Cancer Immune Therapy Edited by G. Stuhler and P. Walden ...

5.3 Determination of the Expression of Classical and Non-classical MHC Antigens
5.2
The Physiology of the Non-classical HLA-G Molecule
The non-classical HLA class Iantigens, e.g. HLA-G, are characterized by a limited
polymorphism, unique structural features and a tissue-restricted distribution. Under
physiological conditions, HLA-G is selectively expressed on trophoblasts of the human
placenta, in fetal eye, liver and thymus, and in adult eye, skin, keratinocytes
and myelomonocytic cells. A major feature of HLA-G is the alternative splicing of
mRNA, thereby encoding four different potentially membrane-bound proteins,
HLA-G1, -G2, -G3 and -G4, and three soluble isoforms, HLA-G5, -G6 and -G7 [19,
20]. The HLA-G1 isoform has the classical MHC class Istructure consisting of the
a1, a2 and a3 extracellular domains non-covalently associated with b2-m. Like the
full-length HLA-G1 molecule, each truncated HLA-G isoform can also be expressed
on the cell surface [21].
The functions of HLA-G are now emerging. HLA-G binds to nonameric peptides and
is capable of presenting them to CD8 + Tcells [22, 23]. In addition, HLA-G is the ligand
of some killer cell immunoglobulin-like receptors presented on lymphocytes, macrophages,
dendritic cells (DCs) and natural killer (NK) cells, such as p49, ILT2 and ILT4,
and can modulate the expression of HLA-E. Studies on the immunological function
have identified HLA-G as a key mediator of immune tolerance, in particular in the interaction
between fetus and mother, possibly by inhibiting alloreactivity of maternal
NK cells against the fetal tissues [2]. Furthermore, it has been suggested that HLA-G
molecules may provide tumor cells with an effective escape mechanism by inhibiting
both the lytic activity of in situ antigen-specific CD8 + CTLs [24] and the NK cellmediated
cytotoxicity through interaction with killer inhibitory receptors (KIRs) [25].
5.3
Determination of the Expression of Classical and Non-classical MHC Antigens
The pattern of MHC class Iand IIantigens as well as HLA-G expression has been
studied in many tumors of distinct origin using different experimental approaches.
In general, immunohistochemical staining of paraffin-embedded, formalin-fixed or
frozen tumor sections has been employed [26±28]. However, the lack of standardized
protocols that can be applied consistently by different investigators rendered it difficult
to compare results obtained in various studies [29]. In addition, such analyses
are significantly hampered by both the availability and reliability of reagents. So far,
only a limited number of well-characterized anti-MHC class I, class II and non-classical
MHC class Ipan-reactive antibodies as well as allele- and locus-specific reagents
are available and can be successfully employed for staining formalin-fixed tissue sections
[30, 31].
Another hurdle is the evaluation and interpretation of the immunohistochemical
staining in terms of its classification and scoring, thereby differentiating between a
highly significant loss or decrease in expression. This is even more difficult since a
heterogeneic MHC expression pattern is often found within the tumor analyzed.
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