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Two TNF receptor types have been characterized. The type I receptor displays a molecular mass of 55 kDa, and is known as TNF-R55 (also P-55, TNFR1 or CD120a). The type II receptor is larger (75 kDa) and is known as TNF-R75 (also P-75, TNFR2 or CD120b). Both receptors bind both TNF-a and TNF-b. A TNF receptor is present on the plasma membrane of almost all nucleated cell types, generally in numbers varying from 100-10000. Although virtually all cells express TNF-R55, the TNF-R75 cell surface receptor distribution is more restricted — being most prominent on haematopoietic cells. Differential regulation of expression of these two receptor types is also apparent. Generally, low constitutive expression of TNF-R55 is observed, with TNF-R75 expression being inducable.
Both receptor types are members of the nerve growth factor receptor superfamily and exhibit the characteristic four (cysteine-rich) repeat units in their extracellular domain. The extracellular domains of TNF-R55 and TNF-R75 exhibit only 28% homology while their intracellular domains are devoid of any homology, indicating the likely existence of distinct signalling mechanisms.
It appears that TNF-R55 is capable of mediating most TNF activities, while the biological activities induced via the TNF-R75 receptor are more limited. For example, TNF’s cytotoxic activity, as well as its ability to induce synthesis of various cytokines and prostaglandins, are all mediated mainly/exclusively by TNF-R55. TNF-R75 appears to play a more prominent role in the induction of synthesis of T lymphocytes. All of the biological activities mediated by TNF-R75 can also be triggered via TNF-R55, and usually at much lower densities of receptors. TNF-R75 thus appears to play more of an accessory role, mainly to enhance effects mediated via TNF-R55.
Figure 5.11. TNF binding to its receptor (TNF-R55), with resulting clustering of the receptor and generation of intracellular signals. Binding of TNF to its other receptor (TNF-R75) also induces receptor clustering (see text for details)
Binding of TNF to either receptor type results in oligomerization of the receptor (Figure 5.11). Indeed, antibodies raised against the extracellular domains of the receptors can induce TNF activity, indicating that the major/sole function of the TNF ligand is to promote such clustering of receptors. In most cases, binding of TNF to TNF-R55 results in rapid internalization of the ligand-receptor complex, followed by lysosomal degradation. In contrast, binding of TNF to TNF-R75 does not induce such receptor internalization. In some cases, ligand-binding appears to activate selective cleavage of the extracellular domain, resulting in the release of soluble TNF-R75. Soluble forms of both receptor types have been found in both the blood and urine.
The exact molecular mechanisms by which TNF-induced signal transduction are mediated remain to be characterized in detail. Oligomerization of the receptors is often followed by their phosphorylation—most likely by accessory kinases that associate with the intracellular domain of the receptor (neither receptor type displays intrinsic protein kinase activity). The existence of several phosphoproteins capable of associating with (the intracellular domain of) TNF-R55 and TNF-R75 has also been established. Following clustering of the TNF receptors, these associated proteins are likely to become activated, thus mediating additional downstream events which eventually trigger characteristic TNF molecular responses. The downstream events are complex and varied. Experimental evidence from various studies implicate a variety of mechanisms, including phosphorylation events, as well as activation of various phospholipases, resulting in the generation of messengers such as diacylglycerol, inositol phosophates and arachidonic acid.
TNF: therapeutic aspects
The initial interest in utilizing TNF as a general anti-cancer agent has diminished, largely due to the realization that:
• many tumours are not susceptible to destruction mediated by TNF (indeed, some tumours produce TNF as an autocrine growth factor);
• tumour cell necrosis is not TNF’s major biological activity;
CYTOKINES: INTERLEUKINS AND TUMOUR NECROSIS FACTOR 251
Figure 5.12. Overview of the likely steps undertaken during the manufacture of the recombinant TNF-a product Beromun. Exact details do not appear to be freely available. It is unclear from the publicly available descriptions whether the product accumulates intracellularly in soluble form or in the form of inclusion bodies
Table 5.11. Some diseases in which TNF is known to mediate many of the symptoms
Cancer Septic shock
Multiple sclerosis Diabetes
Symptoms induced by TNF