The multidisciplinary management of gastrointestinal cancer. The integration of cytotoxics and biologicals in the treatment of metastatic colorectal cancer

The management of patients with metastatic colorectal cancer (CRC) has changed dramatically over the last five years, with increasing chances of prolonged survival. The development of new drugs has contributed to the better outcome of patients with metastatic colorectal cancer. Until the mid-1990s the only available drug, with limited activity in metastatic CRC, was 5-fluorouracil (5-FU). The development of the cytotoxic agents irinotecan, oxaliplatin and capecitabine and of the monoclonal antibodies against the vascular endothelial growth factor (VEGF) bevacizumab and against the epidermal growth factor receptor (EGFR) cetuximab and panitumumab have clearly increased the therapeutic options and have improved the outcome for patients with metastatic colorectal cancer. However, their introduction also raises many new questions and challenges.     

New antiangiogenetic agents and non-small cell lung cancer

New blood vessel formation, known as angiogenesis is a fundamental event in the process of tumor growth and metastatic dissemination. Due to its central role in tumor angiogenesis, the vascular endothelial growth factor (VEGF) and its receptor have been a major focus of basic research and drug development in the field of oncology, including the treatment of non-small cell lung cancer (NSCLC). Approaches targeting VEGF include monoclonal antibodies and vascular endothelial growth factor receptor-tyrosine kinase inhibitors (VEGFR-TKIs). Bevacizumab (Avastin) is an anti-VEGF recombinant humanized monoclonal antibody. A very recent randomized phase III trial demonstrated a statistically significant advantage in median survival favouring the combination of bevacizumab plus chemotherapy versus chemotherapy alone in the treatment of advanced non-squamous NSCLC. This study represents the first evidence of superior efficacy of targeted therapy combined with chemotherapy over chemotherapy alone in the treatment of NSCLC. ZD6474 is an orally bioavailable inhibitor of VEGFR-2 tyrosine kinase. First evidences of antitumor activity and its excellent toxicity profile make it a promising targeted agent for the treatment of NSCLC. A recent phase I/II study examined the combination of Epidermal Growth Factor Receptor (EGFR)-TKI erlotinib and bevacizumab in patients with non-squamous stage IIIB/IV NSCLC. Data on antitumor activity of this combination have to be considered very promising. Clinical trials of multiple targeted therapy may represent the second generation studies in the treatment of NSCLC.     

Inhibitory effects of docetaxel on expression of VEGF,bFGF and MMPs of LS174T cell

AIM: To study the effects of non-cytotoxic concentrations of docetaxel on some important angiogenic factors of LS174T Cells. METHODS: The non-cytotoxic concentration of docetaxel and the activity of gelatinase were determined with MTT and gelatin zymography respectively, the expression of VEGF(vascular endothelial growth factor), bFGF (basic fibroblast growth factor), MMP (matrix metalloproteinase) 2 and MMP 9 was investigated with RT-PCR and Western blot. RESULTS: The maximum non-cytotoxic concentration of docetaxel on LS174T Cells was 1.0 ng/ml. Compared with the solvent control group, 0.1, 0.5, 1.0 ng/ml of docetaxel could downregulate the expression of VEGF, bFGF, MMP 2 and MMP 9 and suppress the activity of gelatinase. CONCLUSION: Our study suggests that the non-cytotoxic concentrations of docetaxel have strong antiangiogenic activity on LS174T Cells, which suggests docetaxel may be a promising antiangiogenic agent.     

Modulation of endothelial cell network formation in vitro by molecular signaling of head and neck squamous cell carcinoma (HNSCC) exposed to cetuximab

Overexpression of EGFR plays a key-role in head and neck squamous cell carcinoma (HNSCC) and justifies the extensive use of cetuximab, a monoclonal anti-EGFR antibody, as well as EGFR-tyrosine kinase inhibitors (EGFR-TKI), which have been reported to inhibit tumor cell growth and the secretion of pro-angiogenic factors by tumor cells, such as VEGF and IL-8. Moreover, vessel normalization in tumors, suggesting a more complex mediation of endothelial cell growth control has also been observed in vivo. The present study was designed to investigate the angiogenic consequences of exposure of HNSCC tumor cell lines to cetuximab and intercellular signaling between tumor and endothelial cells by secretion of pro- and anti-angiogenic mediators in the conditioned media (CM). The results achieved showed that cetuximab decreased the secretion of VEGF by HNSCC cells and that exposure of human umbilical vein endothelial cells (HUVEC) to CM from HNSCC cells exposed to cetuximab induced an increase in endothelial cell network formation. Angiogenesis proteome profiling showed that cetuximab induced a complex alteration of the secretion of pro- and anti-angiogenic factors by HNSCC cells without enabling to identify a unique molecular marker. Expression of endothelial membrane receptors (VEGFR-2, EGFR, PECAM-1 and Notch-4) was investigated and only EGFR expression was found influenced when HUVEC were exposed to CM from cetuximab-exposed HNSCC cells. These results showed that the decrease in the secretion of pro-angiogenic agents like VEGF by HNSCC cells exposed to cetuximab could not be sufficient to justify its anti-angiogenic activity in vitro.     

西咪替丁抑制人肠癌裸鼠移植瘤

目的：研究西咪替丁对人结肠癌裸鼠移植瘤生长的抑制作用及机制。方法：建立人结肠癌裸鼠皮下移植瘤模型。随机分2组,每组5只实验鼠。肿瘤种植前3 d开始分别皮下注射生理盐水(对照组)或西咪替丁(治疗组),每天1次,观察成瘤时间及瘤体成长情况。肿瘤种植后第7周处死实验鼠,测定瘤体大小,并用免疫组化方法测定肿瘤组织内微血管密度(MVD)和血管内皮生长因子(VEGF)的表达。结果：治疗组肿瘤体积明显小于对照组;治疗组肿瘤组织中的VEGF表达程度和MVD计数亦明显低于对照组。结论：西咪替丁通过抑制VEGF表达,减少血管生成,从而抑制肿瘤生长。     

Antitumor activity and pharmacokinetics of a mixed-backbone antisense oligonucleotide targeted to the RIalpha subunit of protein kinase A after oral administration

Overexpression of the RIalpha subunit of cAMP-dependent protein kinase (PKA) has been demonstrated in various human cancers. PKA has been suggested as a potential target for cancer therapy. The goal of the present study was to evaluate an anti-PKA antisense oligonucleotide (mixed-backbone oligonucleotide) as a therapeutic approach to human cancer treatment. The identified oligonucleotide inhibited the growth of cell lines of human colon cancer (LS174T, DLD-1), leukemia (HL-60), breast cancer (MCF-7, MDA-MB-468), and lung cancer (A549) in a time-, concentration-, and sequence-dependent manner. In a dose-dependent manner, the oligonucleotide displayed in vivo antitumor activity in severe combined immunodeficient and nude mice bearing xenografts of human cancers of the colon (LS174T), breast (MDA-MB-468), and lung (A549). The routes of drug administration were intraperitoneal and oral. Synergistic effects were found when the antisense oligonucleotide was used in combination with the cancer chemotherapeutic agent cisplatin. The pharmacokinetics of the oligonucleotide after oral administration of (35)S-labeled oligonucleotide into tumor-bearing mice indicated an accumulation and retention of the oligonucleotide in tumor tissue. This study further provides a basis for clinical studies of the antisense oligonucleotide targeted to the RIalpha subunit of PKA (GEM 231) as a cancer therapeutic agent used alone or in combination with conventional chemotherapy.     

Vascular endothelial growth factor and its relationship to the prognosis and treatment of breast,ovarian, and cervical cancer

Tumor neovascularization is a complex process that plays a crucial role in the development of many different types of cancer. Vascular endothelial growth factor (VEGF) is a potent mitogen that is involved with mitogenesis, angiogenesis, endothelial survival, and the induction of hematopoiesis. By increasing vascular permeability in endothelial cells, it helps tumors recruit wound-healing proteins fibrin and fibrinogen from the plasma, suggesting that tumor formation is a process of abnormal wound healing dependent on the ability to generate a blood supply. The human female reproductive tract is highly dependent on VEGF for normal functions such as endometrial proliferation and development of the corpus luteum. The unique influence of female sex steroid hormones on the expression and activity of VEGF deems angiogenesis an important facet of the development of breast and ovarian cancer. Additionally, the up-regulation of VEGF by the E6 oncoprotein of the human papillomavirus suggests that VEGF plays an important role in the development of cervical cancer. Clinical trials have investigated the humanized monoclonal antibody bevacizumab as potential treatment for all three forms of cancer; the data show that in breast cancer, the use of bevacizumab may lengthen the disease-free survival for women with advanced breast cancer, but does not appear to change their overall survival. It may have a role as salvage chemotherapy for ovarian and cervical cancer, though further research is needed to establish it as a definitive form of treatment.     

Welche Rolle spielen die neuen Therapieoptionen in der palliativen Therapie des Kolonkarzinoms?

Therapeutic options in the treatment of metastatic colorectal cancer have recently been expanded by the introduction of two new monoclonal antibodies: bevacizumab and cetuximab. These antibodies were the proof of principle of two exciting new antitumor strategies: antiangiogenesis and inhibition of epidermal growth factor (EGF) receptor. Bevacizumab binds to vascular endothelial growth factor and thus blocks its angiogenic effects. In a randomized phase III trial bevacizumab in combination with irinotecan + 5-fluorouracil/leucovorin (IFL) was compared to chemotherapy alone as first-line treatment. The combination showed a superior response rate, a prolonged progression-free and overall survival. Cetuximab binds to the EGF receptor and thus inhibits its activation by its natural ligand. In a randomized phase II trial irinotecan refractory patients were treated with cetuximab alone or cetuximab plus irinotecan. The combination showed a response rate of 22,5% and a prolonged progression-free survival identifying cetuximab as an important new option for this patient group.     

Antitumor efficacy of cytotoxic drugs and the monoclonal antibody 806 is enhanced by the EGF receptor inhibitor AG1478

Blockade of epidermal growth factor receptor (EGFR) signaling with specific inhibitors of the EGFR tyrosine kinase retards cellular proliferation and arrests the growth of tumor xenografts. AG1478, an inhibitor of the EGFR tyrosine kinase, is used in laboratory studies; however, its therapeutic potential has not been elucidated. Therefore, we evaluated an aqueous form of AG1478 for its antitumor activity in mice bearing human xenografts expressing the WT EGFR or a naturally occurring ligand-independent truncation of the EGFR [delta2-7 (de2-7) EGFR or EGFRvIII]. Parenteral administration of soluble AG1478 blocked phosphorylation of the EGFR at the tumor site and inhibited the growth of A431 xenografts that overexpress the WT EGFR and glioma xenografts expressing the de2-7 EGFR. Strikingly, even subtherapeutic doses of AG1478 significantly enhanced the efficacy of cytotoxic drugs, with the combination of AG1478 and temozolomide displaying synergistic antitumor activity against human glioma xenografts. AG1478 was also examined in combination with mAb 806, an anti-EGFR antibody that was raised against the de2-7 EGFR but unexpectedly also binds a subset of the EGFR expressed in cells exhibiting amplification of the EGFR gene. The combination of AG1478 and mAb 806 displayed additive, and in some cases synergistic, antitumor activity against tumor xenografts overexpressing the EGFR. Here, we demonstrate that different classes of inhibitors to the EGFR can have synergistic antitumor activity in vivo. These results establish the antitumor efficacy of the EGFR inhibitor AG1478 and provide a rationale for its clinical evaluation in combination with both chemotherapy and other EGFR therapeutics.     

Immunogene therapy of tumors with vaccine based on Xenopus homologous vascular endothelial growth factor as a model antigen

Overcoming immune tolerance of the growth factors associated with tumor growth should be a useful approach to cancer therapy by active immunity. We used vascular endothelial growth factor (VEGF) as a model antigen to explore the feasibility of the immunogene tumor therapy with a vaccine based on a single xenogeneic homologous gene, targeting the growth factors associated with angiogenesis. To test this concept, we constructed a plasmid DNA encoding Xenopus homologous VEGF (XVEGF-p) and control vectors. We found that immunogene tumor therapy with a vaccine based on XVEGF was effective at both protective and therapeutic antitumor immunity in several tumor models in mice. VEGF-specific autoantibodies in sera of mice immunized with XVEGF-p could be found in Western blotting analysis and ELISA assay. The purified immunoglobulins were effective at the inhibition of VEGF-mediated endothelial cell proliferation in vitro, and at antitumor activity and the inhibition of angiogenesis by adoptive transfer in vivo. The elevation of VEGF in the sera of the tumor-bearing mice could be abrogated with XVEGF-p immunization. The antitumor activity and production of VEGF-specific autoantibodies, significantly elevated IgG1 and IgG2b, could be abrogated by the depletion of CD4(+) T lymphocytes. The observations may provide a vaccine strategy for cancer therapy through the induction of autoimmunity against the growth factors associated with tumor growth in a cross reaction with single xenogeneic homologous gene and may be of importance in the further exploration of the applications of other xenogeneic homologous genes identified in human and other animal genome sequence projects in cancer therapy.     

Targeted therapy in advanced non-small-cell lung cancer

Molecularly targeted therapies have recently expanded the options available for patients with advanced non-small-cell lung cancer (NSCLC). Two cancer cell pathways in particular have been exploited, the epidermal growth factor receptor (EGFR) and the vascular endothelial growth factor (VEGF) pathway. The former has emerged as a key regulator of cancer cell proliferation and invasion, and several EGFR inhibitors have been developed. Erlotinib, a small-molecule inhibitor of the EGFR intracellular tyrosine kinase, has been found to improve survival compared with placebo in previously treated patients with advanced NSCLC and is Food and Drug Administration (FDA)-approved in this setting. Clinical and molecular predictors of response to erlotinib, such as a history of never smoking and EGFR gene mutation or amplification, are presently being evaluated to select patients for earlier therapy with erlotinib. Additional EGFR inhibitors are also being examined in randomized trials. The VEGF pathway, a key mediator of angiogenesis, has become an attractive target in multiple malignancies, including lung cancer. Bevacizumab, a monoclonal antibody to VEGF, received FDA approval for use in advanced non-squamous-cell NSCLC in 2006 after a phase III trial reported a significant survival advantage when bevacizumab was added to standard first-line chemotherapy. Small-molecule inhibitors of the VEGF receptor tyrosine kinase, such as sunitinib and sorafenib, have also shown promise in phase II trials and are being further investigated in phase III studies. Because preclinical data suggest a synergistic effect when VEGF and EGFR inhibitors are combined, the concurrent use of erlotinib and bevacizumab has additionally been evaluated in a phase II trial, with encouraging early results suggesting at least equivalent activity to standard salvage chemotherapy, with less toxicity. Several other novel agents are being examined, including inhibitors of histone deacteylases and the 26S proteosome. Research efforts are currently focusing on tailoring such therapies according to predictive clinical and molecular markers.     

Anti-angiogenic therapy for colorectal cancer: on the way to getting better!

Based on the theory that tumor growth and metastasis depend on vessels to provide oxygen and nutrients, antiangiogenic therapy was thought as a promising approach to cure cancer. Bevacizumab is the first validated angiogenesis inhibitor, when combined with conventional treatments, which can enhance antitumor effects and prolong survival for patients with colorectal cancer. However, recent years, bevacizumab and other angiogenesis inhibitors are more discussed with drug resistance and the diverse test results. Fortunately, antiangiogenic strategy is more than bevacizumab and more than anti-vascular endothelial growth factor. Dozens of compounds that potently inhibit neoplastic blood vessels formation with different mechanisms have been developed, and many of them are being tested clinically for colorectal cancer. This review will numerate the principal antiangiogenic drugs with various mechanisms, recapitulate the information of studying these drugs for colorectal cancer treatment and try to clue better usage of antiangiogenic therapy for patients with colorectal cancer in the future.     

Palliativ intendierte First-line-Chemotherapie des metastasierten nichtkleinzelligen Lungenkarzinoms

EGFR-TKI are established as an effective option for second- and third-line-treatment of advanced non-small cell lung cancer. The combination with chemotherapy in nonselected patients did not show an additional benefit compared with chemotherapy alone. With a combination of the EGFR antibody cetuximab or the VEGF antibody bevacizumab with chemotherapy a moderate improvement of overall survival was achieved. Results of comparative trials with molecular targeted therapies and classic chemotherapy as first-line treatment are currently only available for the EGFR-TKI erlotinib and gefitinib which showed impressively the superiority in response rate, progression-free survival, toxicity, and improvement of quality of life for EGFR-TKI in patients harboring an activating mutation of EGFR. For patients with unknown EGFR mutation status or wild-type EGFR the platinum-based chemotherapy combination is still the most effective treatment option.     

Exploiting novel molecular targets in gastrointestinal cancers

Novel molecular targets are being discovered as we learn more about the aberrant processes underlying various cancers. Efforts to translate this knowledge are starting to impact on the care of patients with gastrointestinal cancers. The epidermal growth factor receptor （EGFR） pathway and angiogenesis have been targeted successfully in colorectal cancer with cetuximab, panitunumab and bevacizumab. Similarly, EGFR-targeting with erlotinib yielded significant survival benefit in pancreatic cancer when combined with gemcitabine. The multi-targeting approach with sorafenib has made it the first agent to achieve significant survival benefit in hepatocellular carcinoma. Efforts to exploit the dysregulated Akt/mTOR pathway in GI cancer therapy are ongoing. These molecular targets can be disrupted by various approaches, including the use of monoclonal antibody to intercept extracellular ligands and disrupt receptor-ligand binding, and small molecule inhibitors that interrupt the activation of intracellular kinases.     

Characterization of the CXCR4 signaling in pancreatic cancer cells

CXCL12 and its receptor, CXCR4, are emerging as promising targets for modulating growth, angiogenesis, and metastasis in several human cancers. Indeed, blocking the receptor is sufficient to prevent metastasis and angiogenesis in experimental breast cancer xenografts. Recently, the biological effect of the CXCR4 in pancreatic cancer, one of the most deadly neoplastic diseases, has been reported. However, the molecular mechanism by which CXCR4 contributes to these properties is not completely understood. In this paper, we characterize the signaling pathways activated by CXCR4 in pancreatic cancer. We show that after CXCR4 activation, EGFR becomes tyrosine phosphorylated, and the kinase activity of this receptor, together with the activation of MMPs, Src, and PI3-Kinase, is required for CXCR4-mediated ERK activation. Analysis of this cascade in pancreatic cancer cells revealed that the ERK-mediated pathway regulates genes involved in angiogenesis, such as VEGF, CD44, HIF1α, and IL-8. Furthermore, ERK blockage inhibits the migration and tube formation of endothelial cells induced by CXCL12. Considering that inhibitors for several components of this pathway, including CXCR4 itself, are at different stages of clinical trials, this study provides theoretical justification for the clinical testing of these drugs in pancreatic cancer, thus extending the list of potential targets for treating this dismal disease.     

sFlt-1 gene therapy of follicular thyroid carcinoma

Tumor progression largely depends on blood supply and neovessel formation, and angiogenesis is emerging as a promising target for cancer therapy. Vascular endothelial growth factor (VEGF), a major proangiogenic molecule, stimulates angiogenesis via promoting endothelial proliferation, survival and migration. VEGF has been found to be up-regulated in various types of tumors and to be associated with tumor progression and poor prognosis. Inhibition of VEGF or its signaling pathway has been shown to suppress tumor angiogenesis and tumor growth. In the present study, we tested the antiangiogenic and antitumor effects of soluble VEGF receptor-1 [soluble Flt (sFlt)-1] on the growth of follicular thyroid carcinoma (FTC). We constructed a 293 embryonic kidney cell line (293-Flt1-3d) that expresses sFlt-1, which is composed of the first three extracellular domains of Flt-1. The 293-Flt1-3d cells inhibited the in vitro growth of human umbilical vein endothelial cells in a paracrine manner. The in vivo antitumor and antiangiogenic activities of the 293-Flt1-3d cells were tested. When 293-Flt1-3d cells were inoculated at a site remote to the FTC-133 tumor transplant, the growth of FTC-133 tumors were inhibited by 70.37%, as compared with the control treatment with 293 cells expressing control gene LacZ. Immunohistochemical analysis of microvessel densities in treated tumors demonstrated that 293-Flt1-3d cells robustly suppressed intratumoral angiogenesis. Our data suggest that a mammalian cell-mediated approach could effectively deliver sFlt-1 gene therapy and inhibit tumor angiogenesis and tumor growth.     

VEGF-D expression correlates with colorectal cancer aggressiveness and is downregulated by cetuximab

AIM： To gain mechanistic insights into the role played by epidermal growth factor receptor （EGFR） in the regulation of vascular endothelial growth factors （VEGFs） in colorectal cancer （CRC）. METHODS： The impact of high-level expression of the growth factor receptors EGFR and VEGF receptor （VEGFR）3 and the VEGFR3 ligands VEGF-C and VEGF-D on disease progression and prognosis in human CRC was investigated in 108 patients using immu- nohistochemistry. Furthermore, the expression of the lymphangiogenic factors in response to the modulation of EGFR signalling by the EGFR-targeted monoclonal antibody cetuximab was investigated at the mRNA and protein level in human SW480 and SW620 CRC cell lines and a mouse xenograft model. RESULTS： Human CRC specimens and cell lines displayed EGFR, VEGF-C and VEGF-D expression with varying intensities. VEGF-C expression was associated with histological grade. Strong expression of VEGF-D was significantly associated with lymph node metas- tases and linked to a trend for decreased survival in lymph node-positive patients. EGFR blockade with cetuximab resulted in a significant decrease of VEGF-D expression in vitro and in vivo. CONCLUSION： In conclusion, the expression of VEGF-D in colorectal tumours is significantly associated with lymphatic involvement in CRC patients and such expression might be blocked effectively by cetuximab.     

Oncogenes as regulators of tissue factor expression in cancer: implications for tumor angiogenesis and anti-cancer therapy

Up-regulation of tissue factor (TF) is often observed in cancer. TF is a cell-associated receptor for coagulation factor VII/VIIa, an interaction known to activate the coagulation cascade. At the same time, TF is also known as a mediator of intracellular signaling events that can alter gene expression patterns and cell behavior. Both aspects of TF activity are of possible relevance to tumor growth, metastasis, and angiogenesis, including up-regulation of vascular endothelial growth factor (VEGF). TF up-regulation is often observed on the surfaces of tumor-associated endothelial cells, inflammatory cells, and particularly on cancer cells themselves. In the last case, high TF levels may be associated with poor prognosis and parallel clinical (and genetic) tumor progression. We have proposed elsewhere that TF may be a target of oncogenic events in cancer. Here we discuss our observations suggesting that oncogene-targeting agents may down-regulate TF expression. Such is the effect of treatment with the neutralizing monoclonal antibody (C225) raised against the epidermal growth factor receptor (EGFR) in EGFR-dependent squamous cell carcinoma cells (A431). This two- to threefold TF down-regulation by C225 treatment is paralleled by a decrease in expression of VEGF. It is conceivable that TF participates in signals that regulate VEGF and angiogenesis triggered by activated oncogenic pathways. Therefore, direct targeting of TF in cancer should be considered in combination with other treatment modalities such as oncogene-directed therapies, antiangiogenic agents (e.g., VEGF antagonists), and anti-cancer chemotherapy.