The SSX-2 gene, which is involved in the t(X;18) translocation of synovial sarcomas, codes for the human tumor antigen HOM-MEL-40.

Using autologous serum for the serological analysis of recombinantly expressed clones (SEREX) from a cDNA derived from a human melanoma, several new melanoma antigens were identified that are immunogenic in the autologous host. Sequence analysis revealed that one of these antigens, HOM-MEL-40, was coded for by the SSX2 gene, which has recently been described to be involved in the t(X;18) translocation of human synovial sarcomas. Expression analysis performed by Northern blot and RT-PCR demonstrated the presence of HOM-MEL-40 transcripts in a significant proportion of human melanomas (50%), colon cancers (25 %), hepatocarcinomas (30%), and breast carcinoma (20%) but not in normal tissues except for testis. Sequence comparison with transcripts cloned from testis ruled out mutations in the melanoma-derived HOM-MEL-40. Antibodies against HOM-MEL-40 were found in 10 of 89 patients with melanoma, including 3 of 8 patients with HOM-MEL-40-positive tumors, but not in 41 apparently healthy controls. In view of the specific expression pattern and immunogenicity in cancer patients, HOM-MEL-40 holds promise as a target for immune interventions in a considerable population of patients with HOM-MEL-40-positive tumors.

Mimicry of the Hodgkin-associated IRAC antigen by an anti-idiotype network: potential use in active immunotherapy of Hodgkin's lymphoma.

The murine monoclonal antibody anti-IRac, defining a surface-antigen structure (MW 70 kDa) on Hodgkin-Reed-Sternberg and interdigitating reticulum cells, was used to generate a cascade of anti-idiotypic antibodies as well as a cellular immunity against Hodgkin-Reed-Sternberg cells in syngeneic BALB/c-mice. The anti-idiotypic antibody monoclonal antibody 4B4 demonstrated characteristics of an "internal image" or network antibody (Ab2 beta). Ab2 beta 4B4 bound specifically to anti-IRac and inhibited anti-IRac-binding to antigen-bearing cells effectively. 4B4 induced an IRac-specific humoral polyclonal immune response in BALB/c-mice and New Zealand white rabbits as judged by Flow cytometric and histochemical analysis. Moreover, BALB/c-mice immunized with 4B4 showed statistically significant (p > 0.01) delayed-type hypersensitivity reaction against IRac-expressing Hodgkin cell-lines. Ab2 beta 4B4 induced in syngeneic BALB/c mice a monoclonal anti-anti-idiotypic antibody (Ab3) termed 3G10, which mimicked the specificities of Ab1 anti-IRac and thus confirms the internal image nature of Ab2 beta 4B4. The anti-idiotype-induced tumor cell specific T- and B-cell mediated immune response even across species barriers via the Hodgkin related IRac-antigen may play an important part in active specific immunotherapy of Hodgkin's disease.

The clinical significance of cytokines and soluble forms of membrane-derived activation antigens in the serum of patients with Hodgkin's disease.

In a search for specific serum markers with prognostic impact in Hodgkin's Disease (HD), we evaluated the clinical significance of several cytokines (IL-1 beta, IL-2, IL-3, IL-6, G-CSF, GM-CSF, TNF-alpha) and soluble forms of membrane-derived antigens (sCD4, sCD8, sCD23, sCD25, sCD30) in the serum of patients with untreated HD. Elevations of three groups of serum factors were observed: Firstly, elevations of the hematopoietic cytokines GM-CSF (detected in 39%), IL-6 (57%) and IL-3 (13%), which occurred simultaneously in the majority of the cases; secondly, simultaneous elevations of the inflammatory cytokines TNF-alpha and IL-1 beta (detected in 7%); and finally, elevations of membrane-derived activation antigens sCD8, sCD25, and sCD30. While the cytokine levels did not correlate with other obvious parameters, the membrane-derived activation antigens sCD8, sCD25 and sCD30 were associated with a poor prognosis. Only sCD30 correlated with disease activity and holds promise for the follow-up of patients in remission. Further investigations of these parameters at the cellular level might help to elucidate the enigmatic biology of HD.

CD30-specific AB1-AB2-AB3 internal image antibody network: potential use as anti-idiotype vaccine against Hodgkin's lymphoma.

The tumor-associated CD30 antigen is presently under study as a target for active specific immunotherapy of Hodgkin's lymphoma with anti-idiotypic antibodies. Internal image antibodies (Ab2 beta) 9G10 and 14G9 against the CD30-specific antibody HRS-4 (Ab1) have been described, which induce a CD30-specific T- and B-cell response in BALB/c mice and New Zealand white rabbits. In extension of this work, murine monoclonal anti-idiotypic Ab2 beta 9G10, mimicking structures of the nominal CD30 antigen, was used to generate monoclonal Ab3 in mice and polyclonal Ab3 in rabbits with specificity for CD30. The Ab2 beta 9G10-specific murine monoclonal Ab3 4A4 bound specifically to the 120-kDa band of CD30 present on Hodgkin cell lines and Hodgkin tumor tissue, and effectively inhibited binding of Ab1 HRS-4 to Ab2 9G10 as well as to CD30+ cells. Monoclonal Ab3 4A4 was cytotoxic for CD30+ cell lines in vitro and effectively prevented the s.c. growth of L540 cell tumors after passive i.v. administration in a SCID mouse tumor model. While this cytotoxic effect of the IgM subclass monoclonal Ab3 4A4 was due to complement activation, the murine monoclonal Ab1 HRS-4 and a polyclonal Ab3 preparation of IgG-subclass from New Zealand white rabbits were cytotoxic by an antibody-dependent cell-mediated mechanism in vitro. In conclusion, Ab2 beta 9G10 is able to induce a CD30-specific cytotoxic IgG and IgM response. Cytotoxicity was shown to be mediated by complement activation and antibody-dependent cell-mediated cytotoxicity in vitro and in vivo and across species barriers. Thus, the CD30-like Ab2 beta 9G10 may hold promise for effective active specific immunotherapy of human Hodgkin's lymphoma.

Increased levels of circulating cytokines in patients with untreated Hodgkin's disease.

Expression of several cytokines has been demonstrated in Hodgkin and Reed-Sternberg (H&RS) cells in vitro and in vivo. In order to determine whether interleukin-1 beta (IL-1 beta), IL-3, IL-6, GM-CSF, G-CSF, and TNF-alpha are elevated in Hodgkin's disease (HD), we tested the sera of untreated patients with HD by means of sensitive sandwich ELISAs. GM-CSF was detected in 22/56 patients (39%; range 40-140 pg/ml), IL-3 in 5/40 (13%; range 13-26 pg/ml), and IL-6 in 32/56 patients (57%; range 12-332 pg/ml). TNF-alpha and IL-1 beta were detected in only 3/43 patients (7%; range: TNF-alpha: 36-66 pg/ml; IL-1 beta: 389-1505 pg/ml) and G-CSF not at all. All patients with measurable IL-3 levels had both elevated IL-6 and GM-CSF levels, and the majority of patients with elevated IL-6 also had elevated GM-CSF levels and vice versa. In contrast, the 3/40 patients with both measurable IL-beta and TNF-alpha did not have elevated IL-3, IL-6, or GM-CSF levels. Cytokine levels were independent of stage or the presence of B-symptoms, and there was no correlation with any other clinical or laboratory parameter. Elevations of the respective cytokines might be a means to maintain normal blood cell counts in the respective patients with HD.

Human neoplasms elicit multiple specific immune responses in the autologous host.

Expression of cDNA libraries from human melanoma, renal cancer, astrocytoma, and Hodgkin disease in Escherichia coli and screening for clones reactive with high-titer IgG antibodies in autologous patient serum lead to the discovery of at least four antigens with a restricted expression pattern in each tumor. Besides antigens known to elicit T-cell responses, such as MAGE-1 and tyrosinase, numerous additional antigens that were overexpressed or specifically expressed in tumors of the same type were identified. Sequence analyses suggest that many of these molecules, besides being the target of a specific immune response, might be of relevance for tumor growth. Antibodies to a given antigen were usually confined to patients with the same tumor type. The unexpected frequency of human tumor antigens, which can be readily defined at the molecular level by the serological analysis of autologous tumor cDNA expression cloning, indicates that human neoplasms elicit multiple specific immune responses in the autologous host and provides diagnostic and therapeutic approaches to human cancer.