Reiner Caspari

MUTYH‐associated polyposis: 70 of 71 patients with biallelic mutations present with an attenuated or atypical phenotype

To determine the frequency, mutation spectrum and phenotype of the recently described autosomal recessive MUTYH‐associated polyposis (MAP), we performed a systematic search for MUTYH (MYH) mutations by sequencing the complete coding region of the gene in 329 unselected APC mutation‐negative index patients with the clinical diagnosis of familial adenomatous polyposis (FAP) or attenuated FAP (AFAP). Biallelic germline mutations in MUTYH were identified in 55 of the 329 unselected patients (17%) and in another 9 selected index cases. About one‐fifth (20%) of the 64 unrelated MAP patients harboured none of the 2 hot‐spot missense mutations Y165C and/or G382D. Including 7 affected relatives, almost all MAP patients presented with either an attenuated (80%) or with an atypical phenotype (18%). Fifty percentage of the MAP patients had colorectal cancer at diagnosis. Duodenal polyposis was found in 18%, thyroid and stomach cancer in 1 case, other extraintestinal manifestations associated with FAP were not observed. In 8 families, vertical segregation was suspected; in 2 of these families, biallelic mutations were identified in 2 generations. Monoallelic changes with predicted functional relevance were found in 0.9% of the 329 patients, which is in accordance with the carrier frequency in the general population. In conclusion, biallelic MUTYH mutations are the underlying genetic basis in a substantial fraction of patients with adenomatous polyposis. The phenotype of MAP is best characterised as attenuated or atypical, respectively. Colorectal surveillance starting at about 18 years of age is recommended for biallelic mutation carriers and siblings of MAP patients, who refuse predictive testing.

Attenuated familial adenomatous polyposis due to a mutation in the 3′ part of the APC gene. A clue for understanding the function of the APC protein

The identification of germline mutations in a large number of clinically well-characterised patients with familial adenomatous polyposis (FAP) has allowed the unravelling of several genotype-phenotype relationships that can now be interpreted in the light of the structure and functional domains of the adenomatous polyposis coli (APC) protein. An attenuated phenotype has been found to be associated with mutations at the 5′ end of the gene, while a severe clinical expression was found in patients with the most common mutation at codon 1309. So far, only few mutations in the 3′ half of the gene have been published. We report on two families with a rather mild phenotype due to a frameshift mutation at codon 1597. These families may represent a clue for defining a 5′ border for the occurrence of a second region of attenuated FAP that is localised in the 3′ part of the APC gene. We propose a model to explain the relationship between the severity of the disease and the size of the mutant APC protein.

Allogeneic dendritic cells fused with tumor cells: preclinical results and outcome of a clinical phase I/II trial in patients with metastatic renal cell carcinoma.

Therapeutic vaccination with dendritic cells (DC) can lead to tumor regression in animal models and has shown promising results in the first clinical trials of metastatic renal cell carcinoma and malignant melanoma. In vitro data and results of a clinical phase I/II trial using DC tumor fusions in patients with progressive metastatic renal cell carcinoma are presented here. In addition to toxicity and feasibility, complex immune monitoring was a point of interest. DC precursor cells were obtained from the peripheral blood mononuclear cells (PBMCs) of healthy donors and were fused with either allogeneic (8 patients) or autologous (4 patients) renal tumor cells. In total, 12 patients with progressive metastatic renal cell carcinoma were treated with an average of 2.8 x 10(7) tumor cells fused with 1.8 x 10(7) DC each administered on days 0, 28, and 56 intradermally. Fusion efficacy for the tumor cells used was 14.3% +/- 7.8%. Cell viability was 59.8% +/- 6.8% after fusion and irradiation. We observed no adverse effects and no difference in clinical outcome between the allogeneic and the autologous treatment. Eight patients remained in a progressive disease state and four patients in a stable disease state. T-cell immunity was carefully monitored before, during, and after treatment. Delayed-type hypersensitivity (DTH) reaction using tumor cells was positive after treatment in 7 of 12 patients, 2 of whom were found to have stable disease. An increase in the reactivity against recall antigens was seen in most patients. Interestingly, cytotoxicity of peripheral blood lymphocytes (PBLs) against renal cell carcinoma cells increased during treatment as well as the percentage of interferon-gamma-secreting cells. This effect was significantly enhanced within the group that had stable disease. The lack of adverse effects together with positive immunologic signs justifies further investigation of this novel therapeutic approach. Further studies are necessary to test for clinical effectiveness in patients with tumors, especially those with less advanced disease.

Frequency and parental origin of de novo APC mutations in familial adenomatous polyposis

A predominance of de novo mutations in the paternal germ line has been reported for several disorders; however, in familial adenomatous polyposis (FAP), the parental origin of APC mutations has been scarcely analysed so far. Among 563 unrelated FAP families with known family history, we identified 58 patients with a suspected de novo mutation in the APC gene. A germline mutation was detected in 52 of them; in 38 patients, the mutation could be excluded in both parents. The five base pair deletion at codon 1309 (c.3927_3931delAAAGA) was over-represented in the group of patients with suspected de novo mutations (17/58=29%), when compared to the group of familial cases (26/505=5%); thus, the high frequency of this mutation is not due to a founder effect but rather due to de novo mutation events. Parental origin of de novo mutations could be traced in 16 families, including three families with large chromosomal deletions. Four mutations were of maternal and 12 of paternal origin, pointing to a moderate preponderance towards paternal origin. Sex-related differences of mutation types could be observed: large deletions and single-base substitutions were exclusively of paternal origin, whereas the small deletions were equally distributed (maternal/paternal ratio 4:4).

Familial adenomatous polyposis: a submicroscopic deletion at the APC locus in a family with mentally normal patients.

Cytogenetically visible deletions that include the adenomatosis polyposis coli (APC) locus have repeatedly been reported in mentally handicapped polyposis patients. We report on a family with a submicroscopic deletion of about 200 kb including more than the 3′ half of the APC gene and the adjacent DPI gene. The deletion was detected by linkage analysis with flanking and intragenic markers and proven by in situ hybridisation with intragenic cosmid clones. All the familial adenomatous polyposis (FAP) patients and persons at risk in the family show normal behaviour and intelligence. Thus, it is conceivable that at least some of the FAP patients in whom mutations could not be identified by routine methods may have large but submicroscopic deletions.

Arylamine N-acetyltransferase type 2 and glutathione S-transferases M1 and T1 polymorphisms in familial adenomatous polyposis

Familial adenomatous polyposis (FAP) exhibits a variable phenotype even in carriers of the same adenomatous polyposis coli germline mutation. Xenobiotic-metabolizing enzymes such as N-acetyltransferases (NATs) and glutathione S-transferases (GSTs) were reported to modify the individual risk for colorectal cancer. We examined whether the polymorphisms of the NAT2, GSTM1, and GSTT1 enzymes affect age at diagnosis of first colorectal adenomas or extracolonic manifestations in 411 FAP patients. Neither age at diagnosis of colorectal adenomas nor occurrence of extra-intestinal tumors differed significantly between genotypes at the NAT2 and GSTM1 loci, whereas GSTT1 polymorphism showed an uncertain association with extra-intestinal manifestations. Combinations of supposed at risk genotypes of the three enzymes showed no significant differences either. Thus, NAT2, GSTM1, or GSTT1 are unlikely to modify the disease phenotype in FAP patients.

Mapping of a Gene for nonspecific x-linked mental retardation (MRX 75) to Xq24-q26

Nonspecific X-linked mental retardation is a heterogeneous condition consisting of nonsyndromal mental retardation in males. It is caused by mutation in one of several genes on the X chromosome (MRX genes). Here we report on the localization of a presumptive MRX gene to chromosomal region Xq24-q26 in a German family with nonspecific X-linked mental retardation (MRX 75, HUGO Human Gene Nomenclature Committee). Two point linkage analysis with 23 informative markers gave a lod score of 2.53 at theta = 0 for markers DXS425, DXS1254, DXS1114, and HPRT.

Impact de la génétique moléculaire sur le dépistage du cancer colorectal héréditaire non polypoïde

RésuméLe syndrome HPNCC (Cancer colorectal héréditaire non polypoïde) connu aussi sous l’appellation de «Syndrome de Lynch» est une prédisposition précancéreuse autosomale dominante responsable d’environ 2 à 5 % des cancers colorectaux (CCR). En outre, le risque de développer des lésions malignes extra coliques est très élevé dans ce syndrome. Le syndrome HPNCC résulte de mutations de lignée germinale au niveau d’au moins un des cinq gènes codant les protéines, ce qui conduit à un défaut d’appariement au niveau du complexe MMR. Ce complexe (avec d’autres) assure la stabilité du génome au cours de la division cellulaire. Les mutations au niveau d’un seul des gènes MMR conduit à une perte de capacité réparatrice du complexe MMR avec pour conséquence une instabilité de l’ADN encore appelée instabilité microsatellite (MSI).La reconnaissance des gènes MMR permet l’identification des patients atteints de syndrome HNPCC, entité encore gênée par le manque d’un phénotype non équivoque de cette maladie. Par conséquent, ceci a conduit à une définition clinique du syndrome HNPCC (c’est-à-dire porteurs ou non de tumeurs extra-coliques propres au syndrome). Par ailleurs, la surveillance des personnes à risque de HNPCC pourrait être nettement améliorée.L’identification des gènes MMR et la compréhension de leur rôle favorisent une meilleure connaissance de la carcinogenèse du CCR. En dehors des tumeurs dues au syndrome HNPCC, plus de 20 % des CCR sporadiques sont causés par des déficiences du complexe MMR. Nous savons à présent que ces cancers se développent différemment des 80 % de cancers restants. A l’avenir, ces différences devraient influencer leur traitement.SummaryHereditary non-polyposis colorectal cancer (HNPCC), also known as Lynch syndrome, is an autosomal dominant precancerous condition accounting for about 2–5 % of colorectal cancers. In addition, the risk to develop extracolonic malignancies is markedly elevated in affected persons. HNPCC is caused by germline mutations in one of at least five genes coding for proteins which build up the human mismatch repair (MMR) complex. This complex (among others) cares for stability of the genome during cell division. Mutations in one of the MMR genes lead to loss of repair capacity of the MMR complex and subsequently to a type of DNA instability which is called microsatellite instability (MSI).Identification of the MMR genes enabled the identification of HNPCC patients, which is hampered by lack of an unambiguous phenotype of the disease. Thus, it helped also with the clinical definition of HNPCC (e. g., which extracolonic tumours belong to the syndrome and which do not). Furthermore, surveillance in persons at risk for HNPCC could be clearly improved.Identification of the MMR genes and understanding of their function, on the other side, has led to an expansion of our knowledge about colorectal carcinogenesis. Not only tumours in HNPCC patients, but up to 20 % of sporadic colorectal cancers are caused by defects of the mismatch repair complex. We now know that these tumours develop differently from the remaining 80 % of CRC. In the future, these differences may even influence our treatment.