8q24 risk alleles and prostate cancer in African-Barbadian men.

BACKGROUND: African American men (AA) exhibit a disproportionate share of prostate cancer (PRCA) incidence, morbidity, and mortality. Several genetic association studies have implicated select 8q24 loci in PRCA risk in AA. The objective of this investigation is to evaluate the association between previously reported 8q24 risk alleles and PRCA in African-Barbadian (AB) men known to have high rates of PRCA. METHODS: Ten previously reported candidate tag SNPs were genotyped and/or imputed in the 8q24 region in 532 AB men with PRCA and 513 AB controls from the Prostate Cancer in a Black Population (PCBP) study. RESULTS: Rs2124036 was significant in AB men, (OR = 2.7, 95% CI (1.3-5.3), P = 0.005, Empirical (max (T), corrected for multiple testing) P = 0.03) for the homozygous C/C genotype. Only a single SNP from this region remained statistically significant in our analysis of our AB population. These results may indicate the presence of a founder effect or due to the chosen SNPs not tagging an ancestral haplotype bearing the 8q24 risk allele(s) in this population or could reflect inadequate power to detect an association. We conducted a meta-analysis including our AB population along with two additional African Caribbean populations from Tobago and Jamaica for SNPs rs16901979 and rs1447295. Meta-analysis results were most significant for rs16901979 A allele (Z score 2.73; P = 0.006) with a summary OR = 1.31 (95% CI: 1.09-1.58). CONCLUSIONS: Additional studies are needed to provide deeper genotype coverage to further interrogate the 8q24 region to understand its contribution to PRCA in this population.

Identification of a novel NBN truncating mutation in a family with hereditary prostate cancer.

Nibrin (NBN), located on chromosome 8q21 is a gene involved in DNA double-strand break repair that has been implicated in the rare autosomal recessive chromosomal instability syndrome known as Nijmegen Breakage Syndrome (NBS). NBS is characterized by specific physical characteristics (microcephaly and dysmorphic facies), immunodeficiency, and increased risk of malignancy. Individuals who are heterozygous for NBN mutations are clinically asymptomatic, but may display an elevated risk for certain cancers including, but not limited to, ovarian and prostate cancer as well as various lymphoid malignancies. In this study, 94 unrelated familial prostate cancer cases from the University of Michigan Prostate Cancer Genetics Project (n = 54) and Johns Hopkins University (n = 40) were subjected to targeted next-generation sequencing of the exons, including UTRs, of NBN. One individual of European descent, diagnosed with prostate cancer at age 52, was identified to have a heterozygous 2117 C > G mutation in exon 14 of the gene, that results in a premature stop at codon 706 (S706X). Sequencing of germline DNA from additional male relatives showed partial co-segregation of the NBN S706X mutation with prostate cancer. This NBN mutation was not observed among 2768 unrelated European men (1859 with prostate cancer and 909 controls). NBN is involved in double-strand break repair as a component of the MRE11 (meiotic recombination 11)/RAD50/NBN genomic stability complex. The S706X mutation truncates the protein in a highly conserved region of NBN near the MRE11 binding site, thus suggesting a role for rare NBN mutations in prostate cancer susceptibility.

Germline mutations in HOXB13 and prostate-cancer risk.

BACKGROUND: Family history is a significant risk factor for prostate cancer, although the molecular basis for this association is poorly understood. Linkage studies have implicated chromosome 17q21-22 as a possible location of a prostate-cancer susceptibility gene. METHODS: We screened more than 200 genes in the 17q21-22 region by sequencing germline DNA from 94 unrelated patients with prostate cancer from families selected for linkage to the candidate region. We tested family members, additional case subjects, and control subjects to characterize the frequency of the identified mutations. RESULTS: Probands from four families were discovered to have a rare but recurrent mutation (G84E) in HOXB13 (rs138213197), a homeobox transcription factor gene that is important in prostate development. All 18 men with prostate cancer and available DNA in these four families carried the mutation. The carrier rate of the G84E mutation was increased by a factor of approximately 20 in 5083 unrelated subjects of European descent who had prostate cancer, with the mutation found in 72 subjects (1.4%), as compared with 1 in 1401 control subjects (0.1%) (P=8.5x10(-7)). The mutation was significantly more common in men with early-onset, familial prostate cancer (3.1%) than in those with late-onset, nonfamilial prostate cancer (0.6%) (P=2.0x10(-6)). CONCLUSIONS: The novel HOXB13 G84E variant is associated with a significantly increased risk of hereditary prostate cancer. Although the variant accounts for a small fraction of all prostate cancers, this finding has implications for prostate-cancer risk assessment and may provide new mechanistic insights into this common cancer. (Funded by the National Institutes of Health and others.).

EphB2 SNPs and sporadic prostate cancer risk in African American men.

The EphB2 gene has been implicated as a tumor suppressor gene somatically altered in both prostate cancer (PC) and colorectal cancer. We have previously shown an association between an EphB2 germline nonsense variant and risk of familial prostate cancer among African American Men (AAM). Here we set out to test the hypothesis that common variation within the EphB2 locus is associated with increased risk of sporadic PC in AAM. We genotyped a set of 341 single nucleotide polymorphisms (SNPs) encompassing the EphB2 locus, including known and novel coding and noncoding variants, in 490 AA sporadic PC cases and 567 matched controls. Single marker-based logistical regression analyses revealed seven EphB2 SNPs showing statistically significant association with prostate cancer risk in our population. The most significant association was achieved for a novel synonymous coding SNP, TGen-624, (Odds Ratio (OR)  = 0.22; 95% Confidence Interval (CI) 0.08-0.66, p = 1×10(-5)). Two other SNPs also show significant associations toward a protective effect rs10465543 and rs12090415 (p = 1×10(-4)), OR = 0.49 and 0.7, respectively. Two additional SNPs revealed trends towards an increase in risk of prostate cancer, rs4612601 and rs4263970 (p = 0.001), OR = 1.35 and 1.31, respectively. Furthermore, haplotype analysis revealed low levels of linkage disequilibrium within the region, with two blocks being associated with prostate cancer risk among our population. These data suggest that genetic variation at the EphB2 locus may increase risk of sporadic PC among AAM.

Copy number and targeted mutational analysis reveals novel somatic events in metastatic prostate tumors.

Advanced prostate cancer can progress to systemic metastatic tumors, which are generally androgen insensitive and ultimately lethal. Here, we report a comprehensive genomic survey for somatic events in systemic metastatic prostate tumors using both high-resolution copy number analysis and targeted mutational survey of 3508 exons from 577 cancer-related genes using next generation sequencing. Focal homozygous deletions were detected at 8p22, 10q23.31, 13q13.1, 13q14.11, and 13q14.12. Key genes mapping within these deleted regions include PTEN, BRCA2, C13ORF15, and SIAH3. Focal high-level amplifications were detected at 5p13.2-p12, 14q21.1, 7q22.1, and Xq12. Key amplified genes mapping within these regions include SKP2, FOXA1, and AR. Furthermore, targeted mutational analysis of normal-tumor pairs has identified somatic mutations in genes known to be associated with prostate cancer including AR and TP53, but has also revealed novel somatic point mutations in genes including MTOR, BRCA2, ARHGEF12, and CHD5. Finally, in one patient where multiple independent metastatic tumors were available, we show common and divergent somatic alterations that occur at both the copy number and point mutation level, supporting a model for a common clonal progenitor with metastatic tumor-specific divergence. Our study represents a deep genomic analysis of advanced metastatic prostate tumors and has revealed candidate somatic alterations, possibly contributing to lethal prostate cancer.

Replication of prostate cancer risk loci on 8q24, 11q13, 17q12, 19q33, and Xp11 in African Americans.

BACKGROUND: Prostate cancer (Pca) is a common malignancy that disproportionately affects African American men (AA). Recently there have been several genome-wide association studies (GWAS) implicating new prostate cancer risk loci along chromosomes 2, 3, 6, 7, 8, 10, 11, 12, 17, 19, and X in populations of European ancestry. Given the higher incidence and mortality for AAs, and differences in allele frequencies and haplotype structures between African and European descent populations, it is important to assess the impact of these candidate risk loci in AAs. METHODS: Here we evaluated 20 single nucleotide polymorphisms (SNPs) associated with prostate cancer risk in recent GWAS studies, in AA prostate cancer cases and controls. RESULTS: We replicated five of the SNPs in our AA population, rs10896449 on 11q13.2 (P = 0.009), rs2735839 on 19q33.33 region, (P = 0.04), rs443076 on chromosome 17q12 (P = 0.008), rs5945572 on Xp11.22 (P = 0.05), as well as the rare variant specific to west African ancestry, bd11934905 in region 2 of 8q24 (P = 1 x 10(-4)). CONCLUSIONS: While we were able to replicate a few of the previous GWAS SNPs, we were not able to confirm the vast majority of these associations in our AA population. This finding further supports the need to perform GWAS and additional fine mapping in AAs to locate additional susceptibility loci.

Association of HPC2/ELAC2 and RNASEL non-synonymous variants with prostate cancer risk in African American familial and sporadic cases.

INTRODUCTION: The RNASEL and HPC2/ELAC2 genes have been implicated in hereditary prostate cancer. Further assessment of the role of these genes in sporadic prostate cancer in African American men (AAM) is warranted. METHODS: Genotyping of HPC2/ELAC2 variants (S217L, A541T), along with RNASEL variants (R462Q and E541D) was completed in 155 African American sporadic and 88 familial prostate cancer cases, and 296 healthy male controls. Logistic regression analysis was performed and odds ratios (OR) were calculated, while correcting for both age and population stratification using admixture informative markers. RESULTS: The HPC2/ELAC2 217L allele was significantly associated with risk of prostate cancer when taking all cases into account (OR = 1.6; 1.0-2.6; P = 0.03). The RNASEL 541D allele was associated with a decrease in risk of prostate cancer in sporadic cases (OR = 0.4; 0.2-0.8; P = 0.01). We did not detect an association between prostate cancer risk and the RNASEL R462Q variant. Results from haplotype analyses of the two RNASEL variants revealed highly significant differences in haplotype allele frequencies between cases and controls suggesting a synergistic effect at the RNASEL locus. One haplotype in particular (462R-541D) is far more frequent in our control population and shows a strong protective effect against prostate cancer (OR = 0.47, P = 8.1 x 10(-9)). CONCLUSIONS: These results suggest that HPC2/ELAC2 and RNASEL may play a role, however minor, in prostate cancer risk among AAM.

PI3K/AKT pathway activation in acute myeloid leukaemias is not associated with AKT1 pleckstrin homology domain mutation.

Despite its' central role, the precise mechanisms of the phosphoinositide 3-kinase/Akt (PI3K)/Akt pathway activation in acute myeloid leukaemia (AML) have not been elucidated. Recently, a recurrent novel AKT1 pleckstrin homology domain (PHD) mutation leading to membrane translocation, constitutive AKT activation and leukaemia development in mice was described. To assess AKT1 PHD mutations in AML, we sequenced 57 specimens from 49 AML patients, all of whom showed PI3K/AKT pathway activation by analysis of total and phospho-protein expression for AKT, mTor, p70S6Kinase, S6ribosomal protein and PTEN. No mutations in AKT1 PHD were identified, making this mutation an unlikely cause of PI3K/AKT pathway activation in AML.

Two genome-wide association studies of aggressive prostate cancer implicate putative prostate tumor suppressor gene DAB2IP.

BACKGROUND: The consistent finding of a genetic susceptibility to prostate cancer suggests that there are germline sequence variants predisposing individuals to this disease. These variants could be useful in screening and treatment. METHODS: We performed an exploratory genome-wide association scan in 498 men with aggressive prostate cancer and 494 control subjects selected from a population-based case-control study in Sweden. We combined the results of this scan with those for aggressive prostate cancer from the publicly available Cancer Genetic Markers of Susceptibility (CGEMS) Study. Single-nucleotide polymorphisms (SNPs) that showed statistically significant associations with the risk of aggressive prostate cancer based on two-sided allele tests were tested for their association with aggressive prostate cancer in two independent study populations composed of individuals of European or African American descent using one-sided tests and the genetic model (dominant or additive) associated with the lowest value in the exploratory study. RESULTS: Among the approximately 60,000 SNPs that were common to our study and CGEMS, we identified seven that had a similar (positive or negative) and statistically significant (P<.01) association with the risk of aggressive prostate cancer in both studies. Analysis of the distribution of these SNPs among 1032 prostate cancer patients and 571 control subjects of European descent indicated that one, rs1571801, located in the DAB2IP gene, which encodes a novel Ras GTPase-activating protein and putative prostate tumor suppressor, was associated with aggressive prostate cancer (one-sided P value = .004). The association was also statistically significant in an African American study population that included 210 prostate cancer patients and 346 control subjects (one-sided P value = .02). CONCLUSION: A genetic variant in DAB2IP may be associated with the risk of aggressive prostate cancer and should be evaluated further.

Confirmation study of prostate cancer risk variants at 8q24 in African Americans identifies a novel risk locus.

Prostate cancer is a common complex disease that disproportionately affects men of African descent. Recently, several different common variants on chromosome 8q24 have been shown to be associated with prostate cancer in multiple studies and ethnic groups. The objective of this study was to confirm the association of 8q24 markers with prostate cancer in African Americans. We genotyped 24 markers along 8q24 and 80 unlinked ancestry informative markers in a hospital-based case-control sample of 1057 African American men (490 prostate cancer cases and 567 controls). Association analyses of 8q24 markers with prostate cancer risk were adjusted for both global and local 8q24 admixture stratification using estimates from ancestry informative markers. We report that rs7008482, which maps to the 8q24.13 region, is an additional independent prostate cancer risk variant (P = 5 x 10(-4)), and we also replicate the association of rs16901979 with prostate cancer (P = 0.002). Other published risk variants in the region such as rs1447295 and rs6983267 showed a similar direction and magnitude of effect, but were not significant in our population. Both rs7008482 and rs16901979 independently predicted risk and remained significant (P < 0.001) after controlling for each other. Our data combined with additional replications of 8q24 markers provide compelling support for multiple regions of risk for prostate cancer on 8q24.

A transforming mutation in the pleckstrin homology domain of AKT1 in cancer.

Although AKT1 (v-akt murine thymoma viral oncogene homologue 1) kinase is a central member of possibly the most frequently activated proliferation and survival pathway in cancer, mutation of AKT1 has not been widely reported. Here we report the identification of a somatic mutation in human breast, colorectal and ovarian cancers that results in a glutamic acid to lysine substitution at amino acid 17 (E17K) in the lipid-binding pocket of AKT1. Lys 17 alters the electrostatic interactions of the pocket and forms new hydrogen bonds with a phosphoinositide ligand. This mutation activates AKT1 by means of pathological localization to the plasma membrane, stimulates downstream signalling, transforms cells and induces leukaemia in mice. This mechanism indicates a direct role of AKT1 in human cancer, and adds to the known genetic alterations that promote oncogenesis through the phosphatidylinositol-3-OH kinase/AKT pathway. Furthermore, the E17K substitution decreases the sensitivity to an allosteric kinase inhibitor, so this mutation may have important clinical utility for AKT drug development.

Genome-wide linkage of 77 families from the African American Hereditary Prostate Cancer study (AAHPC).

BACKGROUND: The African American Hereditary Prostate Cancer (AAHPC) Study was designed to recruit families with early-onset disease fulfilling criteria of >or=4 affected. METHODS: We present a approximately 10 cM genome-wide linkage (GWL) analysis on 77 families including 254 affected and 274 unaffected genotyped. RESULTS: Linkage analysis revealed three chromosomal regions with GENEHUNTER multipoint HLOD scores >or=1.3 for all 77 families at 11q22, 17p11, and Xq21. One family yielded genome-wide significant evidence of linkage (LOD = 3.5) to the 17p11 region with seven other families >or=2.3 in this region. Twenty-nine families with no-male-to-male (MM) transmission gave a peak HLOD of 1.62 (alpha = 0.33) at the Xq21 locus. Two novel peaks >or=0.91 for the 16 families with '>6 affected' occurred at 2p21 and 22q12. CONCLUSIONS: These chromosomal regions in the genome warrant further follow-up based on the hypothesis of multiple susceptibility genes with modest effects, or several major genes segregating in small subsets of families.

Fine-mapping the putative chromosome 17q21-22 prostate cancer susceptibility gene to a 10 cM region based on linkage analysis.

Prostate cancer linkage studies have suggested the existence of a prostate cancer susceptibility gene on chromosome 17q21-22. We now report the results of an extended linkage analysis including 95 new multiplex prostate cancer families and 9 additional microsatellite markers resulting in a maximum LOD score of 2.99 at approximately 81-82 cM for all 453 pedigrees. Results from these 95 new pedigrees provide additional support for a chromosome 17q21-22 prostate cancer susceptibility gene. Inclusion of the 9 additional markers significantly reduced the size of the candidate region, as defined using a 1-LOD support interval, especially when focusing analyses on subsets of pedigrees with four or more confirmed affecteds or average age of diagnosis less than or equal to 65 years. A novel subset analysis of only those families (n = 147) that had four or more prostate cancer cases and an average age of prostate cancer diagnosis < or = 65 years results in a maximum LOD score of 5.49 at 78 cM with a 1-LOD support interval of 10 cM. This large set of pedigrees with four more prostate cancer cases characterized by early-onset disease will serve as a useful resource for identifying the putative 17q21-22 prostate cancer susceptibility gene.

Nonsense-mediated decay microarray analysis identifies mutations of EPHB2 in human prostate cancer.

The identification of tumor-suppressor genes in solid tumors by classical cancer genetics methods is difficult and slow. We combined nonsense-mediated RNA decay microarrays and array-based comparative genomic hybridization for the genome-wide identification of genes with biallelic inactivation involving nonsense mutations and loss of the wild-type allele. This approach enabled us to identify previously unknown mutations in the receptor tyrosine kinase gene EPHB2. The DU 145 prostate cancer cell line, originating from a brain metastasis, carries a truncating mutation of EPHB2 and a deletion of the remaining allele. Additional frameshift, splice site, missense and nonsense mutations are present in clinical prostate cancer samples. Transfection of DU 145 cells, which lack functional EphB2, with wild-type EPHB2 suppresses clonogenic growth. Taken together with studies indicating that EphB2 may have an essential role in cell migration and maintenance of normal tissue architecture, our findings suggest that mutational inactivation of EPHB2 may be important in the progression and metastasis of prostate cancer.

Familial isolated hyperparathyroidism is rarely caused by germline mutation in HRPT2, the gene for the hyperparathyroidism-jaw tumor syndrome.

Familial isolated hyperparathyroidism (FIHP) can result occasionally from the incomplete expression of a syndromic form of familial hyperparathyroidism (HPT), specifically multiple endocrine neoplasia type 1 (MEN1), familial hypocalciuric hypercalcemia, or the hyperparathyroidism-jaw tumor syndrome (HPT-JT). The cause of FIHP has not been identified in the majority of families. We investigated 32 families with FIHP to determine the frequency of occult mutation in HRPT2, the gene causing HPT-JT. All families had negative clinical testing for MEN1, hypocalciuric hypercalcemia, and HPT-JT and negative mutational screening of MEN1 and CASR, the gene for the calcium-sensing receptor. Thus, an extended effort was made to exclude each of the principal syndromic causes of FIHP. The families were characterized by young probands (42 +/- 3 yr) and occasionally unusual parathyroid histology, including four families with one case of parathyroid cancer. We had speculated that there was a high frequency of occult mutation in HRPT2 among such carefully screened kindreds. This hypothesis became testable with the recent identification of that gene. Among the 32 FIHP families, only a single one was found to have a mutation in HRPT2 (679insAG); this mutation predicts premature truncation of its gene product, parafibromin, and thus its presumed inactivation. Even accounting for families with one of the three occult syndromes and false negative biochemical or DNA testing, these results indicate that an unexpectedly large fraction of FIHP has currently unrecognized causes.

Mutational analysis of susceptibility genes RNASEL/HPC1, ELAC2/HPC2, and MSR1 in sporadic prostate cancer.

Three putative prostate cancer-susceptibility genes, RNASEL/HPC1 at 1q24, MSR1 at 8p22, and ELAC2/HPC2 at 17p11, have recently been identified. Our objective was to investigate somatic mutations in these genes in sporadic prostate cancer. We analyzed 39 clinical prostate cancer specimens, 10 prostate cancer xenografts (LuCaP series), and 4 prostate cancer cell lines (LNCaP, DU145, PC-3, and MPC-3) for genetic changes using denaturing high-performance liquid chromatography and direct sequencing in order to screen the whole coding regions of RNASEL and MSR1, as well as exons 7 and 17 of ELAC2. The known 471delAAAG truncating mutation was found in the RNASEL gene in cell line LNCaP. The only new missense variation in RNASEL, Gly296Val, was found in cell line DU145, but not in any other samples. RNASEL and ELAC2 also showed the common missense polymorphic changes. A previously reported truncating mutation (Arg293X) was found in MSR1 in the germ line of one individual. Our results indicate that inactivation of the RNASEL, ELAC2, or MSR1 genes by somatic mutation is a rare phenomenon in sporadic prostate cancer.

Somatic and germ-line mutations of the HRPT2 gene in sporadic parathyroid carcinoma.

BACKGROUND: We looked for mutations of the HRPT2 gene, which encodes the parafibromin protein, in sporadic parathyroid carcinoma because germ-line inactivating HRPT2 mutations have been found in a type of familial hyperparathyroidism--hyperparathyroidism-jaw tumor (HPT-JT) syndrome--that carries an increased risk of parathyroid cancer. METHODS: We directly sequenced the full coding and flanking splice-junctional regions of the HRPT2 gene in 21 parathyroid carcinomas from 15 patients who had no known family history of primary hyperparathyroidism or the HPT-JT syndrome at presentation. We also sought to confirm the somatic nature of the identified mutations and tested the carcinomas for tumor-specific loss of heterozygosity at HRPT2. RESULTS: Parathyroid carcinomas from 10 of the 15 patients had HRPT2 mutations, all of which were predicted to inactivate the encoded parafibromin protein. Two distinct HRPT2 mutations were found in tumors from five patients, and biallelic inactivation as a result of a mutation and loss of heterozygosity was found in one tumor. At least one HRPT2 mutation was demonstrably somatic in carcinomas from six patients. Unexpectedly, HRPT2 mutations in the parathyroid carcinomas of three patients were identified as germ-line mutations. CONCLUSIONS: Sporadic parathyroid carcinomas frequently have HRPT2 mutations that are likely to be of pathogenetic importance. Certain patients with apparently sporadic parathyroid carcinoma carry germ-line mutations in HRPT2 and may have the HPT-JT syndrome or a phenotypic variant.

Identification and characterization of mouse Rab32 by mRNA and protein expression analysis.

Rab proteins, a subfamily of the ras superfamily, are low molecular weight GTPases involved in the regulation of intracellular vesicular transport. Cloning of human RAB32 was recently described. Presently, we report the cloning and characterization of the mouse homologue of Rab32. We show that murine Rab32 exhibits a ubiquitous expression pattern, with tissue-specific variation in expression level. Three cell types with highly specialized organelles, melanocytes, platelets and mast cells, exhibit relatively high level of Rab32. We show that in murine amelanotic in vitro transformed melanocytes as well as in human amelanotic metastatic melanoma cell lines, the expression of Rab32 is markedly reduced or absent, in parallel with the loss of expression of two key enzymes for the production of melanin, tyrosinase and Tyrp1. Therefore, in both mouse and human systems, the expression of Rab32 correlates with the expression of genes involved in pigment production. However, in melanoma samples, amelanotic due to a mutation in the tyrosinase gene, the expression of Rab32 remains at levels comparable to those observed in pigmented melanoma samples. Finally, we observed co-localization of Rab32 and the melanosomal proteins, Tyrp1 and Dct, indicating an association of Rab32 with melanosomes. Based on these data, we propose the inclusion of Rab32 to the so-called melanocyte/platelet family of Rab proteins.

Cloning and characterization of a novel gene, SHPRH, encoding a conserved putative protein with SNF2/helicase and PHD-finger domains from the 6q24 region.

Here we report the identification of a novel transcript containing SNF2, PHD-finger, RING-finger, helicase, and linker histone domains mapping to the q24 band region of human chromosome 6. These domains are characteristic of several DNA repair proteins, transcription factors, and helicases. We have cloned both human and mouse homologs of this novel gene using interexon PCR and RACE technologies. The human cDNA, termed SHPRH, is 6018 bp and codes for a putative protein of 1683 amino acids. The mouse cDNA, termed Shprh, is 7225 bp and codes for a putative protein of 1616 amino acids. The deduced amino acid sequences of the two proteins share 86% identity. Both genes are expressed ubiquitously, with a transcript size of approximately 7.5 kb. Mapping of this gene to 6q24, a region reported to contain a tumor suppressor locus, prompted us to evaluate SHPRH by mutation analysis in tumor cell lines. We have identified one truncating and three missense mutations, thus suggesting SHPRH as a possible candidate for the tumor suppressor gene.

Melanoma mouse model implicates metabotropic glutamate signaling in melanocytic neoplasia.

To gain insight into melanoma pathogenesis, we characterized an insertional mouse mutant, TG3, that is predisposed to develop multiple melanomas. Physical mapping identified multiple tandem insertions of the transgene into intron 3 of Grm1 (encoding metabotropic glutamate receptor 1) with concomitant deletion of 70 kb of intronic sequence. To assess whether this insertional mutagenesis event results in alteration of transcriptional regulation, we analyzed Grm1 and two flanking genes for aberrant expression in melanomas from TG3 mice. We observed aberrant expression of only Grm1. Although we did not detect its expression in normal mouse melanocytes, Grm1 was ectopically expressed in the melanomas from TG3 mice. To confirm the involvement of Grm1 in melanocytic neoplasia, we created an additional transgenic line with Grm1 expression driven by the dopachrome tautomerase promoter. Similar to the original TG3, the Tg(Grm1)EPv line was susceptible to melanoma. In contrast to human melanoma, these transgenic mice had a generalized hyperproliferation of melanocytes with limited transformation to fully malignant metastasis. We detected expression of GRM1 in a number of human melanoma biopsies and cell lines but not in benign nevi and melanocytes. This study provides compelling evidence for the importance of metabotropic glutamate signaling in melanocytic neoplasia.