- Email: [email protected]
Tumour necrosis factor α: a potential target for the therapy of solid tumours
Tumour necrosis factor
Review
Tumour necrosis factor : a potential target for the therapy of solid tumours Peter W Szlosarek and Frances R Balkwill
Lancet Oncol 2003; 4: 565–73
Tumour necrosis factor (TNF) is a vital cytokine involved in inflammation, immunity, and cellular organisation.1 It was first isolated from the serum of mice infected with Bacillus Calmette-Guérin treated with endotoxin, and was shown to replicate the ability of endotoxin to induce haemorrhagic tumour necrosis.2 This discovery ended a long search for an important component of “Coley’s mixed toxins”, a crude bacterial filtrate developed by Willam B Coley, a New York surgeon and pioneer of an early systemic cancer treatment at the turn of the 20th century. Coley’s mixed toxins consisted of a filtrate from cultures of Streptococcus pyogenes and Serratia marcescens and induced high fever and tumour necrosis in patients who responded, particularly those with sarcoma, but also patients with carcinoma and lymphoma.3 The phenomenon of tumour necrosis is now attributed, in part, to a vascular thrombotic mechanism, mediated by a surge of macrophage-derived TNF, released after activation by the endotoxin contained within Coley’s toxins. In the 1980s, TNF was also characterised as cachectin4 for its role in wasting—first indentified in rabbits infected with Trypanasoma brucei—and as T-cell differentiation factor.5 Cloning of the TNF gene in 1984 led to a decade of clinical experimentation, culminating in a licence from the European Medicine Evaluation Agency for its use as a locoregional treatment for inoperable sarcoma.6 However, systemic TNF therapy proved ineffective with a lack of objective tumour response and serious side-effects (hypotension and organ THE LANCET Oncology Vol 4 September 2003
Rights were not granted to include this image in electronic media. Please refer to the printed journal. © Wellcome Photo Library/T Blundell and N Campillo
Tumour necrosis factor (TNF), named for its antitumour properties, was isolated almost 30 years ago. It is a vital member of the multifunctional TNF superfamily and has important roles in immunity and cellular remodelling as well as influencing apoptosis and cell survival. Its central role in inflammation has led to the development of TNF antagonists as effective therapies for rheumatoid arthritis and inflammatory bowel disease. In this review, we discuss the evidence which has accumulated during the past decade that implicates TNF in inflammatory pathways that increase tumorigenesis. There is convincing evidence that under specific conditions TNF is a tumour promoter and helps to produce the toxic effects associated with conventional cancer therapy, such as the cytokine release syndrome and cisplatin-induced nephrotoxicity. Several trials have been set up to investigate the role of TNF antagonists in cancer. It is hoped that these agents inhibit the neoplastic process either alone or in combination with other agents, and ameliorate the side effects of cancer therapy.
Figure 1. Ribbon structure of tumour necrosis factor.
failure), which redirected researchers towards alternative forms of local delivery.7 Interest in the use of TNF mutants and heat-shock proteins to increase the therapeutic window of TNF therapy remains experimental. In parallel with the emerging clinical data for TNF therapy in malignant disease, increasing evidence attributed a role for TNF in rheumatoid arthritis, inflammatory bowel disease, diabetes, sepsis, and several infections, including HIV.1 Several studies have implicated endogenous TNF in tumour-cell growth and stromal interactions that facilitate tumour invasion and metastasis. Although TNF is capable of initiating a tumour apoptotic response (with or without an immune-mediated mechanism) the reality is that these pathways are frequently deactivated within tumour cells (figure 2). In some circumstances, TNF can provide a survival signal for the cancer cell and hence it has been referred to as a tumour promoting factor.8 Furthermore, the concept of cancer as a purely genetic disease has been re-evaluated and the importance of the tumour microenvironment and inflammation in cancer progression PWS is a Clinical Research Fellow funded by Cancer Research UK and FRB is Professor of Cancer Biology at Barts and The London and Co-Director of the Translational Oncology Laboratory, Cancer Research UK, London, UK. Correspondence: Dr Frances Balkwill, Cancer Research UK, Translational Oncology Laboratory, Barts and The London, Queen Mary’s School of Medicine and Dentistry, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ, UK. Tel: +44 (0)20 7882 6106. Fax: +44 (0)20 7882 6110. Email: [email protected]
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
565
Tumour necrosis factor
Review has been emphasised.9,10 As a central mediator of inflammation, TNF provides a molecular link between chronic inflammatory stimuli and the subsequent development of malignant disease. Its paradoxical role in cancer progression has stimulated interest in the use of TNF antagonists for the prevention and treatment of cancer. The use of TNF antagonists in haematological cancers has recently been discussed, providing a rationale for their clinical use.11 In this review, we concentrate on the case for modulating TNF in solid tumours.
TNF and TNF receptor signalling TNF is a soluble 17 kDa molecule (157 aminoacids) that binds as a homotrimer to two distinct homotrimeric receptors on the cell surface: TNFRI (p55 receptor) and TNFRII (p75 receptor).12,13 TNF is synthesised as a 26 kDa (233 aminoacids) membrane-bound propeptide (proTNF) and is secreted after cleavage by TNF-converting enzyme (TACE). The 26 kDa form is also functional and binds to TNFRII via direct cell-to-cell contact. Although activated macrophages are a major source of TNF, it can be made by various other cells including fibroblasts, astrocytes, Kupffer cells, smooth-muscle cells, keratinocytes, and tumour cells. The promoter region of the TNF gene contains binding sites for several transcription factors, including NFB, cyclic AMP response element (CRE), activator proteins (AP) 1 and 2, and nuclear-factor-activated T cells. Post-transcriptional regulation of TNF gene expression occurs via an AU-rich element (ARE) within the 3' untranslated region, which interferes with mRNA stability and translation efficiency. For example, deletion of ARE from the genomic TNF locus causes basal concentrations of TNF to increase, producing an inflammatory phenotype in mice (rheumatoid arthritis and inflammatory bowel disease). A similar disorder is observed in mice deficient in tristetraprolin, an endogenous RNA-binding protein that regulates TNF mRNA stability. Lastly TACE, a member of the ADAM family of metalloproteinases, releases the soluble mature form of TNF and its receptors via a protein kinase C and metalloproteinase-dependent signalling pathway. Processed TNFRs provide additional negative regulation in the form of soluble TNF binding proteins. The signalling cascade downstream of TNFRI and TNFRII is complex and involves many adaptor proteins, which are recruited after binding of the TNF ligand to its receptor. Figure 2 summarises the main elements of the TNFR signalling pathway, in particular pathways implicated in inflammation and carcinogenesis. TNFRI is ubiquitously expressed on mammalian cells and is therefore involved in various biological effects after treatment with TNF including: regulation of apoptosis, cell proliferation, differentiation, immunity, and inflammation. Recruitment of TNF to TNFRI results in dissociation of the silencer of death domain protein (SODD), whose function is to repress TNFRI signalling by blocking the binding of adaptor proteins to the death domain of TNFRI. Release of SODD permits the assembly of an activated signalling complex at the death domain of TNFRI, consisting of TNFR-associated death domain (TRADD), TNFR-
566
associated factor 2 (TRAF2), receptor-interacting protein (RIP), and FAS-associated death domain (FADD). These molecules regulate at least four distinct pathways: a proapoptotic pathway that is induced by binding caspase 8 to FADD; an antiapoptotic programme that is activated by the binding of cellular inhibitor of apoptosis protein 1 (cIAP-1) to TRAF2; AP1 activation mediated through TRAF2 via a JNK-dependent kinase cascade; and NFB activation by RIP.14 The regulation of NFB by TNF is an example of a signalling pathway of fundamental importance in carcinogenesis.15 NFB activation requires phosphorylationdependent degradation of inhibitor of B (IB) proteins, which sequester NFB in the cytoplasm. This step is mediated via a multiprotein IB kinase (IKK) complex consisting of two catalytic subunits (IKK and IKK) and a regulatory subunit, IKK, which is in turn activated by RIP. Through this pathway, TNF targets IB to the proteosome for ubiquitination and stimulates translocation of NFB to the nucleus. Although elucidation of TNFRI signalling has in recent years progressed significantly, the factors controlling outcome in a particular cell (eg, the dominance of NFB over caspase activation in a specific cell) are poorly understood. TNFRII signalling is largely confined to endothelial and haematopoietic cells and not much is known about its structure and function. However, pro-TNF, in particular, has a higher affinity for TNFRII and faster dissociation kinetics from this receptor than soluble TNF. There is also evidence of reverse signalling, which involves binding of pro-TNF to TNFRII to initiate retrograde signal transduction. Despite the lack of a death domain, and the recruitment of fewer adaptor molecules than TNRI, TNFRII is able to affect NFB and JNK signalling.
TNF and experimental cancers TNF knockout mice (TNF-/-) have provided evidence of a tumour immunosurveillance role for TNF16,17 and implicated the cytokine in tumour development and metastasis.18,19 Moore and colleagues found that TNF-/mice had ten times fewer skin tumours than wild-type mice after initiation with a carcinogen and repeated application of 12-O-tetradecanoylphorbol-13-acetate (TPA). Although TNF was important for de-novo carcinogenesis, the later stages of tumour progression were similar in TNF-/- and wild-type mice. The resistance of TNF-/- mice to skin carcinogenesis is likely to be related to a temporal delay in the activation of protein kinase C and AP1, which are important TPA-responsive signalling molecules.20 This delay seems to affect the expression of genes involved in tumour development, eg, granulocyte-macrophage colonystimulating factor and matrix metalloproteinases 3 and 9, which are suppressed in the TNF-/- mice but not in wildtype mice exposed to carcinogen. TNFRI-/- mice, and to a lesser extent TNFRII-/- mice, also show resistance to skin carcinogenesis (unpublished data). The tumour-promoting role of TNF was also investigated in models of hepatic carcinogenesis,21 in which TNFRI-/- and TNFRII-/- mice have proven useful for elucidating the mechanism of action involved.22 TNFRI-/-
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
Review
mice had reduced oval cell (hepatic stem cell) proliferation during the preneoplastic phase of liver carcinogenesis, which was associated with fewer tumours. Liver tumorigenesis was unaffected in TNFRII-/- mice. Kitakata and co-workers used TNFRI-/- mice to show that endogenous TNF is vital for promoting liver metastasis after intrasplenic administration of a colonic adenocarcinoma cell line.23 Although there was no difference in the size of the splenic tumours, more than 90% of wild-type mice developed liver metastases compared with less than 50% of TNFRI-/- mice. Furthermore, TNFRI-/- mice had smaller intrahepatic metastases. In wildtype mice, TNF (mRNA and protein) was induced around the hepatic central and portal veins by day 3 and in hepatic metastases after 24 days. The difference in metastatic behaviour was accounted for by increased VCAM1 mRNA and protein expression in the livers of wild-type animals, which was significantly attenuated in TNFRI-/- mice. Lastly, the fatal lymphoproliferative disorder that develops in FASL-deficient mice was attenuated by crossing these animals with TNF-/- mice.24 TNF may have induced the expression of chemokines (chemotactic cytokines), which regulate trafficking and accumulation of tumour cells into lymph nodes.24 Transgenic mice overexpressing TNF are unsuitable for studying the role of TNF in tumorigenesis because they are affected by early and severe inflammatory pathologies, such as pulmonary hypertension, cardiac failure, neurodegeneration, and arthritis.13 Nevertheless, several animal experiments that manipulate concentrations of TNF within the tumour microenvironment have confirmed that an increase in TNF promotes cancer development and spread. Orosz and colleagues showed increased lung metastases from
an experimental fibrosarcoma after pretreatment of the animals with TNF.25 Alternatively, lung metastases were reduced by the addition of an anti-TNF antibody, which neutralised endogenous levels of TNF. Malik and colleagues reported that overexpression of TNF conferred invasive properties on xenograft tumours.26 However, the effect is clearly dependent on tumour type because the TNF transgene could either induce tumour rejection or decrease survival, for example, by promoting hepatic metastases.27 Monoclonal antibodies to TNF reversed the hepatic metastases and increased survival. Further evidence for a tumour-promoting role of TNF has been provided by chemical carcinogenesis experiments with BALB/3T3 fibroblasts. BALB/3T3 cells in nude mice that were treated with a chemical carcinogen and then exposed for 2 weeks to TNF underwent transformation and yielded tumours.28 Human xenograft studies of ovarian cancer in nude mice have also emphasised the paradoxical effects of TNF with conversion of ascites into peritoneal deposits.29 TNF and cachexia
Studies of cancer cachexia in mice have confirmed that TNF is an important mediator of the phenomenon— alongside interleukins 1 and 6 and interferon .30,31 A study of TNFRI-/- and TNFRII-/- mice assessed the contribution of tumour and host TNF to cancer cachexia. Cachexia was not suppressed in the TNFR knockout mice, suggesting that tumour-derived TNF is more important than host TNF.32 Human xenograft data supports a role for TNF in mediating cachexia and other paraneoplastic features, such as hypercalcaemia and leucocytosis.33 These features could all be reversed with anti-TNF antibodies.
TNF as a tumour promoter TNF bound to cell membrane
Soluble TNFR
TACE TNF
Pro-TNF
TNFRI
TACE
Cell membrane
TNFRII
SODD TRADD cIAPS
FADD
Caspase 8
RIP
TRAF2
MAPKK
IKK complex NFB-kB(cytostolic)
JNK/p38
APOPTOSIS
AP1
NFB(nuclear)
Proliferation Cancer cell survival Inflammation
Figure 2. TNF binds to TNFRI and TNFRII activating several pathways associated with inflammation, apoptosis, and cell survival.
THE LANCET Oncology Vol 4 September 2003
Evidence for a role of TNF has come from several preclinical and clinical studies. Expression studies have confirmed abnormally high concentrations of TNF in tumours, implicating the cytokine in specific preneoplastic lesions and in advanced malignant disease. Data from studies in vitro and in vivo suggests that TNF, far from being an epiphenomenon, is an active player in tumorigenesis. Furthermore, several well-known paraneoplastic syndromes that commonly occur in patients with cancer seem to be caused by secreted tumour/host factors, including TNF. Finally, several single nucleotide polymorphisms (SNPs) that may influence the extent of TNF expression are associated with an increased risk of cancer. TNF in the tumour microenvironment
In-situ hybridisation and immunohistochemical studies have confirmed
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
567
Tumour necrosis factor
Review tumour and stromal expression of TNF in breast, ovarian, colorectal, prostate, bladder, oesophageal, and renal-cell cancers, and in melanoma, lymphoma, and leukaemia. The extent of TNF expression varies between tumours, as does its location, and the accompanying cytokine profile. For example, in ovarian cancer TNF is expressed mainly within tumour epithelial islands (70% of assessed tumours)34 whereas predominant expression in colorectal cancer is within tumour-associated macrophages.35 In ovarian cancer, TNFRI is found on tumour and stromal cells and TNFRII localises to the leukocyte infiltrate, suggesting potential for both autocrine and paracrine actions of endogenous TNF. Moreover, TNF activity seems to be deregulated with inappropriate constitutive expression in ovarian tumour epithelium compared with normal ovarian surface epithelium.36 Studies of serous ovarian carcinoma, prostate carcinoma, non-Hodgkin lymphoma, acute leukaemia, chronic myelogenous leukaemia, and chronic lymphocytic leukaemia have found an association between expression of the cytokine, poor survival, and resistance to therapy.34,37–41 By use of immunohistochemical analysis, researchers found overexpression of TNF, inducible nitric oxide synthetase (iNOS) and adrenocorticotrophic hormone peritumoral lymph nodes and not in mesenteric (control) lymph nodes (the two are morphologically identical) from patients with colorectal cancer,42 suggesting a role for these molecules in early lymphatic spread. Recent data from a study of gastrooesophageal cancer showed progression in TNF and TNFRI epithelial expression along the Barrett’s metaplasia-dysplasiacarcinoma sequence (73% of tumours; figure 3).43 Tselepsis and colleagues reported that TNF mediated upregulation of the c-MYC oncogene, via -catenin and a MAPK (p38 and ERK) dependent pathway in CACO2, a cell line which differentiates into small-intestinal-like enterocytes in vitro and simulates Barrett’s metaplasia. Moreover, E cadherin, an important cell-adhesion molecule is negatively regulated by TNF and in common with several other epithelial cancers, is downregulated in Barrett’s adenocarcinoma, with corresponding increases in cytoplasmic and nuclear pools of -catenin. Mechanism of tumour promotion
TNF is rarely cytotoxic to tumour cells in vitro unless an additional stressor (eg, a RNA or protein synthesis inhibitor) is provided. Tumour cells are often resistant to the cytokine via a mechanism involving manganese superoxide dismutase, which scavenges oxygen free radicals induced by TNF.44 The various mechanisms by which TNF promotes cancer growth, invasion, and metastasis are shown in the panel. TNF acts as a growth factor in certain tumour types, increasing concentrations of positive cell-cycle regulators (and decreasing levels of CDK inhibitors) and components of growth-factor-receptor signalling pathways such as RAS or c-MYC.43,45,46 Other direct roles of endogenous TNF include inducing chemoresistance in several cancers47 and mediating androgen independence in prostate cancer.37 TNF also promotes DNA damage and inhibits DNA repair by upregulating nitric oxide (NO) dependent (non-cGMP) pathways.48 Moreover, both argininosuccinate synthetase (the rate-limiting enzyme in NO synthesis) and iNOS are
568
Tumour-promoting effects of TNF l
Production of NO (DNA/enzyme damage, cGMP-mediated tumour promotion)
l
Autocrine growth and survival factor for malignant cells
l
Activation of E6/E7 mRNA in HPV-infected cells
l
Tissue remodelling via induction of matrix metalloproteinases
l
Control of leukocyte infiltration in tumours via modulation of chemokines and their receptors
l
Downregulation of E cadherin, increased nuclear pool of catenin
l
Increase tumour-cell motility and invasion
l
Induction of angiogenic factors
l
Loss of androgen responsiveness
l
Resistance to cytotoxic drugs
induced by various cytokines in tumour cell lines, with increased iNOS present in various solid tumours, including gynaecological cancers and breast cancer. Low concentrations of NO are implicated in many aspects of tumorigenesis. The tumour stroma is modulated in several ways by TNF. Although TNF can induce collapse of tumour vasculature via downregulation of V3 signalling and an increase in angiostatin,7 the cytokine can also promote angiogenesis.49 Various factors mediate the proangiogenic effects of TNF including VEGF and its receptor VEGFR2, bFGF, interleukin 8, platelet activating factor, ephrin A (a ligand for the Eph family of receptor tyrosine kinases), NO, E selectin, ICAM1, and thymidine phosphorylase.50–54 The paradoxical effects of TNF on tumour vasculature may reflect the difference in chronic synthesis (which favours angiogenesis) and acute high-dose local administration (which triggers vascular thrombosis). TNF promotes further tumour remodelling by stimulating fibroblast activity, tumour-cell motility, and tumour invasion via the induction of matrix metalloproteinases.55–57 Paraneoplastic phenomena
An increasing tumour burden is characterised by progressive weight loss with muscle and lipid wasting known as cachexia. This syndrome is driven by several tumour and host factors including cytokines, such as TNF, interleukins 1 and 6, and interferon . Although data from animal studies of the involvement of TNF is convincing, clinical studies evaluating the cytokine in human cachexia have been less consistent.58 However, the data for specific tumours, for example pancreatic carcinoma, is more robust and recent studies have confirmed increased TNF (mRNA and protein) concentrations in blood from patients with cachexia. Other paraneoplastic features have been linked to TNF including fever, hypercalcaemia, anaemia, and fatigue typical of many patients with advanced disease. TNF has been characterised as an endogenous pyrogen, osteoclast activating factor facilitating skeletal metastases, and as a suppressor of erythropoietin production.13 SNPs and susceptibility to cancer
If TNF is an endogenous tumour promoter, it is possible that functional SNPs in the genes for TNF, TNFRs, or
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
Review Treatment-related complications
B
A
Courtesy of Janusz Jankowski
Several proinflammatory cytokines, such as TNF and interleukin 6, mediate cytokine release syndromes, which complicate antibody-based cancer therapy. For example, several studies have shown frequent first-dose infusion-related toxic effects with rituximab (a monoclonal antibody to the CD20 cell-surface antigen) in lowD C grade non-Hodgkin lymphoma— symptoms consist mainly of fever, rigors, and chills. Studies of rituximab in patients with haematological cancer have confirmed TNF release as a major biological event, correlating with these clinical features.65 More severe reactions of dyspnoea, Figure 3. Immunolocalisation of TNF in Barrett's oesophagus (TNF antibody, DAB and hypotension, abdominal pain, and haematoxylin counterstain). TNF expression increases with progression from normal squamous vomiting are less common with epithelium (weak staining in basal layers and submucosa only) along the Barrett's metaplasiarituximab (less than 10%) and have dysplasia-adenocarcinoma sequence. A benign gland entrapped within the tumour (arrow) with no also been reported in solid tumour TNF expression, contrasts with strong expression in the adenocarcinoma cells. therapy with a similar patterns of events downstream in the TNF pathway may be TNF production. The mechanism downstream of TNF is associated with cancer severity and susceptibility. Several unclear, but is thought to involve the release of NO and studies of small numbers of patients have reported leukostasis in pulmonary vasculature. associations between several TNF SNPs in various Ramesh and Reeves showed a major role for TNF in cancers. For example, several studies have linked SNPs in mediating cisplatin-induced nephrotoxicity in mice.66 the promoter region of TNF and the related cytokine Although several proinflammatory cytokines and chemolymphotoxin to cancer susceptibility and severity. In a kines are upregulated during cisplatin-induced injury, study of 273 patients with lymphoma, the TNF308 including TGF, RANTES, MIP2, and MCP1, the protection polymorphism was associated with high plasma afforded by TNF antagonists or deletion of the TNF gene, concentrations of the TNF at first presentation. shows the absolute requirement for TNF. The cytokine Furthermore, possession of at least two TNF/ initiates nephrotoxicity by two mechanisms: a direct lymphotoxin high producer alleles was a risk factor for apoptotic effect on renal epithelial cells (via caspase 8); and first-line treatment failure, shorter progression-free by regulating a cytokine network, which promotes a large survival, and overall survival.59 In addition, individuals influx of leukocytes within the kidney cortex. The latter with polymorphisms associated with a high production of produces oxidant stress causing further damage and release TNF or lymphotoxin are also at significantly increased of TNF. Bleomycin-induced pulmonary fibrosis is another risk of developing monoclonal gammopathy of uncertain example of a toxic effect that is linked to proinflammatory significance and myeloma.60 Associations have also been cytokines and an alveolar leukocyte reaction.67 Moreover, the found between SNPs in the promoter region of TNF and polymorphic TNF2 microsatellite, an allele in the susceptibility to renal and prostate cancer. Patients with promoter region of the TNF gene, is associated with an genotype GA at locus -238 had a 6·5 times higher risk of increased risk of bleomycin-induced lung injury.68 renal cancer and a GA genotype at locus 488 was associated with a three times higher risk. The incidence of prostate TNF antagonists cancer with genotype GA at locus 488 was 17 times Specific TNF antagonists, such as infliximab and etanercept, are now accepted as successful treatments for higher.61,62 There are also two associations between TNF various inflammatory and autoimmune conditions.69 In light microsatellite polymorphisms and human cancer. In of the evidence outlined in this review, and the similarities in cutaneous basal-cell carcinoma, individuals with the TNF1 inflammatory pathways in cancer and rheumatoid arthritis and 7 containing genotypes showed an increased (figure 4), a case can be made for the use of TNF susceptibility to the disease.63 Those with cutaneous basal- antagonists in solid tumours. The behaviour of the cell carcinoma with TNF alleles d4 and d6 and the TNF2- inflammatory pannus of rheumatoid arthritis may be b4-d5 haplotype had more tumours. Secondly, a relative risk likened to the invasion and metastasis of cancer, with the of 15 for cervical intraepithelial neoplasia was conferred by upregulation of angiogenic factors, matrix metalloproteases, the combination of TNF11, HLA-DQ, and HPV16 cytokines, and chemokines. In addition, studies in patients positivity.64 with rheumatoid arthritis have shown that infliximab THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
569
Tumour necrosis factor
Review
RA autoimmune trigger
Tumour initiation RhF Trigger for TNF
Helper T cell Bystander activated T cell
B cell +
Trigger for TNF Transformed cell
TNF
Macrophage
+
+ TNF
+
Endothelial cell Macrophage
Endothelial cell
Fibroblast
Extracellular matrix Fibroblast
Chondrocyte Osteoclast
Erosion of bone and joint cartilage
Cell proliferation/survival Angiogenesis Leukocyte infiltration Matrix metalloproteinases Cytokine deregulation
Cancer progression
Figure 4. TNF pathways common to rheumatoid arthritis (RA) and tumourigenesis. Both RA and certain epithelial cancers (eg, ovarian cancer) are characterised by TNF-dependent processes such as cellular proliferation, angiogenesis, leukocyte inflitation, and the upregulation of matrix metalloproteases.
treatment reduces production of inflammatory cytokines, angiogenic factors, and matrix metalloproteinases, inhibits leukocyte trafficking to inflammed joints, and improves bone-marrow function.69 All these actions could be useful in a biological cancer therapy. TNF antagonists used to treat haematological cancers have confirmed previous safety data. Reports of activity are beginning to emerge, with improvements in bone-marrow function in myelodysplastic syndrome, myelofibrosis, and acute myelogenous leukaemia, and indications of a significant role in acute and chronic graft versus host disease.11 Several studies of TNF antagonists alone or in combination with chemotherapy have now begun in various solid tumours, and also for the palliative treatment of cancer cachexia. Preliminary reports of patients with breast cancer treated with TNF antagonists alone suggest biological activity with no significant toxic effects as determined by a whole blood cytokine release assay for MCP1 and interleukin 6, but no evidence of clinical response.70 A phase II trial of etanercept in patients with advanced ovarian cancer is currently underway assessing various cytokine parameters and CA125 as surrogate endpoints. Other studies of TNF antagonists are in progress in patients with non-small-cell
570
lung cancer (in combination with docetaxel) and pancreatic cancer (in combination with gemcitabine). There are unpublished case reports of infliximab with activity in patients with breast cancer and bone metastases as assessed by a reduction in pain symptoms and improvement on bone scintigraphy (DeWitte M, personal communication). A recent report of infliximab improving pulmonary fibrosis in a patient with rheumatoid arthritis would also be worth pursuing in the context of bleomycin-induced lung injury.71 Infliximab
Infliximab is a humanised monoclonal antibody (murine Fv, human IgG1) with both direct anti-TNF effects and the potential for antibody-dependent cellular cytotoxicity. It is licensed for use in rheumatoid arthritis and Crohn’s disease, with experimental use in various other inflammatory conditions, including juvenile Still’s disease, psoriatic arthropathy, and Behçet’s disease. The antibody is usually given at doses of 3–5 mg/kg every few weeks intravenously (in rheumatoid arthritis and irritable bowel disease). There is an increased risk of infection (particularly tuberculosis) and of certain autoimmune conditions (eg, exacerbation of demyelinating disease and a lupus-like syndrome with
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
Review
Search strategy and selection criteria A search of PubMed was carried out using combinations of the terms “TNF-alpha”, “TNFRI”, “TNFRII”, “knockout”, “tumour”, “carcinogenesis”, “inflammation”, “cachexia”, and “cytokine release syndrome”. Additional information on infliximab, etanercept and rituximab was obtained from the Electronic Medicines Compendium (http://emc.vhn.net/) and from investigators involved in clinical trials of these agents.
autoantibodies to double-stranded DNA), in which TNF has an important immunoregulatory role. This effect is consistent with data from TNF knockout mice. Because of infliximab’s slow elimination from the body and the potential for infective and autoimmune complications close monitoring is recommended for 6 months after cessation of therapy. Antibodies to infliximab have been detected in patients experiencing infusion-related reactions but these reactions lessen with repeated administration. Over 100 000 patients have been treated to date with infliximab and the observed incidence of malignant disease has been similar to those expected for the populations studied. Several other antibodies targeting TNF have recently become available: CDP571, a humanised murine IgG4 monoclonal antibody and D2E7, a human IgG1 monoclonal antibody. Etanercept
Etanercept, a dimeric molecule consisting of two molecules of the extracellular portion of the human TNFRII fused to the Fc portion of human IgG, is synthesised using recombinant DNA technology in the Chinese hamster ovary mammalian cell expression system. The drug is given in twice weekly subcutaneous injections (25 mg adult dose); it has a half life of 70 h and is renally excreted. Although antibodies to etanercept are seen, these are non-neutralising and have no effect on clinical outcome. Etanercept is currently licensed for use in rheumatoid arthritis but lacks efficacy in irritable bowel disease. Both drugs are associated with similar adverse events and contraindications. A pegylated truncated form of TNFRI is an another example of a TNF antagonist under clinical development. Non-specific TNF inhibitors
Thalidomide, dexamethasone, aspirin, and non-steroidal anti-inflammatory drugs (NSAIDs) are all non-specific TNF inhibitors, which modulate TNF-dependent and independent pathways. Dexamethasone, a potent antiinflammatory and immunosuppressive drug and one of the first non-specific TNF inhibitors clinically available, has widespread application in oncology. It inhibits TNF transcription and translation. Thalidomide, with more subtle immunomodulatory actions, also represses TNF in several ways, promoting decay of the mRNA transcript and decreasing NFB binding activity.72 Several studies have confirmed that thalidomide has activity in a range of tumours, including multiple myeloma, renal cell and prostate carcinoma, melanoma, glioma, and Kaposi’s sarcoma. Patients have also reported improvements in symptoms of anorexia, weight loss, nausea, insomnia, and profuse sweating in the palliative-care setting. THE LANCET Oncology Vol 4 September 2003
There is good epidemiological evidence for a protective role of aspirin and NSAIDs against colorectal cancer, and emerging data suggest a benefit in several other epithelial tumours.73 Both groups of anti-inflammatory drugs inhibit cyclo-oxygenase 2 (COX2), which is induced by TNF and other inflammatory cytokines via a NFB pathway, and is implicated in tumorigenesis. A recent randomised doubleblind study in patients with familial adenomatous polyposis of celecoxib, a specific COX2 antagonist, showed a reduction in colorectal polyps. Several studies of aspirin and other COX2 inhibitors for chemoprevention and treatment of cancer are currently underway. Interest has increased in other agents with anti-TNF activity, namely curcumin (a constituent of turmeric) and green tea, which may yet prove to have important roles in cancer chemoprevention.74,75
Conclusion TNF has a paradoxical role in cancer, inducing destruction of blood vessels and cell-mediated killing of certain tumours, as well as acting as a tumour promoter. Chronic endogenous low production of TNF favours the latter. TNF may contribute to the development of tissue architecture necessary for tumour growth and metastasis, may induce other cytokines, angiogenic factors, and matrix metalloproteinases, contributing to DNA damage and increase growth and survival of tumour cells. Thus, TNF provides a molecular bridge between inflammation and cancer. On the basis of the success of TNF antagonists in inflammatory disease, further investigation of these therapies is warranted to combat cancer, complications of cancer, and the adverse events of conventional treatment. The study of TNF has come a long way from the early days of William Coley and we can now manipulate concentrations within tumours by using supraphysiological concentrations or antagonising endogenously secreted TNF. Only time will tell whether TNF is a useful target for cancer therapy or whether tumour promoting factor would be a more appropriate name. Conflicts of interest
FRB is a consultant for Centocor Inc, manufacturers of infliximab, who also provide funding for research in the Translational Oncology Lab (Cancer Research UK) into preclinical studies of TNF antagonists in cancer models. All the published work on TNF and cancer done in our laboratory was carried out before industry collaboration. Acknowledgments
We thank Jannusz Janowski and Fionnula Brennan for assistance with figures 3 and 4, respectively. References
1 Locksley RM, Killeen N, Lenardo MJ, et al. The TNF and TNF receptor superfamilies: integrating mammalian biology. Cell 2001; 104: 487–501. 2 Carswell EA, Old LJ, Kassel RL, et al. An endotoxin-induced serum factor that causes necrosis of tumors. Proc Natl Acad Sci USA 1975; 72: 3666–70. 3 Nauts HC, Fowler GA, Bogatko FH. A review of the influence of bacterial infection and of bacterial products (Coley’s toxins) on malignant tumors in man. Acta Medica Scandinavia 1953; 145: S276. 4 Beutler B, Greenwald D, Hulmes JD, et al. Identity of tumour necrosis factor and the macrophage-secreted factor cachectin. Nature 1985; 316: 552–54. 5 Takeda K, Iwamoto S, Sugimoto H, et al. Identity of differentiation
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
571
Tumour necrosis factor
Review
6 7
8 9 10 11 12 13
14 15 16
17 18 19 20 21 22 23 24 25 26 27
28 29 30 31
inducing factor and tumour necrosis factor. Nature 1986; 323: 338–40. Lejeune FJ, Ruegg C, Lienard D. Clinical applications of TNF-alpha in cancer. Curr Opin Immunol 1998; 10: 573–80. Weichselbaum RR, Kufe DW, Hellman S, et al. Radiation-induced tumour necrosis factor- expression: clinical application of transcriptional and physical targeting of gene therapy. Lancet Oncol 2002; 3: 665–71. Balkwill F. Tumor necrosis factor or tumor promoting factor? Cytokine Growth Factor Rev 2002; 13: 135–41. Hanahan D, Weinberg RA, The hallmarks of cancer. Cell 2000; 100: 57–70. Balkwill F, Mantovani A. Inflammation and cancer: back to Virchow? Lancet 2001; 357: 539–45. Tsimberidou AM, Giles FJ. TNF-alpha targeted therapeutic approaches in patients with hematologic malignancies. Expert Rev Anticancer Ther 2002; 2: 277–86. MacEwan DJ. TNF receptor subtype signalling: differences and cellular consequences. Cell Signal 2002; 14: 477–92. Aggarwal BB, Samanta A, Feldmann M. Chapters on TNF and TNF receptors. In: Oppenheim JJ, Feldmann ME (Eds). The Cytokine Reference. London: Academic Press 2000; 413–34 and 1619–32. Chen G, Goeddel DV. TNF-R1 signaling: a beautiful pathway. Science 2002; 296: 1634–35. Baldwin AS. Control of oncogenesis and cancer therapy resistance by the transcription factor NF-kappaB. J Clin Invest 2001; 107: 241–46. Havell, EA, Fiers W, North RJ. The antitumor function of tumor necrosis factor (TNF), I. Therapeutic action of TNF against an established murine sarcoma is indirect, immunologically dependent, and limited by severe toxicity. J Exp Med 1988; 167: 1067–85. Baxevanis CN, Voutsas IF, Tsitsilonis OE, et al. Compromised antitumor responses in tumor necrosis factor-alpha knockout mice. Eur J Immunol 2000; 30: 1957–66. Moore RJ, Owens DM, Stamp G, et al. Mice deficient in tumor necrosis factor-alpha are resistant to skin carcinogenesis. Nat Med 1999; 5: 828–31. Suganuma M, Okabe S, Marino MW, et al. Essential role of tumor necrosis factor alpha (TNF-alpha) in tumor promotion as revealed by TNF-alpha-deficient mice. Cancer Res 1999; 59: 4516–18. Arnott CH, Scott KA, Moore RJ, et al. Tumour necrosis factor-alpha mediates tumour promotion via a PKC alpha- and AP-1-dependent pathway. Oncogene 2002; 21: 4728–38. Roberts RA, Kimber I. Cytokines in non-genotoxic hepatocarcinogenesis. Carcinogenesis 1999; 20: 1397–401. Knight B, Yeoh GC, Husk KL, et al. Impaired preneoplastic changes and liver tumor formation in tumor necrosis factor receptor type 1 knockout mice. J Exp Med 2000; 192: 1809–18. Kitakata H, Nemoto-Sasaki Y, Takahashi Y, et al. Essential roles of tumor necrosis factor receptor p55 in liver metastasis of intrasplenic administration of colon 26 cells. Cancer Res 2002; 62: 6682–87. Korner H, Cretney E, Wilhelm P, et al. Tumor necrosis factor sustains the generalized lymphoproliferative disorder (gld) phenotype. J Exp Med 2000; 191: 89–96. Orosz P, Echtenacher B, Falk W, et al. Enhancement of experimental metastasis by tumor necrosis factor. J Exp Med 1993; 177: 1391–98. Malik ST, Naylor MS, East N, et al. Cells secreting tumour necrosis factor show enhanced metastasis in nude mice. Eur J Cancer 1990; 26: 1031–34. Qin Z, Kruger-Krasagakes S, Kunzendorf U, et al. Expression of tumor necrosis factor by different tumor cell lines results either in tumor suppression or augmented metastasis. J Exp Med 1993; 178: 355–60. Komori A, Yatsunami J, Suganuma M, et al. Tumor necrosis factor acts as a tumor promoter in BALB/3T3 cell transformation. Cancer Res 1993; 53: 1982–85. Malik ST, Griffin DB, Fiers W, et al. Paradoxical effects of tumour necrosis factor in experimental ovarian cancer. Int J Cancer 1989; 44: 918–25. Oliff A, Defeo-Jones D, Boyer M, et al. Tumors secreting human TNF/cachectin induce cachexia in mice. Cell 1987; 50: 555–63. Sherry BA, Gelin J, Fong Y, et al. Anticachectin/tumor necrosis factor-alpha antibodies attenuate development of cachexia in tumor models. Faseb J 1989; 3: 1956–62.
572
32 Cahlin C, Korner A, Axelsson H, et al. Experimental cancer cachexia: the role of host-derived cytokines interleukin (IL)-6, IL12, interferon-gamma, and tumor necrosis factor alpha evaluated in gene knockout, tumor-bearing mice on C57 Bl background and eicosanoid-dependent cachexia. Cancer Res 2000; 60: 5488–93. 33 Yoneda T, Alsina MA, Chavez JB, et al. Evidence that tumor necrosis factor plays a pathogenetic role in the paraneoplastic syndromes of cachexia, hypercalcemia, and leukocytosis in a human tumor in nude mice. J Clin Invest 1991; 87: 977–85. 34 Naylor MS, Stamp GW, Foulkes WD, et al. Tumor necrosis factor and its receptors in human ovarian cancer: potential role in disease progression. J Clin Invest 1993; 91: 2194–206. 35 Naylor MS, Stamp GW, Balkwill FR. Investigation of cytokine gene expression in human colorectal cancer. Cancer Res 1990; 50: 4436–40. 36 Szlosarek PW, Grimshaw MJ, Wilbanks GD, et al. Tumor necrosis factor- regulation in normal and malignant ovarian epithelium. Proc Am Assoc Cancer Res 2003; 44: (abstr) R5053. 37 Mizokami A, Gotoh A, Yamada H, et al. Tumor necrosis factoralpha represses androgen sensitivity in the LNCaP prostate cancer cell line. J Urol 2000; 164: 800–05. 38 Vinante F, Rigo A, Tecchio C, et al. Serum levels of p55 and p75 soluble TNF receptors in adult acute leukaemia at diagnosis: correlation with clinical and biological features and outcome. Br J Haematol 1998; 102: 1025–34. 39 Warzocha K, Salles G, Bienvenu J, et al. Tumor necrosis factor ligand-receptor system can predict treatment outcome in lymphoma patients. J Clin Oncol 1997; 15: 499–508. 40 Foa R, Massaia M, Cardona S, et al. Production of tumor necrosis factor-alpha by B-cell chronic lymphocytic leukemia cells: a possible regulatory role of TNF in the progression of the disease. Blood 1990; 76: 393–400. 41 Herrmann F, Helfrich SG, Lindemann A, et al. Elevated circulating levels of tumor necrosis factor predict unresponsiveness to treatment with interferon alfa-2b in chronic myelogenous leukaemia. J Clin Oncol 1992; 10: 631–34. 42 Rossi A, Landolfo G, Sassi D, et al. Immunocytochemical increased evidence of inducible nitric oxide synthase, tumor necrosis factoralpha, and adrenocorticotropic hormone in human peritumoral lymph nodes. Appl Immunohistochem Mol Morphol 2002; 10: 52–56. 43 Tselepis C, Perry I, Dawson C, et al. Tumour necrosis factor-alpha in Barrett’s oesophagus: a potential novel mechanism of action. Oncogene 2002; 21: 6071–81. 44 Wong GH, Elwell JH, Oberley LW, et al. Manganous superoxide dismutase is essential for cellular resistance to cytotoxicity of tumor necrosis factor. Cell 1989; 58: 923–31. 45 Wu S, Boyer CM, Whitaker RS, et al. Tumor necrosis factor alpha as an autocrine and paracrine growth factor for ovarian cancer: monokine induction of tumor cell proliferation and tumor necrosis factor alpha expression. Cancer Res 1993; 53: 1939–44. 46 Gaiotti D, Chung J, Iglesias M, et al. Tumor necrosis factor-alpha promotes human papillomavirus (HPV) E6/E7 RNA expression and cyclin-dependent kinase activity in HPV-immortalized keratinocytes by a ras-dependent pathway. Mol Carcinog 2000; 27: 97–109. 47 Maeda M, Watanabe N, Okamoto T, et al. Endogenous tumor necrosis factor functions as a resistant factor against adriamycin. Int J Cancer 1994; 58: 376–79. 48 Jaiswal M, LaRusso NF, Burgart LJ, et al. Inflammatory cytokines induce DNA damage and inhibit DNA repair in cholangiocarcinoma cells by a nitric oxide-dependent mechanism. Cancer Res 2000; 60: 184–90. 49 Leibovich SJ, Polverini PJ, Shepard HM, et al. Macrophage-induced angiogenesis is mediated by tumour necrosis factor-alpha. Nature 1987; 329: 630–32. 50 Yoshida S, Ono M, Shono T, et al. Involvement of interleukin-8, vascular endothelial growth factor, and basic fibroblast growth factor in tumor necrosis factor alpha-dependent angiogenesis. Mol Cell Biol 1997; 17: 4015–23. 51 Cheng N, Chen J. Tumor necrosis factor-alpha induction of endothelial ephrin A1 expression is mediated by a p38 MAPK- and SAPK/JNK-dependent but nuclear factor-kappa B-independent mechanism. J Biol Chem 2001; 276: 13771–77. 52 De Cesaris P, Starace D, Starace G, et al. Activation of Jun Nterminal kinase/stress-activated protein kinase pathway by tumor necrosis factor alpha leads to intercellular adhesion molecule-1 expression. J Biol Chem 1999; 274: 28978–82. 53 Bussolino F, Camussi G, Baglioni C. Synthesis and release of
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
54
55
56 57 58 59 60
61 62
63
Review
platelet-activating factor by human vascular endothelial cells treated with tumor necrosis factor or interleukin 1 alpha. J Biol Chem 1988; 263: 11856–61. Leek RD, Landers R, Fox SB, et al. Association of tumour necrosis factor alpha and its receptors with thymidine phosphorylase expression in invasive breast carcinoma. Br J Cancer 1998; 77: 2246–51. Battegay EJ, Raines EW, Colbert T, et al. TNF-alpha stimulation of fibroblast proliferation: dependence on platelet-derived growth factor (PDGF) secretion and alteration of PDGF receptor expression. J Immunol 1995; 154: 6040–47. Leber TM, Balkwill FR. Regulation of monocyte MMP-9 production by TNF-alpha and a tumour-derived soluble factor (MMPSF). Br J Cancer 1998; 78: 724–32. Rosen EM, Goldberg ID, Liu D et al. Tumor necrosis factor stimulates epithelial tumor cell motility. Cancer Res 1991; 51: 5315–21. Tisdale MJ. Cachexia in cancer patients. Nat Rev Cancer 2002; 2: 862–71. Warzocha K, Ribeiro P, Bienvenu J, et al. Genetic polymorphisms in the tumor necrosis factor locus influence non-Hodgkin’s lymphoma outcome. Blood 1998; 91: 3574–81. Davies FE, Rollinson SJ, Rawstron AC, et al. High-producer haplotypes of tumor necrosis factor alpha and lymphotoxin alpha are associated with an increased risk of myeloma and have an improved progression-free survival after treatment. J Clin Oncol 2000; 18: 2843–51. Nakajima K, Sasaki M, Nojima D, et al. Tumor necrosis factoralpha gene mutations and genotype changes in renal cell carcinoma. J Urol 2001; 165: 612–15. Oh BR, Sasaki M, Perinchery G, et al. Frequent genotype changes at -308, and 488 regions of the tumor necrosis factor-alpha (TNFalpha) gene in patients with prostate cancer. J Urol 2000; 163: 1584–87. Hajeer AH, Lear JT, Ollier WE, et al. Preliminary evidence of an association of tumour necrosis factor microsatellites with increased risk of multiple basal cell carcinomas. Br J Dermatol 2000; 142: 441–45.
THE LANCET Oncology Vol 4 September 2003
64 Ghaderi M, Nikitina L, Peacock CS, et al. Tumor necrosis factor a11 and DR15-DQ6 (B*0602) haplotype increase the risk for cervical intraepithelial neoplasia in human papillomavirus 16 seropositive women in Northern Sweden. Cancer Epidemiol Biomarkers Prev 2000; 9: 1067–70. 65 Bienvenu J, Chvetzoff R, Salles G et al. Tumor necrosis factor alpha release is a major biological event associated with rituximab treatment. J Haematol 2001; 2: 378–84. 66 Ramesh G, Reeves WB. TNF-alpha mediates chemokine and cytokine expression and renal injury in cisplatin nephrotoxicity. J Clin Invest 2002; 110: 835–42. 67 Piquet PF, Collart MA, Grau GE et al. Tumor necrosis factor/cachectin plays a key role in bleomycin-induced pneumopathy and fibrosis. J Exp Med 1989; 170: 655–63. 68 Libura J, Bettens F, Radkowski A, et al. Risk of chemotherapyinduced pulmonary fibrosis is associated with polymorphic tumour necrosis factor-a2 gene. Eur Respir J 2002; 19: 912–18. 69 Feldmann M. Development of anti-TNF therapy for rheumatoid arthritis. Nat Rev Immunol 2002; 2: 364–71. 70 Forster MD, Braybrooke J, Madhusudan S, et al. A Phase II trial using etanercept, a recombinant tumour necrosis factor anatgonist, in metastatic breast cancer. Br J Cancer 2002; 86 (suppl): 15 71 Vassallo R, Matteson E, Thomas CF Jr. Clinical response of rheumatoid arthritis-associated pulmonary fibrosis to tumor necrosis factor-alpha inhibition. Chest 2002; 122: 1093–96. 72 Richardson P, Hideshima T, Anderson K. Thalidomide: emerging role in cancer medicine. Annu Rev Med 2002; 53: 629–57. 73 Turini ME, DuBois RN. Cyclooxygenase-2: a therapeutic target. Annu Rev Med 2002; 53: 35–57. 74 Plummer SM, Holloway KA, Manson MM, et al. Inhibition of cyclo-oxygenase 2 expression in colon cells by the chemopreventative agent curcumin involves inhibition of NFkappaB activation via the NIK/IKK signalling complex. Oncogene 1999; 18: 6013–20. 75 Sueoka N, Suganuma M, Sueoka E, et al. A new function of green tea: prevention of lifestyle-related diseases. Ann N Y Acad Sci 2001; 928: 274–80.
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
573
Review
Tumour necrosis factor : a potential target for the therapy of solid tumours Peter W Szlosarek and Frances R Balkwill
Lancet Oncol 2003; 4: 565–73
Tumour necrosis factor (TNF) is a vital cytokine involved in inflammation, immunity, and cellular organisation.1 It was first isolated from the serum of mice infected with Bacillus Calmette-Guérin treated with endotoxin, and was shown to replicate the ability of endotoxin to induce haemorrhagic tumour necrosis.2 This discovery ended a long search for an important component of “Coley’s mixed toxins”, a crude bacterial filtrate developed by Willam B Coley, a New York surgeon and pioneer of an early systemic cancer treatment at the turn of the 20th century. Coley’s mixed toxins consisted of a filtrate from cultures of Streptococcus pyogenes and Serratia marcescens and induced high fever and tumour necrosis in patients who responded, particularly those with sarcoma, but also patients with carcinoma and lymphoma.3 The phenomenon of tumour necrosis is now attributed, in part, to a vascular thrombotic mechanism, mediated by a surge of macrophage-derived TNF, released after activation by the endotoxin contained within Coley’s toxins. In the 1980s, TNF was also characterised as cachectin4 for its role in wasting—first indentified in rabbits infected with Trypanasoma brucei—and as T-cell differentiation factor.5 Cloning of the TNF gene in 1984 led to a decade of clinical experimentation, culminating in a licence from the European Medicine Evaluation Agency for its use as a locoregional treatment for inoperable sarcoma.6 However, systemic TNF therapy proved ineffective with a lack of objective tumour response and serious side-effects (hypotension and organ THE LANCET Oncology Vol 4 September 2003
Rights were not granted to include this image in electronic media. Please refer to the printed journal. © Wellcome Photo Library/T Blundell and N Campillo
Tumour necrosis factor (TNF), named for its antitumour properties, was isolated almost 30 years ago. It is a vital member of the multifunctional TNF superfamily and has important roles in immunity and cellular remodelling as well as influencing apoptosis and cell survival. Its central role in inflammation has led to the development of TNF antagonists as effective therapies for rheumatoid arthritis and inflammatory bowel disease. In this review, we discuss the evidence which has accumulated during the past decade that implicates TNF in inflammatory pathways that increase tumorigenesis. There is convincing evidence that under specific conditions TNF is a tumour promoter and helps to produce the toxic effects associated with conventional cancer therapy, such as the cytokine release syndrome and cisplatin-induced nephrotoxicity. Several trials have been set up to investigate the role of TNF antagonists in cancer. It is hoped that these agents inhibit the neoplastic process either alone or in combination with other agents, and ameliorate the side effects of cancer therapy.
Figure 1. Ribbon structure of tumour necrosis factor.
failure), which redirected researchers towards alternative forms of local delivery.7 Interest in the use of TNF mutants and heat-shock proteins to increase the therapeutic window of TNF therapy remains experimental. In parallel with the emerging clinical data for TNF therapy in malignant disease, increasing evidence attributed a role for TNF in rheumatoid arthritis, inflammatory bowel disease, diabetes, sepsis, and several infections, including HIV.1 Several studies have implicated endogenous TNF in tumour-cell growth and stromal interactions that facilitate tumour invasion and metastasis. Although TNF is capable of initiating a tumour apoptotic response (with or without an immune-mediated mechanism) the reality is that these pathways are frequently deactivated within tumour cells (figure 2). In some circumstances, TNF can provide a survival signal for the cancer cell and hence it has been referred to as a tumour promoting factor.8 Furthermore, the concept of cancer as a purely genetic disease has been re-evaluated and the importance of the tumour microenvironment and inflammation in cancer progression PWS is a Clinical Research Fellow funded by Cancer Research UK and FRB is Professor of Cancer Biology at Barts and The London and Co-Director of the Translational Oncology Laboratory, Cancer Research UK, London, UK. Correspondence: Dr Frances Balkwill, Cancer Research UK, Translational Oncology Laboratory, Barts and The London, Queen Mary’s School of Medicine and Dentistry, John Vane Science Centre, Charterhouse Square, London EC1M 6BQ, UK. Tel: +44 (0)20 7882 6106. Fax: +44 (0)20 7882 6110. Email: [email protected]
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
565
Tumour necrosis factor
Review has been emphasised.9,10 As a central mediator of inflammation, TNF provides a molecular link between chronic inflammatory stimuli and the subsequent development of malignant disease. Its paradoxical role in cancer progression has stimulated interest in the use of TNF antagonists for the prevention and treatment of cancer. The use of TNF antagonists in haematological cancers has recently been discussed, providing a rationale for their clinical use.11 In this review, we concentrate on the case for modulating TNF in solid tumours.
TNF and TNF receptor signalling TNF is a soluble 17 kDa molecule (157 aminoacids) that binds as a homotrimer to two distinct homotrimeric receptors on the cell surface: TNFRI (p55 receptor) and TNFRII (p75 receptor).12,13 TNF is synthesised as a 26 kDa (233 aminoacids) membrane-bound propeptide (proTNF) and is secreted after cleavage by TNF-converting enzyme (TACE). The 26 kDa form is also functional and binds to TNFRII via direct cell-to-cell contact. Although activated macrophages are a major source of TNF, it can be made by various other cells including fibroblasts, astrocytes, Kupffer cells, smooth-muscle cells, keratinocytes, and tumour cells. The promoter region of the TNF gene contains binding sites for several transcription factors, including NFB, cyclic AMP response element (CRE), activator proteins (AP) 1 and 2, and nuclear-factor-activated T cells. Post-transcriptional regulation of TNF gene expression occurs via an AU-rich element (ARE) within the 3' untranslated region, which interferes with mRNA stability and translation efficiency. For example, deletion of ARE from the genomic TNF locus causes basal concentrations of TNF to increase, producing an inflammatory phenotype in mice (rheumatoid arthritis and inflammatory bowel disease). A similar disorder is observed in mice deficient in tristetraprolin, an endogenous RNA-binding protein that regulates TNF mRNA stability. Lastly TACE, a member of the ADAM family of metalloproteinases, releases the soluble mature form of TNF and its receptors via a protein kinase C and metalloproteinase-dependent signalling pathway. Processed TNFRs provide additional negative regulation in the form of soluble TNF binding proteins. The signalling cascade downstream of TNFRI and TNFRII is complex and involves many adaptor proteins, which are recruited after binding of the TNF ligand to its receptor. Figure 2 summarises the main elements of the TNFR signalling pathway, in particular pathways implicated in inflammation and carcinogenesis. TNFRI is ubiquitously expressed on mammalian cells and is therefore involved in various biological effects after treatment with TNF including: regulation of apoptosis, cell proliferation, differentiation, immunity, and inflammation. Recruitment of TNF to TNFRI results in dissociation of the silencer of death domain protein (SODD), whose function is to repress TNFRI signalling by blocking the binding of adaptor proteins to the death domain of TNFRI. Release of SODD permits the assembly of an activated signalling complex at the death domain of TNFRI, consisting of TNFR-associated death domain (TRADD), TNFR-
566
associated factor 2 (TRAF2), receptor-interacting protein (RIP), and FAS-associated death domain (FADD). These molecules regulate at least four distinct pathways: a proapoptotic pathway that is induced by binding caspase 8 to FADD; an antiapoptotic programme that is activated by the binding of cellular inhibitor of apoptosis protein 1 (cIAP-1) to TRAF2; AP1 activation mediated through TRAF2 via a JNK-dependent kinase cascade; and NFB activation by RIP.14 The regulation of NFB by TNF is an example of a signalling pathway of fundamental importance in carcinogenesis.15 NFB activation requires phosphorylationdependent degradation of inhibitor of B (IB) proteins, which sequester NFB in the cytoplasm. This step is mediated via a multiprotein IB kinase (IKK) complex consisting of two catalytic subunits (IKK and IKK) and a regulatory subunit, IKK, which is in turn activated by RIP. Through this pathway, TNF targets IB to the proteosome for ubiquitination and stimulates translocation of NFB to the nucleus. Although elucidation of TNFRI signalling has in recent years progressed significantly, the factors controlling outcome in a particular cell (eg, the dominance of NFB over caspase activation in a specific cell) are poorly understood. TNFRII signalling is largely confined to endothelial and haematopoietic cells and not much is known about its structure and function. However, pro-TNF, in particular, has a higher affinity for TNFRII and faster dissociation kinetics from this receptor than soluble TNF. There is also evidence of reverse signalling, which involves binding of pro-TNF to TNFRII to initiate retrograde signal transduction. Despite the lack of a death domain, and the recruitment of fewer adaptor molecules than TNRI, TNFRII is able to affect NFB and JNK signalling.
TNF and experimental cancers TNF knockout mice (TNF-/-) have provided evidence of a tumour immunosurveillance role for TNF16,17 and implicated the cytokine in tumour development and metastasis.18,19 Moore and colleagues found that TNF-/mice had ten times fewer skin tumours than wild-type mice after initiation with a carcinogen and repeated application of 12-O-tetradecanoylphorbol-13-acetate (TPA). Although TNF was important for de-novo carcinogenesis, the later stages of tumour progression were similar in TNF-/- and wild-type mice. The resistance of TNF-/- mice to skin carcinogenesis is likely to be related to a temporal delay in the activation of protein kinase C and AP1, which are important TPA-responsive signalling molecules.20 This delay seems to affect the expression of genes involved in tumour development, eg, granulocyte-macrophage colonystimulating factor and matrix metalloproteinases 3 and 9, which are suppressed in the TNF-/- mice but not in wildtype mice exposed to carcinogen. TNFRI-/- mice, and to a lesser extent TNFRII-/- mice, also show resistance to skin carcinogenesis (unpublished data). The tumour-promoting role of TNF was also investigated in models of hepatic carcinogenesis,21 in which TNFRI-/- and TNFRII-/- mice have proven useful for elucidating the mechanism of action involved.22 TNFRI-/-
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
Review
mice had reduced oval cell (hepatic stem cell) proliferation during the preneoplastic phase of liver carcinogenesis, which was associated with fewer tumours. Liver tumorigenesis was unaffected in TNFRII-/- mice. Kitakata and co-workers used TNFRI-/- mice to show that endogenous TNF is vital for promoting liver metastasis after intrasplenic administration of a colonic adenocarcinoma cell line.23 Although there was no difference in the size of the splenic tumours, more than 90% of wild-type mice developed liver metastases compared with less than 50% of TNFRI-/- mice. Furthermore, TNFRI-/- mice had smaller intrahepatic metastases. In wildtype mice, TNF (mRNA and protein) was induced around the hepatic central and portal veins by day 3 and in hepatic metastases after 24 days. The difference in metastatic behaviour was accounted for by increased VCAM1 mRNA and protein expression in the livers of wild-type animals, which was significantly attenuated in TNFRI-/- mice. Lastly, the fatal lymphoproliferative disorder that develops in FASL-deficient mice was attenuated by crossing these animals with TNF-/- mice.24 TNF may have induced the expression of chemokines (chemotactic cytokines), which regulate trafficking and accumulation of tumour cells into lymph nodes.24 Transgenic mice overexpressing TNF are unsuitable for studying the role of TNF in tumorigenesis because they are affected by early and severe inflammatory pathologies, such as pulmonary hypertension, cardiac failure, neurodegeneration, and arthritis.13 Nevertheless, several animal experiments that manipulate concentrations of TNF within the tumour microenvironment have confirmed that an increase in TNF promotes cancer development and spread. Orosz and colleagues showed increased lung metastases from
an experimental fibrosarcoma after pretreatment of the animals with TNF.25 Alternatively, lung metastases were reduced by the addition of an anti-TNF antibody, which neutralised endogenous levels of TNF. Malik and colleagues reported that overexpression of TNF conferred invasive properties on xenograft tumours.26 However, the effect is clearly dependent on tumour type because the TNF transgene could either induce tumour rejection or decrease survival, for example, by promoting hepatic metastases.27 Monoclonal antibodies to TNF reversed the hepatic metastases and increased survival. Further evidence for a tumour-promoting role of TNF has been provided by chemical carcinogenesis experiments with BALB/3T3 fibroblasts. BALB/3T3 cells in nude mice that were treated with a chemical carcinogen and then exposed for 2 weeks to TNF underwent transformation and yielded tumours.28 Human xenograft studies of ovarian cancer in nude mice have also emphasised the paradoxical effects of TNF with conversion of ascites into peritoneal deposits.29 TNF and cachexia
Studies of cancer cachexia in mice have confirmed that TNF is an important mediator of the phenomenon— alongside interleukins 1 and 6 and interferon .30,31 A study of TNFRI-/- and TNFRII-/- mice assessed the contribution of tumour and host TNF to cancer cachexia. Cachexia was not suppressed in the TNFR knockout mice, suggesting that tumour-derived TNF is more important than host TNF.32 Human xenograft data supports a role for TNF in mediating cachexia and other paraneoplastic features, such as hypercalcaemia and leucocytosis.33 These features could all be reversed with anti-TNF antibodies.
TNF as a tumour promoter TNF bound to cell membrane
Soluble TNFR
TACE TNF
Pro-TNF
TNFRI
TACE
Cell membrane
TNFRII
SODD TRADD cIAPS
FADD
Caspase 8
RIP
TRAF2
MAPKK
IKK complex NFB-kB(cytostolic)
JNK/p38
APOPTOSIS
AP1
NFB(nuclear)
Proliferation Cancer cell survival Inflammation
Figure 2. TNF binds to TNFRI and TNFRII activating several pathways associated with inflammation, apoptosis, and cell survival.
THE LANCET Oncology Vol 4 September 2003
Evidence for a role of TNF has come from several preclinical and clinical studies. Expression studies have confirmed abnormally high concentrations of TNF in tumours, implicating the cytokine in specific preneoplastic lesions and in advanced malignant disease. Data from studies in vitro and in vivo suggests that TNF, far from being an epiphenomenon, is an active player in tumorigenesis. Furthermore, several well-known paraneoplastic syndromes that commonly occur in patients with cancer seem to be caused by secreted tumour/host factors, including TNF. Finally, several single nucleotide polymorphisms (SNPs) that may influence the extent of TNF expression are associated with an increased risk of cancer. TNF in the tumour microenvironment
In-situ hybridisation and immunohistochemical studies have confirmed
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
567
Tumour necrosis factor
Review tumour and stromal expression of TNF in breast, ovarian, colorectal, prostate, bladder, oesophageal, and renal-cell cancers, and in melanoma, lymphoma, and leukaemia. The extent of TNF expression varies between tumours, as does its location, and the accompanying cytokine profile. For example, in ovarian cancer TNF is expressed mainly within tumour epithelial islands (70% of assessed tumours)34 whereas predominant expression in colorectal cancer is within tumour-associated macrophages.35 In ovarian cancer, TNFRI is found on tumour and stromal cells and TNFRII localises to the leukocyte infiltrate, suggesting potential for both autocrine and paracrine actions of endogenous TNF. Moreover, TNF activity seems to be deregulated with inappropriate constitutive expression in ovarian tumour epithelium compared with normal ovarian surface epithelium.36 Studies of serous ovarian carcinoma, prostate carcinoma, non-Hodgkin lymphoma, acute leukaemia, chronic myelogenous leukaemia, and chronic lymphocytic leukaemia have found an association between expression of the cytokine, poor survival, and resistance to therapy.34,37–41 By use of immunohistochemical analysis, researchers found overexpression of TNF, inducible nitric oxide synthetase (iNOS) and adrenocorticotrophic hormone peritumoral lymph nodes and not in mesenteric (control) lymph nodes (the two are morphologically identical) from patients with colorectal cancer,42 suggesting a role for these molecules in early lymphatic spread. Recent data from a study of gastrooesophageal cancer showed progression in TNF and TNFRI epithelial expression along the Barrett’s metaplasia-dysplasiacarcinoma sequence (73% of tumours; figure 3).43 Tselepsis and colleagues reported that TNF mediated upregulation of the c-MYC oncogene, via -catenin and a MAPK (p38 and ERK) dependent pathway in CACO2, a cell line which differentiates into small-intestinal-like enterocytes in vitro and simulates Barrett’s metaplasia. Moreover, E cadherin, an important cell-adhesion molecule is negatively regulated by TNF and in common with several other epithelial cancers, is downregulated in Barrett’s adenocarcinoma, with corresponding increases in cytoplasmic and nuclear pools of -catenin. Mechanism of tumour promotion
TNF is rarely cytotoxic to tumour cells in vitro unless an additional stressor (eg, a RNA or protein synthesis inhibitor) is provided. Tumour cells are often resistant to the cytokine via a mechanism involving manganese superoxide dismutase, which scavenges oxygen free radicals induced by TNF.44 The various mechanisms by which TNF promotes cancer growth, invasion, and metastasis are shown in the panel. TNF acts as a growth factor in certain tumour types, increasing concentrations of positive cell-cycle regulators (and decreasing levels of CDK inhibitors) and components of growth-factor-receptor signalling pathways such as RAS or c-MYC.43,45,46 Other direct roles of endogenous TNF include inducing chemoresistance in several cancers47 and mediating androgen independence in prostate cancer.37 TNF also promotes DNA damage and inhibits DNA repair by upregulating nitric oxide (NO) dependent (non-cGMP) pathways.48 Moreover, both argininosuccinate synthetase (the rate-limiting enzyme in NO synthesis) and iNOS are
568
Tumour-promoting effects of TNF l
Production of NO (DNA/enzyme damage, cGMP-mediated tumour promotion)
l
Autocrine growth and survival factor for malignant cells
l
Activation of E6/E7 mRNA in HPV-infected cells
l
Tissue remodelling via induction of matrix metalloproteinases
l
Control of leukocyte infiltration in tumours via modulation of chemokines and their receptors
l
Downregulation of E cadherin, increased nuclear pool of catenin
l
Increase tumour-cell motility and invasion
l
Induction of angiogenic factors
l
Loss of androgen responsiveness
l
Resistance to cytotoxic drugs
induced by various cytokines in tumour cell lines, with increased iNOS present in various solid tumours, including gynaecological cancers and breast cancer. Low concentrations of NO are implicated in many aspects of tumorigenesis. The tumour stroma is modulated in several ways by TNF. Although TNF can induce collapse of tumour vasculature via downregulation of V3 signalling and an increase in angiostatin,7 the cytokine can also promote angiogenesis.49 Various factors mediate the proangiogenic effects of TNF including VEGF and its receptor VEGFR2, bFGF, interleukin 8, platelet activating factor, ephrin A (a ligand for the Eph family of receptor tyrosine kinases), NO, E selectin, ICAM1, and thymidine phosphorylase.50–54 The paradoxical effects of TNF on tumour vasculature may reflect the difference in chronic synthesis (which favours angiogenesis) and acute high-dose local administration (which triggers vascular thrombosis). TNF promotes further tumour remodelling by stimulating fibroblast activity, tumour-cell motility, and tumour invasion via the induction of matrix metalloproteinases.55–57 Paraneoplastic phenomena
An increasing tumour burden is characterised by progressive weight loss with muscle and lipid wasting known as cachexia. This syndrome is driven by several tumour and host factors including cytokines, such as TNF, interleukins 1 and 6, and interferon . Although data from animal studies of the involvement of TNF is convincing, clinical studies evaluating the cytokine in human cachexia have been less consistent.58 However, the data for specific tumours, for example pancreatic carcinoma, is more robust and recent studies have confirmed increased TNF (mRNA and protein) concentrations in blood from patients with cachexia. Other paraneoplastic features have been linked to TNF including fever, hypercalcaemia, anaemia, and fatigue typical of many patients with advanced disease. TNF has been characterised as an endogenous pyrogen, osteoclast activating factor facilitating skeletal metastases, and as a suppressor of erythropoietin production.13 SNPs and susceptibility to cancer
If TNF is an endogenous tumour promoter, it is possible that functional SNPs in the genes for TNF, TNFRs, or
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
Review Treatment-related complications
B
A
Courtesy of Janusz Jankowski
Several proinflammatory cytokines, such as TNF and interleukin 6, mediate cytokine release syndromes, which complicate antibody-based cancer therapy. For example, several studies have shown frequent first-dose infusion-related toxic effects with rituximab (a monoclonal antibody to the CD20 cell-surface antigen) in lowD C grade non-Hodgkin lymphoma— symptoms consist mainly of fever, rigors, and chills. Studies of rituximab in patients with haematological cancer have confirmed TNF release as a major biological event, correlating with these clinical features.65 More severe reactions of dyspnoea, Figure 3. Immunolocalisation of TNF in Barrett's oesophagus (TNF antibody, DAB and hypotension, abdominal pain, and haematoxylin counterstain). TNF expression increases with progression from normal squamous vomiting are less common with epithelium (weak staining in basal layers and submucosa only) along the Barrett's metaplasiarituximab (less than 10%) and have dysplasia-adenocarcinoma sequence. A benign gland entrapped within the tumour (arrow) with no also been reported in solid tumour TNF expression, contrasts with strong expression in the adenocarcinoma cells. therapy with a similar patterns of events downstream in the TNF pathway may be TNF production. The mechanism downstream of TNF is associated with cancer severity and susceptibility. Several unclear, but is thought to involve the release of NO and studies of small numbers of patients have reported leukostasis in pulmonary vasculature. associations between several TNF SNPs in various Ramesh and Reeves showed a major role for TNF in cancers. For example, several studies have linked SNPs in mediating cisplatin-induced nephrotoxicity in mice.66 the promoter region of TNF and the related cytokine Although several proinflammatory cytokines and chemolymphotoxin to cancer susceptibility and severity. In a kines are upregulated during cisplatin-induced injury, study of 273 patients with lymphoma, the TNF308 including TGF, RANTES, MIP2, and MCP1, the protection polymorphism was associated with high plasma afforded by TNF antagonists or deletion of the TNF gene, concentrations of the TNF at first presentation. shows the absolute requirement for TNF. The cytokine Furthermore, possession of at least two TNF/ initiates nephrotoxicity by two mechanisms: a direct lymphotoxin high producer alleles was a risk factor for apoptotic effect on renal epithelial cells (via caspase 8); and first-line treatment failure, shorter progression-free by regulating a cytokine network, which promotes a large survival, and overall survival.59 In addition, individuals influx of leukocytes within the kidney cortex. The latter with polymorphisms associated with a high production of produces oxidant stress causing further damage and release TNF or lymphotoxin are also at significantly increased of TNF. Bleomycin-induced pulmonary fibrosis is another risk of developing monoclonal gammopathy of uncertain example of a toxic effect that is linked to proinflammatory significance and myeloma.60 Associations have also been cytokines and an alveolar leukocyte reaction.67 Moreover, the found between SNPs in the promoter region of TNF and polymorphic TNF2 microsatellite, an allele in the susceptibility to renal and prostate cancer. Patients with promoter region of the TNF gene, is associated with an genotype GA at locus -238 had a 6·5 times higher risk of increased risk of bleomycin-induced lung injury.68 renal cancer and a GA genotype at locus 488 was associated with a three times higher risk. The incidence of prostate TNF antagonists cancer with genotype GA at locus 488 was 17 times Specific TNF antagonists, such as infliximab and etanercept, are now accepted as successful treatments for higher.61,62 There are also two associations between TNF various inflammatory and autoimmune conditions.69 In light microsatellite polymorphisms and human cancer. In of the evidence outlined in this review, and the similarities in cutaneous basal-cell carcinoma, individuals with the TNF1 inflammatory pathways in cancer and rheumatoid arthritis and 7 containing genotypes showed an increased (figure 4), a case can be made for the use of TNF susceptibility to the disease.63 Those with cutaneous basal- antagonists in solid tumours. The behaviour of the cell carcinoma with TNF alleles d4 and d6 and the TNF2- inflammatory pannus of rheumatoid arthritis may be b4-d5 haplotype had more tumours. Secondly, a relative risk likened to the invasion and metastasis of cancer, with the of 15 for cervical intraepithelial neoplasia was conferred by upregulation of angiogenic factors, matrix metalloproteases, the combination of TNF11, HLA-DQ, and HPV16 cytokines, and chemokines. In addition, studies in patients positivity.64 with rheumatoid arthritis have shown that infliximab THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
569
Tumour necrosis factor
Review
RA autoimmune trigger
Tumour initiation RhF Trigger for TNF
Helper T cell Bystander activated T cell
B cell +
Trigger for TNF Transformed cell
TNF
Macrophage
+
+ TNF
+
Endothelial cell Macrophage
Endothelial cell
Fibroblast
Extracellular matrix Fibroblast
Chondrocyte Osteoclast
Erosion of bone and joint cartilage
Cell proliferation/survival Angiogenesis Leukocyte infiltration Matrix metalloproteinases Cytokine deregulation
Cancer progression
Figure 4. TNF pathways common to rheumatoid arthritis (RA) and tumourigenesis. Both RA and certain epithelial cancers (eg, ovarian cancer) are characterised by TNF-dependent processes such as cellular proliferation, angiogenesis, leukocyte inflitation, and the upregulation of matrix metalloproteases.
treatment reduces production of inflammatory cytokines, angiogenic factors, and matrix metalloproteinases, inhibits leukocyte trafficking to inflammed joints, and improves bone-marrow function.69 All these actions could be useful in a biological cancer therapy. TNF antagonists used to treat haematological cancers have confirmed previous safety data. Reports of activity are beginning to emerge, with improvements in bone-marrow function in myelodysplastic syndrome, myelofibrosis, and acute myelogenous leukaemia, and indications of a significant role in acute and chronic graft versus host disease.11 Several studies of TNF antagonists alone or in combination with chemotherapy have now begun in various solid tumours, and also for the palliative treatment of cancer cachexia. Preliminary reports of patients with breast cancer treated with TNF antagonists alone suggest biological activity with no significant toxic effects as determined by a whole blood cytokine release assay for MCP1 and interleukin 6, but no evidence of clinical response.70 A phase II trial of etanercept in patients with advanced ovarian cancer is currently underway assessing various cytokine parameters and CA125 as surrogate endpoints. Other studies of TNF antagonists are in progress in patients with non-small-cell
570
lung cancer (in combination with docetaxel) and pancreatic cancer (in combination with gemcitabine). There are unpublished case reports of infliximab with activity in patients with breast cancer and bone metastases as assessed by a reduction in pain symptoms and improvement on bone scintigraphy (DeWitte M, personal communication). A recent report of infliximab improving pulmonary fibrosis in a patient with rheumatoid arthritis would also be worth pursuing in the context of bleomycin-induced lung injury.71 Infliximab
Infliximab is a humanised monoclonal antibody (murine Fv, human IgG1) with both direct anti-TNF effects and the potential for antibody-dependent cellular cytotoxicity. It is licensed for use in rheumatoid arthritis and Crohn’s disease, with experimental use in various other inflammatory conditions, including juvenile Still’s disease, psoriatic arthropathy, and Behçet’s disease. The antibody is usually given at doses of 3–5 mg/kg every few weeks intravenously (in rheumatoid arthritis and irritable bowel disease). There is an increased risk of infection (particularly tuberculosis) and of certain autoimmune conditions (eg, exacerbation of demyelinating disease and a lupus-like syndrome with
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
Review
Search strategy and selection criteria A search of PubMed was carried out using combinations of the terms “TNF-alpha”, “TNFRI”, “TNFRII”, “knockout”, “tumour”, “carcinogenesis”, “inflammation”, “cachexia”, and “cytokine release syndrome”. Additional information on infliximab, etanercept and rituximab was obtained from the Electronic Medicines Compendium (http://emc.vhn.net/) and from investigators involved in clinical trials of these agents.
autoantibodies to double-stranded DNA), in which TNF has an important immunoregulatory role. This effect is consistent with data from TNF knockout mice. Because of infliximab’s slow elimination from the body and the potential for infective and autoimmune complications close monitoring is recommended for 6 months after cessation of therapy. Antibodies to infliximab have been detected in patients experiencing infusion-related reactions but these reactions lessen with repeated administration. Over 100 000 patients have been treated to date with infliximab and the observed incidence of malignant disease has been similar to those expected for the populations studied. Several other antibodies targeting TNF have recently become available: CDP571, a humanised murine IgG4 monoclonal antibody and D2E7, a human IgG1 monoclonal antibody. Etanercept
Etanercept, a dimeric molecule consisting of two molecules of the extracellular portion of the human TNFRII fused to the Fc portion of human IgG, is synthesised using recombinant DNA technology in the Chinese hamster ovary mammalian cell expression system. The drug is given in twice weekly subcutaneous injections (25 mg adult dose); it has a half life of 70 h and is renally excreted. Although antibodies to etanercept are seen, these are non-neutralising and have no effect on clinical outcome. Etanercept is currently licensed for use in rheumatoid arthritis but lacks efficacy in irritable bowel disease. Both drugs are associated with similar adverse events and contraindications. A pegylated truncated form of TNFRI is an another example of a TNF antagonist under clinical development. Non-specific TNF inhibitors
Thalidomide, dexamethasone, aspirin, and non-steroidal anti-inflammatory drugs (NSAIDs) are all non-specific TNF inhibitors, which modulate TNF-dependent and independent pathways. Dexamethasone, a potent antiinflammatory and immunosuppressive drug and one of the first non-specific TNF inhibitors clinically available, has widespread application in oncology. It inhibits TNF transcription and translation. Thalidomide, with more subtle immunomodulatory actions, also represses TNF in several ways, promoting decay of the mRNA transcript and decreasing NFB binding activity.72 Several studies have confirmed that thalidomide has activity in a range of tumours, including multiple myeloma, renal cell and prostate carcinoma, melanoma, glioma, and Kaposi’s sarcoma. Patients have also reported improvements in symptoms of anorexia, weight loss, nausea, insomnia, and profuse sweating in the palliative-care setting. THE LANCET Oncology Vol 4 September 2003
There is good epidemiological evidence for a protective role of aspirin and NSAIDs against colorectal cancer, and emerging data suggest a benefit in several other epithelial tumours.73 Both groups of anti-inflammatory drugs inhibit cyclo-oxygenase 2 (COX2), which is induced by TNF and other inflammatory cytokines via a NFB pathway, and is implicated in tumorigenesis. A recent randomised doubleblind study in patients with familial adenomatous polyposis of celecoxib, a specific COX2 antagonist, showed a reduction in colorectal polyps. Several studies of aspirin and other COX2 inhibitors for chemoprevention and treatment of cancer are currently underway. Interest has increased in other agents with anti-TNF activity, namely curcumin (a constituent of turmeric) and green tea, which may yet prove to have important roles in cancer chemoprevention.74,75
Conclusion TNF has a paradoxical role in cancer, inducing destruction of blood vessels and cell-mediated killing of certain tumours, as well as acting as a tumour promoter. Chronic endogenous low production of TNF favours the latter. TNF may contribute to the development of tissue architecture necessary for tumour growth and metastasis, may induce other cytokines, angiogenic factors, and matrix metalloproteinases, contributing to DNA damage and increase growth and survival of tumour cells. Thus, TNF provides a molecular bridge between inflammation and cancer. On the basis of the success of TNF antagonists in inflammatory disease, further investigation of these therapies is warranted to combat cancer, complications of cancer, and the adverse events of conventional treatment. The study of TNF has come a long way from the early days of William Coley and we can now manipulate concentrations within tumours by using supraphysiological concentrations or antagonising endogenously secreted TNF. Only time will tell whether TNF is a useful target for cancer therapy or whether tumour promoting factor would be a more appropriate name. Conflicts of interest
FRB is a consultant for Centocor Inc, manufacturers of infliximab, who also provide funding for research in the Translational Oncology Lab (Cancer Research UK) into preclinical studies of TNF antagonists in cancer models. All the published work on TNF and cancer done in our laboratory was carried out before industry collaboration. Acknowledgments
We thank Jannusz Janowski and Fionnula Brennan for assistance with figures 3 and 4, respectively. References
1 Locksley RM, Killeen N, Lenardo MJ, et al. The TNF and TNF receptor superfamilies: integrating mammalian biology. Cell 2001; 104: 487–501. 2 Carswell EA, Old LJ, Kassel RL, et al. An endotoxin-induced serum factor that causes necrosis of tumors. Proc Natl Acad Sci USA 1975; 72: 3666–70. 3 Nauts HC, Fowler GA, Bogatko FH. A review of the influence of bacterial infection and of bacterial products (Coley’s toxins) on malignant tumors in man. Acta Medica Scandinavia 1953; 145: S276. 4 Beutler B, Greenwald D, Hulmes JD, et al. Identity of tumour necrosis factor and the macrophage-secreted factor cachectin. Nature 1985; 316: 552–54. 5 Takeda K, Iwamoto S, Sugimoto H, et al. Identity of differentiation
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
571
Tumour necrosis factor
Review
6 7
8 9 10 11 12 13
14 15 16
17 18 19 20 21 22 23 24 25 26 27
28 29 30 31
inducing factor and tumour necrosis factor. Nature 1986; 323: 338–40. Lejeune FJ, Ruegg C, Lienard D. Clinical applications of TNF-alpha in cancer. Curr Opin Immunol 1998; 10: 573–80. Weichselbaum RR, Kufe DW, Hellman S, et al. Radiation-induced tumour necrosis factor- expression: clinical application of transcriptional and physical targeting of gene therapy. Lancet Oncol 2002; 3: 665–71. Balkwill F. Tumor necrosis factor or tumor promoting factor? Cytokine Growth Factor Rev 2002; 13: 135–41. Hanahan D, Weinberg RA, The hallmarks of cancer. Cell 2000; 100: 57–70. Balkwill F, Mantovani A. Inflammation and cancer: back to Virchow? Lancet 2001; 357: 539–45. Tsimberidou AM, Giles FJ. TNF-alpha targeted therapeutic approaches in patients with hematologic malignancies. Expert Rev Anticancer Ther 2002; 2: 277–86. MacEwan DJ. TNF receptor subtype signalling: differences and cellular consequences. Cell Signal 2002; 14: 477–92. Aggarwal BB, Samanta A, Feldmann M. Chapters on TNF and TNF receptors. In: Oppenheim JJ, Feldmann ME (Eds). The Cytokine Reference. London: Academic Press 2000; 413–34 and 1619–32. Chen G, Goeddel DV. TNF-R1 signaling: a beautiful pathway. Science 2002; 296: 1634–35. Baldwin AS. Control of oncogenesis and cancer therapy resistance by the transcription factor NF-kappaB. J Clin Invest 2001; 107: 241–46. Havell, EA, Fiers W, North RJ. The antitumor function of tumor necrosis factor (TNF), I. Therapeutic action of TNF against an established murine sarcoma is indirect, immunologically dependent, and limited by severe toxicity. J Exp Med 1988; 167: 1067–85. Baxevanis CN, Voutsas IF, Tsitsilonis OE, et al. Compromised antitumor responses in tumor necrosis factor-alpha knockout mice. Eur J Immunol 2000; 30: 1957–66. Moore RJ, Owens DM, Stamp G, et al. Mice deficient in tumor necrosis factor-alpha are resistant to skin carcinogenesis. Nat Med 1999; 5: 828–31. Suganuma M, Okabe S, Marino MW, et al. Essential role of tumor necrosis factor alpha (TNF-alpha) in tumor promotion as revealed by TNF-alpha-deficient mice. Cancer Res 1999; 59: 4516–18. Arnott CH, Scott KA, Moore RJ, et al. Tumour necrosis factor-alpha mediates tumour promotion via a PKC alpha- and AP-1-dependent pathway. Oncogene 2002; 21: 4728–38. Roberts RA, Kimber I. Cytokines in non-genotoxic hepatocarcinogenesis. Carcinogenesis 1999; 20: 1397–401. Knight B, Yeoh GC, Husk KL, et al. Impaired preneoplastic changes and liver tumor formation in tumor necrosis factor receptor type 1 knockout mice. J Exp Med 2000; 192: 1809–18. Kitakata H, Nemoto-Sasaki Y, Takahashi Y, et al. Essential roles of tumor necrosis factor receptor p55 in liver metastasis of intrasplenic administration of colon 26 cells. Cancer Res 2002; 62: 6682–87. Korner H, Cretney E, Wilhelm P, et al. Tumor necrosis factor sustains the generalized lymphoproliferative disorder (gld) phenotype. J Exp Med 2000; 191: 89–96. Orosz P, Echtenacher B, Falk W, et al. Enhancement of experimental metastasis by tumor necrosis factor. J Exp Med 1993; 177: 1391–98. Malik ST, Naylor MS, East N, et al. Cells secreting tumour necrosis factor show enhanced metastasis in nude mice. Eur J Cancer 1990; 26: 1031–34. Qin Z, Kruger-Krasagakes S, Kunzendorf U, et al. Expression of tumor necrosis factor by different tumor cell lines results either in tumor suppression or augmented metastasis. J Exp Med 1993; 178: 355–60. Komori A, Yatsunami J, Suganuma M, et al. Tumor necrosis factor acts as a tumor promoter in BALB/3T3 cell transformation. Cancer Res 1993; 53: 1982–85. Malik ST, Griffin DB, Fiers W, et al. Paradoxical effects of tumour necrosis factor in experimental ovarian cancer. Int J Cancer 1989; 44: 918–25. Oliff A, Defeo-Jones D, Boyer M, et al. Tumors secreting human TNF/cachectin induce cachexia in mice. Cell 1987; 50: 555–63. Sherry BA, Gelin J, Fong Y, et al. Anticachectin/tumor necrosis factor-alpha antibodies attenuate development of cachexia in tumor models. Faseb J 1989; 3: 1956–62.
572
32 Cahlin C, Korner A, Axelsson H, et al. Experimental cancer cachexia: the role of host-derived cytokines interleukin (IL)-6, IL12, interferon-gamma, and tumor necrosis factor alpha evaluated in gene knockout, tumor-bearing mice on C57 Bl background and eicosanoid-dependent cachexia. Cancer Res 2000; 60: 5488–93. 33 Yoneda T, Alsina MA, Chavez JB, et al. Evidence that tumor necrosis factor plays a pathogenetic role in the paraneoplastic syndromes of cachexia, hypercalcemia, and leukocytosis in a human tumor in nude mice. J Clin Invest 1991; 87: 977–85. 34 Naylor MS, Stamp GW, Foulkes WD, et al. Tumor necrosis factor and its receptors in human ovarian cancer: potential role in disease progression. J Clin Invest 1993; 91: 2194–206. 35 Naylor MS, Stamp GW, Balkwill FR. Investigation of cytokine gene expression in human colorectal cancer. Cancer Res 1990; 50: 4436–40. 36 Szlosarek PW, Grimshaw MJ, Wilbanks GD, et al. Tumor necrosis factor- regulation in normal and malignant ovarian epithelium. Proc Am Assoc Cancer Res 2003; 44: (abstr) R5053. 37 Mizokami A, Gotoh A, Yamada H, et al. Tumor necrosis factoralpha represses androgen sensitivity in the LNCaP prostate cancer cell line. J Urol 2000; 164: 800–05. 38 Vinante F, Rigo A, Tecchio C, et al. Serum levels of p55 and p75 soluble TNF receptors in adult acute leukaemia at diagnosis: correlation with clinical and biological features and outcome. Br J Haematol 1998; 102: 1025–34. 39 Warzocha K, Salles G, Bienvenu J, et al. Tumor necrosis factor ligand-receptor system can predict treatment outcome in lymphoma patients. J Clin Oncol 1997; 15: 499–508. 40 Foa R, Massaia M, Cardona S, et al. Production of tumor necrosis factor-alpha by B-cell chronic lymphocytic leukemia cells: a possible regulatory role of TNF in the progression of the disease. Blood 1990; 76: 393–400. 41 Herrmann F, Helfrich SG, Lindemann A, et al. Elevated circulating levels of tumor necrosis factor predict unresponsiveness to treatment with interferon alfa-2b in chronic myelogenous leukaemia. J Clin Oncol 1992; 10: 631–34. 42 Rossi A, Landolfo G, Sassi D, et al. Immunocytochemical increased evidence of inducible nitric oxide synthase, tumor necrosis factoralpha, and adrenocorticotropic hormone in human peritumoral lymph nodes. Appl Immunohistochem Mol Morphol 2002; 10: 52–56. 43 Tselepis C, Perry I, Dawson C, et al. Tumour necrosis factor-alpha in Barrett’s oesophagus: a potential novel mechanism of action. Oncogene 2002; 21: 6071–81. 44 Wong GH, Elwell JH, Oberley LW, et al. Manganous superoxide dismutase is essential for cellular resistance to cytotoxicity of tumor necrosis factor. Cell 1989; 58: 923–31. 45 Wu S, Boyer CM, Whitaker RS, et al. Tumor necrosis factor alpha as an autocrine and paracrine growth factor for ovarian cancer: monokine induction of tumor cell proliferation and tumor necrosis factor alpha expression. Cancer Res 1993; 53: 1939–44. 46 Gaiotti D, Chung J, Iglesias M, et al. Tumor necrosis factor-alpha promotes human papillomavirus (HPV) E6/E7 RNA expression and cyclin-dependent kinase activity in HPV-immortalized keratinocytes by a ras-dependent pathway. Mol Carcinog 2000; 27: 97–109. 47 Maeda M, Watanabe N, Okamoto T, et al. Endogenous tumor necrosis factor functions as a resistant factor against adriamycin. Int J Cancer 1994; 58: 376–79. 48 Jaiswal M, LaRusso NF, Burgart LJ, et al. Inflammatory cytokines induce DNA damage and inhibit DNA repair in cholangiocarcinoma cells by a nitric oxide-dependent mechanism. Cancer Res 2000; 60: 184–90. 49 Leibovich SJ, Polverini PJ, Shepard HM, et al. Macrophage-induced angiogenesis is mediated by tumour necrosis factor-alpha. Nature 1987; 329: 630–32. 50 Yoshida S, Ono M, Shono T, et al. Involvement of interleukin-8, vascular endothelial growth factor, and basic fibroblast growth factor in tumor necrosis factor alpha-dependent angiogenesis. Mol Cell Biol 1997; 17: 4015–23. 51 Cheng N, Chen J. Tumor necrosis factor-alpha induction of endothelial ephrin A1 expression is mediated by a p38 MAPK- and SAPK/JNK-dependent but nuclear factor-kappa B-independent mechanism. J Biol Chem 2001; 276: 13771–77. 52 De Cesaris P, Starace D, Starace G, et al. Activation of Jun Nterminal kinase/stress-activated protein kinase pathway by tumor necrosis factor alpha leads to intercellular adhesion molecule-1 expression. J Biol Chem 1999; 274: 28978–82. 53 Bussolino F, Camussi G, Baglioni C. Synthesis and release of
THE LANCET Oncology Vol 4 September 2003
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
Tumour necrosis factor
54
55
56 57 58 59 60
61 62
63
Review
platelet-activating factor by human vascular endothelial cells treated with tumor necrosis factor or interleukin 1 alpha. J Biol Chem 1988; 263: 11856–61. Leek RD, Landers R, Fox SB, et al. Association of tumour necrosis factor alpha and its receptors with thymidine phosphorylase expression in invasive breast carcinoma. Br J Cancer 1998; 77: 2246–51. Battegay EJ, Raines EW, Colbert T, et al. TNF-alpha stimulation of fibroblast proliferation: dependence on platelet-derived growth factor (PDGF) secretion and alteration of PDGF receptor expression. J Immunol 1995; 154: 6040–47. Leber TM, Balkwill FR. Regulation of monocyte MMP-9 production by TNF-alpha and a tumour-derived soluble factor (MMPSF). Br J Cancer 1998; 78: 724–32. Rosen EM, Goldberg ID, Liu D et al. Tumor necrosis factor stimulates epithelial tumor cell motility. Cancer Res 1991; 51: 5315–21. Tisdale MJ. Cachexia in cancer patients. Nat Rev Cancer 2002; 2: 862–71. Warzocha K, Ribeiro P, Bienvenu J, et al. Genetic polymorphisms in the tumor necrosis factor locus influence non-Hodgkin’s lymphoma outcome. Blood 1998; 91: 3574–81. Davies FE, Rollinson SJ, Rawstron AC, et al. High-producer haplotypes of tumor necrosis factor alpha and lymphotoxin alpha are associated with an increased risk of myeloma and have an improved progression-free survival after treatment. J Clin Oncol 2000; 18: 2843–51. Nakajima K, Sasaki M, Nojima D, et al. Tumor necrosis factoralpha gene mutations and genotype changes in renal cell carcinoma. J Urol 2001; 165: 612–15. Oh BR, Sasaki M, Perinchery G, et al. Frequent genotype changes at -308, and 488 regions of the tumor necrosis factor-alpha (TNFalpha) gene in patients with prostate cancer. J Urol 2000; 163: 1584–87. Hajeer AH, Lear JT, Ollier WE, et al. Preliminary evidence of an association of tumour necrosis factor microsatellites with increased risk of multiple basal cell carcinomas. Br J Dermatol 2000; 142: 441–45.
THE LANCET Oncology Vol 4 September 2003
64 Ghaderi M, Nikitina L, Peacock CS, et al. Tumor necrosis factor a11 and DR15-DQ6 (B*0602) haplotype increase the risk for cervical intraepithelial neoplasia in human papillomavirus 16 seropositive women in Northern Sweden. Cancer Epidemiol Biomarkers Prev 2000; 9: 1067–70. 65 Bienvenu J, Chvetzoff R, Salles G et al. Tumor necrosis factor alpha release is a major biological event associated with rituximab treatment. J Haematol 2001; 2: 378–84. 66 Ramesh G, Reeves WB. TNF-alpha mediates chemokine and cytokine expression and renal injury in cisplatin nephrotoxicity. J Clin Invest 2002; 110: 835–42. 67 Piquet PF, Collart MA, Grau GE et al. Tumor necrosis factor/cachectin plays a key role in bleomycin-induced pneumopathy and fibrosis. J Exp Med 1989; 170: 655–63. 68 Libura J, Bettens F, Radkowski A, et al. Risk of chemotherapyinduced pulmonary fibrosis is associated with polymorphic tumour necrosis factor-a2 gene. Eur Respir J 2002; 19: 912–18. 69 Feldmann M. Development of anti-TNF therapy for rheumatoid arthritis. Nat Rev Immunol 2002; 2: 364–71. 70 Forster MD, Braybrooke J, Madhusudan S, et al. A Phase II trial using etanercept, a recombinant tumour necrosis factor anatgonist, in metastatic breast cancer. Br J Cancer 2002; 86 (suppl): 15 71 Vassallo R, Matteson E, Thomas CF Jr. Clinical response of rheumatoid arthritis-associated pulmonary fibrosis to tumor necrosis factor-alpha inhibition. Chest 2002; 122: 1093–96. 72 Richardson P, Hideshima T, Anderson K. Thalidomide: emerging role in cancer medicine. Annu Rev Med 2002; 53: 629–57. 73 Turini ME, DuBois RN. Cyclooxygenase-2: a therapeutic target. Annu Rev Med 2002; 53: 35–57. 74 Plummer SM, Holloway KA, Manson MM, et al. Inhibition of cyclo-oxygenase 2 expression in colon cells by the chemopreventative agent curcumin involves inhibition of NFkappaB activation via the NIK/IKK signalling complex. Oncogene 1999; 18: 6013–20. 75 Sueoka N, Suganuma M, Sueoka E, et al. A new function of green tea: prevention of lifestyle-related diseases. Ann N Y Acad Sci 2001; 928: 274–80.
http://oncology.thelancet.com
For personal use. Only reproduce with permission from The Lancet.
573