nordic FLOX regimen in first-line treatment of mCRC.

Plasma microRNAs predicting clinical outcome in metastatic colorectal cancer patients receiving first‐line oxaliplatin‐based treatment

The conventional first‐line chemotherapy for metastatic colorectal cancer (mCRC) consists of fluorouracil (5‐FU) in combination with either oxaliplatin or irinotecan. We have explored microRNAs (miRNAs) in plasma as potential predictive markers to oxaliplatin‐based chemotherapy. The expression of 742 miRNAs was examined in plasma samples from 24 mCRC patients (12 responders and 12 non‐responders) before onset and after four cycles of 5‐FU/oxaliplatin. The top differentially expressed miRNAs between responders and non‐responders were selected for further analysis in a validation cohort of 150 patients. In the validation cohort, there was a significant overrepresentation of miRNAs with higher mean expression in the non‐responder group than in the responder group before treatment (p < 0.002). Moreover, we found three miRNAs (miR‐106a, miR‐484, and miR‐130b) to be significantly differentially expressed before treatment (p = 0.008, 0.008, and 0.008, respectively). All three miRNAs were upregulated in non‐responders. High expression of miR‐27b, miR‐148a, and miR‐326 were associated with decreased progression‐free survival (Hazard ratios (HR) of 1.4 (95% CI 1.1–1.8, p = 0.004), 1.3 (95% CI 1.1–1.6, p = 0.007), and 1.4 (95% CI 1.1–1.8, p = 0.008), respectively). miR‐326 was also associated with decreased overall survival (HR 1.5 (95% CI 1.1–2.0, p = 0.003)). There were no significantly differentially expressed miRNAs in association with clinical outcome after four cycles of chemotherapy. The present study demonstrates that plasma miRNAs analyzed before treatment may serve as non‐invasive markers predicting outcome in mCRC patients treated with 5‐FU and oxaliplatin‐based chemotherapy.

Portrait of the PI3K/AKT pathway in colorectal cancer ☆

PI3K/AKT signaling leads to reduced apoptosis, stimulates cell growth and increases proliferation. Under normal conditions, PI3K/AKT activation is tightly controlled and dependent on both extracellular growth signals and the availability of amino acids and glucose. Genetic aberrations leading to PI3K/AKT hyper-activation are observed at considerable frequency in all major nodes in most tumors. In colorectal cancer the most commonly observed pathway changes are IGF2 overexpression, PIK3CA mutations and PTEN mutations and deletions. Combined, these alterations are found in about 40% of large bowel tumors. In addition, but not mutually exclusive to these, KRAS mutations are observed at a similar frequency. There are however additional, less frequent and more poorly understood events that may also push the PI3K/AKT pathway into overdrive and thus promote malignant growth. Here we discuss aberrations of components at the genetic, epigenetic, transcriptional, post-transcriptional, translational and post-translational level where perturbations may drive excessive PI3K/AKT signaling. Integrating multiple molecular levels will advance our understanding of this cancer critical circuit and more importantly, improve our ability to pharmacologically target the pathway in view of clonal development, tumor heterogeneity and drug resistance mechanisms. In this review, we revisit the PI3K/AKT pathway cancer susceptibility syndromes, summarize the known aberrations at the different regulatory levels and the prognostic and predictive values of these alterations in colorectal cancer.

Response to transforming growth factor α (TGFα) and epidermal growth factor (EGF) in hepatocytes: Lower EGF receptor affinity of TGFα is associated with more sustained activation of p42/p44 mitogen-activated protein kinase and greater efficacy in stimulation of DNA synthesis

The epidermal growth factor (EGF) receptor mediates the effects of both EGF and transforming growth factor α (TGFα). Recent data suggested that EGF acts as a partial agonist/antagonist in hepatocytes, TGFα exerting a larger maximal stimulation of DNA synthesis than EGF. To further study the mechanisms involved in mediating the different effects of EGF and TGFα, we have examined receptor binding of the two growth factors and their action on the p42/p44 mitogen‐activated protein (MAP) kinase activity in hepatocytes. Single‐ligand concentration curves and competition experiments showed that the binding affinity to a common population of surface binding sites was about 20‐fold lower for TGFα than for EGF. MAP kinase activity responded to EGF and TGFα with different kinetics. While the two agents produced almost identical acute (5 min) stimulation (peak about fivefold), TGFα produced a more sustained MAP kinase activity than EGF. The difference between EGF and TGFα was still detectable 24 h after growth factor addition. The results show that in hepatocytes a lower receptor affinity of TGFα, as compared to EGF, is associated with a more sustained activation of the MAP kinase and a greater efficacy in the stimulation of DNA synthesis. This suggests that differential interaction of these two agents with the EGF receptor results in differences in the downstream events elicited at a given level of receptor occupancy. The data also are compatible with a role of a prolonged MAP kinase activity in the mitogenic effects of EGF and TGFα. J. Cell. Physiol. 175:10–18, 1998.

Prostaglandin E2 upregulates EGF-stimulated signaling in mitogenic pathways involving Akt and ERK in hepatocytes.

Prostaglandins (PGs) such as PGE2 enhance proliferation in many cells, apparently through several distinct mechanisms, including transactivation of the epidermal growth factor (EGF) receptor (EGFR) as well as EGFR‐independent pathways. In this study we found that in primary cultures of rat hepatocytes PGE2 did not induce phosphorylation of the EGFR, and the EGFR tyrosine kinase blockers gefitinib and AG1478 did not affect PGE2‐stimulated phosphorylation of ERK1/2. In contrast, PGE2 elicited EGFR phosphorylation and EGFR tyrosine kinase inhibitor‐sensitive ERK phosphorylation in MH1C1 hepatoma cells. These findings suggest that PGE2 elicits EGFR transactivation in MH1C1 cells but not in hepatocytes. Treatment of the hepatocytes with PGE2 at 3 h after plating amplified the stimulatory effect on DNA synthesis of EGF administered at 24 h and advanced and augmented the cyclin D1 expression in response to EGF in hepatocytes. The pretreatment of the hepatocytes with PGE2 resulted in an increase in the magnitude of EGF‐stimulated Akt phosphorylation and ERK1/2 phosphorylation and kinase activity, including an extended duration of the responses, particularly of ERK, to EGF in PGE2‐treated cells. Pertussis toxin abolished the ability of PGE2 to enhance the Akt and ERK responses to EGF. The results suggest that in hepatocytes, unlike MH1C1 hepatoma cells, PGE2 does not transactivate the EGFR, but instead acts in synergism with EGF by modulating mitogenic mechanisms downstream of the EGFR. These effects seem to be at least in part Gi protein‐mediated and include upregulation of signaling in the PI3K/Akt and the Ras/ERK pathways. J. Cell. Physiol. 214: 371–380, 2008.

Role of ERK, p38 and PI3-kinase in EGF receptor-mediated mitogenic signalling in cultured rat hepatocytes: requirement for sustained ERK activation.

Previous data disagree as to the role of extracellular signal-regulated kinase (ERK) in the EGF receptor-mediated stimulation of proliferation in hepatocytes. Using cultured rat hepatocytes, we here show that EGF receptor stimulation in mid/late G1 phase caused a sustained ERK activation that lasted for at least 48 h. Inhibition of the early part of this activity by a single addition of the MEK inhibitor PD98059 did not affect the DNA synthesis. However, inhibition of both the early and the sustained phase of ERK activation by wash-out and repeated administrations of PD98059 abolished the DNA synthesis induced by EGF and TGFα. EGF receptor stimulation also transiently activated p38, and inhibition of p38 by SB203580 markedly decreased the DNA synthesis. Furthermore, EGF and TGFα stimulated phosphorylation of Akt, a downstream target of the PI3-kinase pathway, and the PI3-kinase inhibitors wortmannin and LY294002 blocked the EGF-induced DNA synthesis. These results support a mechanism for EGF receptor-mediated mitogenic signalling in hepatocytes where ERK has an obligatory role, acting in concert with PI3-kinase, and augmented by p38. Furthermore, the data suggest that to perform this role ERK has to be activated for a prolonged period.

Let-7 miRNA-binding site polymorphism in the KRAS 3′UTR; colorectal cancer screening population prevalence and influence on clinical outcome in patients with metastatic colorectal cancer treated with 5-fluorouracil and oxaliplatin +/− cetuximab

BackgroundRecent studies have reported associations between a variant allele in a let-7 microRNA complementary site (LCS6) within the 3′untranslated region (3′UTR) of KRAS (rs61764370) and clinical outcome in metastatic colorectal cancer (mCRC) patients receiving cetuximab. The variant allele has also been associated with increased cancer risk. We aimed to reveal the incidence of the variant allele in a colorectal cancer screening population and to investigate the clinical relevance of the variant allele in mCRC patients treated with 1st line Nordic FLOX (bolus 5-fluorouracil/folinic acid and oxaliplatin) +/− cetuximab.MethodsThe feasibility of the variant allele as a risk factor for CRC was investigated by comparing the LCS6 gene frequencies in 197 CRC patients, 1060 individuals with colorectal polyps, and 358 healthy controls. The relationship between clinical outcome and LCS6 genotype was analyzed in 180 mCRC patients receiving Nordic FLOX and 355 patients receiving Nordic FLOX + cetuximab in the NORDIC-VII trial (NCT00145314).ResultsLCS6 frequencies did not vary between CRC patients (23%), individuals with polyps (20%), and healthy controls (20%) (P = 0.50). No statistically significant differences were demonstrated in the NORDIC-VII cohort even if numerically increased progression-free survival (PFS) and overall survival (OS) were found in patients with the LCS6 variant allele (8.5 (95% CI: 7.3-9.7 months) versus 7.8 months (95% CI: 7.4-8.3 months), P = 0.16 and 23.5 (95% CI: 21.6-25.4 months) versus 19.5 months (95% CI: 17.8-21.2 months), P = 0.31, respectively). Addition of cetuximab seemed to improve response rate more in variant carriers than in wild-type carriers (from 35% to 57% versus 44% to 47%), however the difference was not statistically significant (interaction P = 0.16).ConclusionsThe LCS6 variant allele does not seem to be a risk factor for development of colorectal polyps or CRC. No statistically significant effect of the LCS6 variant allele on response rate, PFS or OS was found in mCRC patients treated with 1st line Nordic FLOX +/− cetuximab.

Chemotherapy or Liver Transplantation for Nonresectable Liver Metastases from Colorectal Cancer

OBJECTIVE The primary objective was to compare overall survival (OS) in patients with colorectal cancer (CRC) with nonresectable liver-only metastases treated by liver transplantation or chemotherapy. BACKGROUND CRC is the third most common cancer worldwide. About 50% of patients will develop metastatic disease primarily to the liver and the lung. The majority of patients with liver metastases receive palliative chemotherapy, with a median OS of trial patients of about 2 years, and less than 10% are alive at 5 years. METHODS Patients with nonresectable liver-only CRC metastases underwent liver transplantation in the SECA study (n = 21). Disease-free survival (DFS) and OS of patients included in the SECA study were compared with progression-free survival (PFS) and OS in a similar cohort of CRC patients with liver-only disease included in a first-line chemotherapy study, the NORDIC VII study (n = 47). PFS/DFS and OS were estimated by the Kaplan-Meier method. RESULTS DFS/PFS in both groups were 8 to 10 months. However, a dramatic difference in OS was observed. The 5-year OS rate was 56% in patients undergoing liver transplantation compared with 9% in patients starting first-line chemotherapy. The reason for the large difference in OS despite similar DFS/PFS is likely different metastatic patterns at relapse/progression. Relapse in the liver transplantation group was often detected as small, slowly growing lung metastases, whereas progression of nonresectable liver metastases was observed in the chemotherapy group. CONCLUSIONS Compared with chemotherapy, liver transplantation resulted in a marked increased OS in CRC patients with nonresectable liver-only metastases.

Cancer therapy targeted at cellular signal transduction mechanisms: Strategies, clinical results, and unresolved issues

Much effort is currently being spent on developing anticancer drugs targeted at cellular signal transduction mechanisms, and several signal inhibitors have also been introduced into clinical practice. The rationale for such therapy is the realization that, in general, oncogenes and tumour suppressor genes encode proteins that are mutated or dysregulated forms of key components in major regulatory pathways. However, while we have witnessed striking clinical effects in certain malignancies, as in the treatment of chronic myelogeneous leukemia, the results in many other cancers have been rather disappointing, and this has raised the question of whether major advances can realistically be expected by the use of signal-targeted therapies on a broad scale. Here we briefly review the cellular and molecular basis for treatment with pharmacological signal inhibitors and the clinical experience with their use in different malignancies. We also discuss general strategies to improve the treatment, including approaches to the problem of resistance development. Clinical results vary greatly, from little or no treatment success in some cancers to an increasing number of examples of very promising responses. Progress has primarily been achieved in those malignancies and subsets of patients where the underlying molecular mechanisms have been explored in detail and the targets have been well defined. In conclusion, it is likely that novel signal-directed approaches will lead to further important advances in cancer therapy, but this will require continued efforts to identify targets that are oncogenic determinants, and systematic work to select patients for more individualized therapies.

EGF receptor-mediated, c-Src-dependent, activation of Stat5b is downregulated in mitogenically responsive hepatocytes.

Signal transducer and activator of transcription (STAT) proteins may be activated by epidermal growth factor (EGF), but their role in EGF receptor‐mediated mitogenic signaling is not clear. We previously showed that Stat5b was activated by EGF in rat hepatocytes in primary monolayer culture. In the present study, we found that EGF induced tyrosine phosphorylation of Stat5b both on Tyr‐699, which correlated with specific DNA binding activity, and also on other tyrosine residues. The Src tyrosine kinase inhibitor CGP77675 blocked the EGF‐induced activation of Stat5b, but did not affect the Stat5b activation by growth hormone (GH) or prolactin (PRL). The Stat5b response to EGF was most pronounced soon (3 h) after plating (early G1) and at high cell density (50,000 hepatocytes per cm2). However, at this cell density EGF did not stimulate DNA synthesis. In hepatocytes at 24 h of culturing (mid/late G1) with 20,000 cells per cm2, EGF induced strong phosphorylation of the EGF receptor, as well as Shc and ERK, and stimulated DNA synthesis, but did not activate Stat5b, although the Stat5b response to GH or PRL was retained. A strong GH‐induced Stat5b activation neither influenced the DNA synthesis alone nor enhanced the mitogenic effect of EGF. The results show that EGF induces tyrosine phosphorylation and DNA‐binding activity of Stat5b in a manner different from GH and PRL, apparently by a Src‐dependent mechanism. The data also provide further evidence that Stat5b is not required for mitogenic signaling from the EGF receptor.