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A Hemminki

Publications and source records attributed to A Hemminki.

At least 37 records · Page 2Linked to original sources
PubMed1999

The molecular basis and clinical aspects of Peutz-Jeghers syndrome.

Peutz-Jeghers syndrome (PJS) is a classic, but not widely known hereditary trait [1, 2]. Its clinical hallmarks are intestinal hamartomatous polyposis and melanin pigmentation of the skin and mucous membranes. In addition, PJS predisposes to cancer [3, 4]. The most common malignancies are small intestinal, colorectal, stomach and pancreatic adenocarcinomas. Other cancer types that probably occur in excess in PJS families include breast and uterine cervical cancer, as well as testicular and ovarian sex cord tumors. The relative risk of cancer may be as high as 18 times that of the general population, and the cancer patients' prognosis is reduced. Recently, the predisposing locus was mapped to 19p13.3 using a novel method [5]. Subsequently, the causative gene was shown to be LKB1 (a.k.a. STK11), a serine/threonine kinase of unknown function [6]. Although preliminary reports seem to suggest a minor role for LKB1 in sporadic tumorigenesis [7-12], further investigations are needed.

AMP-Activated Protein Kinase Kinases↗
PubMed1999

LKB1 somatic mutations in sporadic tumors.

Germline mutations of LKB1/Peutz-Jeghers syndrome gene predispose carriers to hamartomatous polyposis of the gastrointestinal tract as well as to cancer of different organ systems. Although Peutz-Jeghers syndrome patients frequently present with neoplasms of the colon, stomach, small intestine, pancreas, breast, ovaries, and cervix, somatic mutations appear to be rare in the sporadic tumor types thus far studied (colorectal, gastric, testicular, and breast cancers). To evaluate whether somatic mutations of LKB1 contribute to the tumorigenesis of yet unstudied tumor types, we screened 14 cell lines and 129 tumor specimens from different cancers for a genetic defect in LKB1. Six melanoma and eight myeloma cell lines were scrutinized for LKB1 somatic mutations by genomic sequencing. No changes were found in the coding LKB1 sequence and exon/intron boundaries. Next, we analyzed 12 pancreatic, 8 gastric, 12 ovarian granulosa cell, 26 cervical, 28 lung, 24 soft tissue, and 19 renal tumors by single-strand conformational polymorphism analysis. Three changes in LKB1 coding nucleotide sequence were identified. One base pair deletion at A957 and G958 substitution by T occurred in a cervical adenocarcinoma sample, resulting in a frameshift and premature stop codon at position 335. Substitution of A581 by T occurred in a lung adenocarcinoma sample, resulting in the change of aspartic acid at position 194 to valine. A loss of another allele was detected in this sample. One silent change, C1257T, was found in a pancreatic carcinoma sample. The changes were not present in the matched normal tissue DNA samples. Our results suggest that mutational inactivation of LKB1 is a rare event in most sporadic tumor types.

AMP-Activated Protein Kinase Kinases↗
PubMed1999

Mutations and impaired function of LKB1 in familial and non-familial Peutz-Jeghers syndrome and a sporadic testicular cancer.

Germline mutations in LKB1 have been reported to underlie familial Peutz-Jeghers syndrome (PJS) with intestinal hamartomatous polyps and an elevated risk of various neoplasms. To investigate the prevalence of LKB1 germline mutations in PJS more generally, we studied samples from 33 unrelated PJS patients including eight non-familial sporadic patients, 20 familial patients and five patients with unknown family history. Nineteen germline mutations were identified, 12 (60%) in familial and four (50%) in sporadic cases. LKB1 mutations were not detected in 14 (42%) patients, indicating that the existence of additional minor PJS loci cannot be excluded. LKB1 is predicted to encode a serine/threonine kinase. To demonstrate the putative Lkb1 kinase function and to study the consequences of LKB1 mutations in PJS and sporadic tumors, we have analyzed the kinase activity of wild-type and mutant Lkb1 proteins. Interestingly, while most of the small deletions or missense mutations resulted in loss-of-function alleles, one missense mutation (G163D) previously identified in a sporadic testicular tumor demonstrated severely impaired but detectable kinase activity.

AMP-Activated Protein Kinase Kinases↗
PubMed1998

Incidence of hereditary nonpolyposis colorectal cancer and the feasibility of molecular screening for the disease.

BACKGROUND: Genetic disorders that predispose people to colorectal cancer include the polyposis syndromes and hereditary nonpolyposis colorectal cancer. In contrast to the polyposis syndromes, hereditary nonpolyposis colorectal cancer lacks distinctive clinical features. However, a germ-line mutation of DNA mismatch-repair genes is a characteristic molecular feature of the disease. Since clinical screening of carriers of such mutations can help prevent cancer, it is important to devise strategies applicable to molecular screening for this disease. METHODS: We prospectively screened tumor specimens obtained from 509 consecutive patients with colorectal adenocarcinomas for DNA replication errors, which are characteristic of hereditary colorectal cancers. These replication errors were detected through microsatellite-marker analyses of tumor DNA. DNA from normal tissue from the patients with replication errors was screened for germ-line mutations of the mismatch-repair genes MLH1 and MSH2. RESULTS: Among the 509 patients, 63 (12 percent) had replication errors. Specimens of normal tissue from 10 of these 63 patients had a germ-line mutation of MLH1 or MSH2. Of these 10 patients (2 percent of the 509 patients), 9 had a first-degree relative with endometrial or colorectal cancer, 7 were under 50 years of age, and 4 had had colorectal or endometrial cancer previously. CONCLUSIONS: In this series of patients with colorectal cancer in Finland, at least 2 percent had hereditary nonpolyposis colorectal cancer. We recommend testing for replication errors in all patients with colorectal cancer who meet one or more of the following criteria: a family history of colorectal or endometrial cancer, an age of less than 50 years, and a history of multiple colorectal or endometrial cancers. Patients found to have replication errors should undergo further analysis for germ-line mutations in DNA mismatch-repair genes.

Adaptor Proteins, Signal Transducing↗
PubMed1998

Somatic mutations in LKB1 are rare in sporadic colorectal and testicular tumors.

Germ-line mutations in a serine/threonine kinase gene, LKB1, were recently shown to underlie Peutz-Jeghers syndrome (PJS), a hereditary disorder that predisposes to benign and malignant tumors of multiple organ systems. Most mutations that have been described thus far dramatically change the predicted protein and are likely to be of an inactivating nature. This observation and a previous observation that the LKB1 locus is often deleted in PJS polyps suggest that the gene may function as a tumor suppressor. We examined whether somatic mutations in this gene are present in sporadic carcinomas of the colon and testis, tumors that are characteristic of PJS. First, 20 randomly selected colorectal and 28 testicular tumors were analyzed by single-strand conformation polymorphism analysis. No mutations in LKB1 were found in colorectal tumors. One testicular tumor displayed a heterozygous missense type variant, in which glycine 163 was changed to aspartic acid. This change was absent in the DNA of normal tissue. To better focus our efforts, we tested 75 additional colon carcinomas for loss of heterozygosity at 19p, where LKB1 is localized. Of 75 samples analyzed, 50 were informative with a closely linked marker, D19S886, and 13 (26%) of these displayed loss of heterozygosity. The 13 tumors were scrutinized for LKB1 mutations by genomic sequencing. This analysis revealed no changes. Together, these findings suggest that somatic mutations of LKB1 are not frequent in colorectal and testicular cancer.

AMP-Activated Protein Kinase Kinases↗
PubMed1998

Intestinal cancer in patients with a germline mutation in the down-regulated in adenoma (DRA) gene.

A recent study has revealed that germline mutations of the down-regulated in adenoma (DRA) gene are a likely cause of a recessive intestinal absorption defect, congenital chloride diarrhea. This finding was in accordance with previous works showing that DRA encodes a sodium independent transporter for sulfate and oxalate. Although DRA was originally reported as a candidate tumor suppressor, these studies have questioned the relevance of DRA in cancer. To evaluate whether further studies on the role of DRA in tumorigenesis are still of interest, we examined whether individuals carrying germline DRA mutations have an excess of intestinal cancer. Cancer status of 229 members of 36 Finnish congenital chloride diarrhea families (44 homozygous patients, 70 heterozygous parents, and 115 grandparents at 50% risk of being a DRA mutation carrier) was checked at the Finnish Cancer Registry and the risk of intestinal cancer was found slightly elevated (standardized incidence ratio 3.4, 95% confidence interval 1.4-7.0, P < 0.05). While this result does not unambiguously demonstrate an increased intestinal cancer risk in DRA mutation carriers, it should promote further studies to determine the possible role of DRA in cancer.

Adenoma↗
PubMed1998

A serine/threonine kinase gene defective in Peutz-Jeghers syndrome.

Studies of hereditary cancer syndromes have contributed greatly to our understanding of molecular events involved in tumorigenesis. Here we investigate the molecular background of the Peutz-Jeghers syndrome (PJS), a rare hereditary disease in which there is predisposition to benign and malignant tumours of many organ systems. A locus for this condition was recently assigned to chromosome 19p. We have identified truncating germline mutations in a gene residing on chromosome 19p in multiple individuals affected by PJS. This previously identified but unmapped gene, LKB1, has strong homology to a cytoplasmic Xenopus serine/threonine protein kinase XEEK1, and weaker similarity to many other protein kinases. Peutz-Jeghers syndrome is therefore the first cancer-susceptibility syndrome to be identified that is due to inactivating mutations in a protein kinase.

AMP-Activated Protein Kinase Kinases↗
PubMed1998

A novel splice site mutation of the EXT2 gene in a Finnish hereditary multiple exostoses family. Mutations in brief no. 197. Online.

Hereditary multiple exostoses is a dominantly inherited disease characterized by multiple benign osteochondromas. The affected individuals have an increased risk of developing sarcoma. A large Finnish family with hereditary multiple exostosis was analyzed to find the disease-causing mutation. Blood samples were obtained from 35 family members, including 21 affected and 14 unaffected individuals. Using 2-point linkage analysis the EXT phenotype was shown to be linked to the recently cloned EXT2 gene on chromosome 11p11. The coding region of the gene was sequenced and a previously unreported splice site mutation found. This G to T transversion within a 5-prime splice donor site following exon 6 was shown to cause aberrant splicing of RNA. The described change is considered to be a novel disease-causing mutation in the EXT2 gene.

Alternative Splicing↗
PubMed1998

Fine tuning of an anti-testosterone antibody binding site by stepwise optimisation of the CDRs.

BACKGROUND: We have previously reported specificity improvement of an anti-testosterone monoclonal antibody (3-C4F5) by random mutagenesis of the third complementarity determining regions (CDR3s) and by phage display selection. OBJECTIVES: Here we extend the mutagenesis strategy to the other CDRs and select the mutant libraries using two different approaches in order to further fine-tune the binding properties of this recombinant Fab fragment. STUDY DESIGN: To improve the affinity the new mutant libraries were selected by using limiting, decreasing concentrations of biotinylated testosterone (TES) in solution and capturing the binders on streptavidin-coated microtiter plate. The specificity was improved by preincubating the mutant libraries in solution with a high concentration of the most problematic cross-reacting steroid, dehydroepiandrosterone sulfate (DHEAS). RESULTS: In two different light chain CDR1 mutant clones isolated from the affinity pannings, the relative TES affinity was increased over 10-fold while the cross-reactivities to related steroids were preserved at the same level as in the parental combined CDR3 mutant clone. New heavy chain CDR1 and light chain CDR2 mutants showing slightly decreased cross-reactivities were isolated from specificity selections. By combining compatible mutant CDRs together we were able to create a Fab fragment with over 12-fold higher relative TES affinity and significantly lower cross-reactivity to DHEAS when compared to the original monoclonal antibody 3-C4F5. CONCLUSIONS: Our results demonstrate that a high-affinity and selective recombinant Fab fragment working over a wide TES concentration range with clinical samples could be generated by CDR mutagenesis and phage display selection.

Antibody Affinity↗
PubMed1998

Specificity improvement of a recombinant anti-testosterone Fab fragment by CDRIII mutagenesis and phage display selection.

The monoclonal antibodies so far developed by hybridoma technology have not had high enough specificity or affinity to distinguish the closely related steroid hormones in routine clinical assays. We have employed random mutagenesis and phage display approaches to improve the specificity of one anti-testosterone monoclonal antibody (3-C4F5). The affinity of the antibody is 0.3 x 10(9) M(-1) and the cross-reactivities with most of the related steroids are low. However, the antibody cross-reacts about 1% with dehydroepiandrosterone sulfate (DHEAS) and owing to the high DHEAS serum concentration this is about 1000-fold too high for clinical immunoassays. The complementarity-determining regions (CDRs) of the heavy and light chains, which were predicted by molecular modelling to be in close contact with the testosterone (TES) ligand, were randomized and mutant Fab libraries were cloned into a phagemid vector. Binders were selected by a competitive panning procedure. By combining the identified light and heavy chain CDRIII mutations the TES affinity was preserved at the wild-type level but DHEAS cross-reactivity was decreased to 0.03%. An important finding was that by the competitive panning procedure the overall binding specificity of the 3-C4F5 antibody was refined, since the cross-reactivities to related steroids were also significantly decreased in the combined mutant.

Amino Acid Sequence↗
PubMed1998

Peutz-Jeghers disease: most, but not all, families are compatible with linkage to 19p13.3.

A locus for Peutz-Jeghers syndrome (PJS) was recently mapped to chromosome 19p13.3. Each of 12 families studied was compatible with linkage to the marker D19S886. We have analysed 20 further families and found that the majority of these are consistent with a PJS gene on 19p13.3. Three families were, however, unlinked to 19p13.3 and none of the available PJS polyps from these families showed allele loss at D19S886. There were no obvious clinicopathological or ethnic differences between the 19p13.3 linked and unlinked families. There appears, therefore, to be a major PJS locus on chromosome 19p13.3 and the possibility exists of a minor locus (or loci) elsewhere.

Chromosomes, Human, Pair 19↗
PubMed1997

Exclusion of PTEN and 10q22-24 as the susceptibility locus for juvenile polyposis syndrome.

Juvenile polyposis syndrome (JPS; MIM 174900) is an autosomal dominant condition with incomplete penetrance characterized by hamartomatous polyps of the gastrointestinal tract and a risk of gastrointestinal cancer. Gastrointestinal hamartomatous polyps are also present in Cowden syndrome (CS; MIM 158350) and Bannayan-Zonana syndrome (BZS; also called Ruvalcaba-Myhre-Smith syndrome; MIM 153480). The susceptibility locus for both CS and BZS has recently been identified as the novel tumor suppressor gene PTEN, encoding a dual specificity phosphatase, located at 10q23.3. A putative JPS locus, JP1, which most likely functions as a tumor suppressor, had previously been mapped to 10q22-24 in both familial and sporadic juvenile polyps. Given the shared clinical features of gastrointestinal hamartomatous polyps among the three syndromes and the coincident mapping of JP1 to the region of PTEN, we sought to determine whether JPS was allelic to CS and BZS by mutation analysis of PTEN and linkage approaches. Microsatellite markers spanning the CS/BZS locus (D10S219, D10S551, D10S579, and D10S541) were used to compute multipoint lod scores in eight informative families with JPS. Lod scores of < -2.0 were generated for the entire region, thus excluding PTEN and any genes within the flanking 20-cM interval as candidate loci for familial JPS under our statistical models. In addition, analysis of PTEN using a combination of denaturing gradient gel electrophoresis and direct sequencing was unable to identify a germline mutation in 14 families with JPS and 11 sporadic cases. Therefore, at least a proportion of JPS cases are not caused by germline PTEN alteration or by an alternative locus at 10q22-24.

Chromosomes, Human, Pair 10↗
PubMed1997

Localization of a susceptibility locus for Peutz-Jeghers syndrome to 19p using comparative genomic hybridization and targeted linkage analysis.

Many human cancer susceptibility genes have been successfully mapped by genetic linkage studies. One that has so far eluded researchers is that for Peutz-Jeghers (P-J) syndrome, a condition characterized by intestinal hamartomatous polyposis and melanin spots of the lips, buccal mucosa and digits. A dramatically elevated risk of malignancy has also been documented. Gastrointestinal tumours as well as cancers of the breast, ovary, testis and uterine cervix appear to be overrepresented in families with this syndrome. The nature of hamartomatous polyps is equivicol. Hamartomas are usually considered histologically benign, but in the case of Peutz-Jeghers patients, there are reports of adenomatous and malignant changes in the polyps, and the possibility of a hamartoma-carcinoma sequence has been discussed. A search for a putative tumour suppressor locus was made using comparative genomic hybridization (CGH) of Peutz-Jeghers polyps, combined with loss of heterozygosity (LOH) study. Genetic linkage analysis in 12 families using markers from a deletion site demonstrated the presence of a high-penetrance locus in distal 19p with a multipoint lod score of 7.00 at marker D19S886 without evidence of genetic heterogeneity. The study demonstrates the power of CGH combined with LOH analysis in identifying putative tumour suppressor loci, and provides molecular evidence of malignant potential in hamartomas.

Adenocarcinoma↗
PubMed1996

Semiautomated assessment of loss of heterozygosity and replication error in tumors.

Loss of heterozygosity (LOH) and replication error (RER) are important phenomena in tumor development, with diagnostic and prognostic relevance. Therefore, screening for LOH and RER is a desirable first step in the molecular analysis of tumors. We used semiautomated procedures based on multicolor fluorescently labeled microsatellite markers and an automated sequencer for PCR amplification, electrophoresis of PCR products, and allele detection with a set of 16 microsatellites in 56 colorectal tumors. We improved existing software for computer-assisted assessment of LOH and RER. A comparison of these results with those of a conventional, radioactive technique and visual interpretation shows a high degree of correlation between the two methods. The detection rates of LOH and RER are similar to those reported previously. The main advantages of the semiautomated fluorescence-based typing are in the objective, observer-unrelated, easy, and rapid computer-based scoring, and the resulting quantitative assessment of RER.

Automation↗
PubMed1995

Introduction of lysine residues on the light chain constant domain improves the labelling properties of a recombinant Fab fragment.

Europium chelates provide a non-radioactive alternative for sensitive labelling of antibodies for diagnostic immunoassays. Lysine residues at antibody surfaces are ready targets for labelling by an isothiocyanate derivative of the europium chelate (Eu3+). Here the labelling efficiency of a recombinant anti-human alpha-fetoprotein (hAFP) Fab fragment has been improved by increasing its lysine content by protein engineering. Molecular modelling was used to identify three light chain constant domain surface arginine residues, R154, R187 and R210, which were mutated to lysine residues. The mutations did not influence the affinity of the lysine-enriched Fab fragment and its labelling efficiency was found to be approximately 40% higher than that of the wild-type Fab fragment. With low degree of labelling, the affinities of the two Fab fragments were identical and comparable with that of the original monoclonal anti-hAFP IgG. With a higher degree of labelling the affinities of both Fab fragments decreased more than that of the intact IgG since more lysine residues are available for labelling in the additional heavy chain constant domains of the larger molecule. Electrostatic adsorption and covalent immobilization of the Fab fragments were characterized by BIAcore and the lysine-enriched Fab fragment was found to be more efficiently immobilized to an activated carboxymethyl surface.

Amino Acid Sequence↗
PubMed1994

Reaction kinetics of alkyl epoxides with DNA and other nucleophiles.

1,2-Epoxy alkanes from C3 to C8 were reacted with DNA, deoxyguanosine and 4-(p-nitrobenzyl) pyridine (NBP). DNA was hydrolyzed at neutral pH to release 7-alkylguanines. The products were analyzed by HPLC. The epoxides reacted largely according to the chain length, shorter epoxides being more reactive. Substitutions through carbon 1 predominated. Reactivity with NBP was almost equal between the epoxides.

Alkanes↗
PubMed1994

Loss of the wild type MLH1 gene is a feature of hereditary nonpolyposis colorectal cancer.

The mechanism by which germline mutations of DNA mismatch repair genes cause susceptibility to tumour formation is not yet understood. Studies in vitro indicate that heterozygosity for these mutations, unlike homozygosity, does not affect mismatch repair. Surprisingly, no loss of heterozygosity at the predisposing loci has so far been described in hereditary nonpolyposis colorectal cancers. Here, we show that loss of heterozygosity (LOH) of markers within or adjacent to the MLH1 gene on chromosome 3p occurs nonrandomly in tumours from members of families in which the disease phenotype cosegregates with MLH1. In every informative case, the loss affects the wild type allele. These results suggest that DNA mismatch repair genes resemble tumour suppressor genes in that two hits are required to cause a phenotypic effect.

Base Sequence↗