Larisa V. Debelenko

The Insulin-like Growth Factor-1 Receptor–Targeting Antibody, CP-751,871, Suppresses Tumor-Derived VEGF and Synergizes with Rapamycin in Models of Childhood Sarcoma

Signaling through the type 1 insulin-like growth factor receptor (IGF-1R) occurs in many human cancers, including childhood sarcomas. As a consequence, targeting the IGF-1R has become a focus for cancer drug development. We examined the antitumor activity of CP-751,871, a human antibody that blocks IGF-1R ligand binding, alone and in combination with rapamycin against sarcoma cell lines in vitro and xenograft models in vivo. In Ewing sarcoma (EWS) cell lines, CP751,871 inhibited growth poorly (<50%), but prevented rapamycin-induced hyperphosphorylation of AKT(Ser473) and induced greater than additive apoptosis. Rapamycin treatment also increased secretion of IGF-1 resulting in phosphorylation of IGF-1R (Tyr1131) that was blocked by CP751,871. In vivo CP-751,871, rapamycin, or the combination were evaluated against EWS, osteosarcoma, and rhabdomyosarcoma xenografts. CP751871 induced significant growth inhibition [EFS(T/C) >2] in four models. Rapamycin induced significant growth inhibition [EFS(T/C) >2] in nine models. Although neither agent given alone caused tumor regressions, in combination, these agents had greater than additive activity against 5 of 13 xenografts and induced complete remissions in one model each of rhabdomyosarcoma and EWS, and in three of four osteosarcoma models. CP751,871 caused complete IGF-1R down-regulation, suppression of AKT phosphorylation, and dramatically suppressed tumor-derived vascular endothelial growth factor (VEGF) in some sarcoma xenografts. Rapamycin treatment did not markedly suppress VEGF in tumors and synergized only in tumor lines where VEGF was dramatically inhibited by CP751,871. These data suggest a model in which blockade of IGF-1R suppresses tumor-derived VEGF to a level where rapamycin can effectively suppress the response in vascular endothelial cells.

The multiple endocrine neoplasia type I gene locus is involved in the pathogenesis of type II gastric carcinoids

BACKGROUND & AIMS Both gastrin and genetic factors were suggested to underlie the pathogenesis of multiple gastric enterochromaffin-like (ECL) cell carcinoids. To assess the role of genetic alterations in carcinoid tumorigenesis, loss of heterozygosity (LOH) at the locus of the multiple endocrine neoplasia type 1 (MEN-1) gene was studied in gastric carcinoids of patients with MEN-1 and chronic atrophic type A gastritis (A-CAG), as well as in sporadically arising intestinal carcinoids. METHODS DNA extracted from archival tissue sections of 35 carcinoid tumors was assessed for LOH with eight polymorphic markers on chromosome 11q13. A combined tumor and family study was performed in 1 patient with MEN-1-Zollinger-Ellison syndrome (ZES). RESULTS LOH at 11q13 loci was detected in 15 of 20 (75%) MEN-1-ZES carcinoids, and each ECL-cell carcinoid with LOH showed deletion of the wild-type allele. Only 1 of 6 A-CAG carcinoids displayed LOH at the MEN-1 gene locus, and none of the 9 intestinal and rectal carcinoids showed 11q13 LOH. CONCLUSIONS Gastric ECL-cell carcinoid is an independent tumor type of MEN-1 that shares a common developmental mechanism (via inactivation of the MEN-1 gene) with enteropancreatic and parathyroid MEN-1 tumors. Further analysis of sporadic and A-CAG carcinoids is needed to elucidate genetic factors involved in their tumorigenesis.

MEN1 gene mutation analysis of high‐grade neuroendocrine lung carcinoma

Neuroendocrine tumors of the lung consist of a spectrum of neoplasms, including typical carcinoids, atypical carcinoids, large‐cell neuroendocrine carcinomas (LCNEC), and small‐cell lung carcinomas (SCLC). We previously reported frequent inactivation of the gene responsible for multiple endocrine neoplasia type 1 (MEN1) in both typical and atypical carcinoid tumors. In the present study, we extend the analysis of human NE lung tumors to include 9 primary SCLCs, 36 SCLC cell lines, and 13 primary LCNECs for MEN1 gene inactivation. In SCLC, loss of heterozygosity (LOH) at the MEN1 gene on chromosome band 11q13 was detected in one primary tumor and two cell lines. The coding sequence and splice junctions of the MEN1 gene were screened for mutations in all 44 tumors and cell lines, and no mutations were detected. Northern blot analysis of 13 SCLC cell lines showed the MEN1 transcript to be present and of normal size. In LCNECs, a somatic frameshift in the MEN1 gene (1226delC) was found in one of 13 tumors, representing the first mutation observed outside the spectrum of neoplasms associated with MEN1. Interestingly, neither a deletion nor a mutation was detected in the other allele, and wild‐type mRNA sequence was expressed in the tumor, suggesting that the MEN1 gene was not inactivated by a conventional two‐hit mechanism. The data support the hypothesis that SCLC and lung carcinoids develop via distinct molecular pathways; however, further investigation is necessary to determine the significance of the MEN1 gene mutation observed in a single case of LCNEC. Genes Chromosomes Cancer 28:58–65, 2000. Published 2000 Wiley‐Liss, Inc.

Eighteen new polymorphic markers in the multiple endocrine neoplasia type 1 (MEN1) region

Abstract Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder in which affected individuals develop tumors primarily in the parathyroids, anterior pituitary, endocrine pancreas, and duodenum. The locus for MEN1 is tightly linked to the marker PYGM on chromosome 11q13, and linkage analysis has previously placed the MEN1 gene within a 2-Mb interval flanked by markers D11S1883 and D11S449. Loss of heterozygosity (LOH) studies in MEN1 and sporadic tumors have helped narrow the location of the gene to a 600-kb interval between PYGM and D11S449. Eighteen new polymerase chain reaction (PCR)-based polymorphic markers were generated for the MEN1 region, with ten mapping to the PYGM-D11S449 interval. These new markers, along with 14 previously known polymorphic markers, were precisely mapped on a 2.8-Mb (D11S480–D11S913) high-density clone contig-based, physical map generated for the MEN1 region.

Germline and somatic mutation of the gene for multiple endocrine neoplasia type 1 (MEN1)

Marx SJ, Agarwal SK, Kester MB, Heppner C, Kim YS, Emmert‐Buck MR, Debelenko LV, Lubensky IA, Zhuang Z, Guru SC, Manickam P, Olufemi SE, Skarulis MC, Doppman JL, Alexander RH, Liotta LA, Collins FS, Chandrasekharappa SC, Spiegel AM, Burns AL (National Institutes of Health, Bethesda, USA). Germline and somatic mutation of the gene for multiple endocrine neoplasia type 1 (MEN1) (Minisymposium: MEN & VHL). J Intern Med 1998; 243: 447–53.

The gene for multiple endocrine neoplasia type 1: recent findings

Multiple endocrine neoplasia type 1 (MENI) is a promising model to understand endocrine and other tumors. Its most common endocrine expressions are tumors of parathyroids, entero-pancreatic neuro-endocrine tissue, and anterior pituitary. Recently, collagenomas and multiple angiofibromas of the dermis also have been recognized as very common. MEN1 can be characterized from different perspectives: (a) as a hormone (parathyroid hormone, gastrin, prolactin, etc.) excess syndrome with excellent therapeutic options; (b) as a syndrome with sometimes lethal outcomes from malignancy of entero-pancreatic neuro-endocrine or foregut carcinoid tissues; or (c) as a disorder than can give insight about cell regulation in the endocrine, the dermal, and perhaps other tissue systems. The MEN1 gene was identified recently by positional cloning, a comprehensive strategy of narrowing the candidate interval and evaluating all or most genes in that interval. This discovery has opened new approaches to basic and clinical issues. Germline MEN1 mutations have been identified in most MEN1 families. Germline MENI mutations were generally not found in families with isolated hyperparathyroidism or with isolated pituitary tumor. Thus, studies with the MENI gene helped establish that mutation of other gene(s) is likely causative of these two MEN1 phenocopies. MEN1 proved to be the gene most frequent L4 mutated in common-variety, nonhereditary parathyroid tumor, gastrinoma, insulinoma, or bronchial carcinoid. For example, in common-variety parathyroid tumors, mutation of several other genes (such as cyclin D1 and P53) has been found, but much less frequently than MEN1 mutation. The majority of germline and somatic MEN1 mutations predicted truncation of the encoded protein (menin). Such inactivating mutations strongly supported prior predictions that MEN1 is a tumor suppressor gene insofar as stepwise mutational inactivation of both copies can release a cell from normal growth suppression. Menin is principally a nuclear protein; menin interacts with junD. Future studies, such as discovery of menins metabolic pathway, could lead to new opportunities in cell biology and in tumor therapy.

Analysis of recurrent germline mutations in the MEN1 gene encountered in apparently unrelated families.

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder that manifests as varying combinations of tumors of endocrine and other tissues (parathyroids, pancreatic islets, duodenal endocrine cells, the anterior pituitary and others). The MEN1 gene is on chromosome 11q13; it was recently identified by positional cloning. We previously reported 32 different germline mutations in 47 of the 50 familial MEN1 probands studied at the NIH. Eight different germline MEN1 mutations were encountered repeatedly in two or more apparently unrelated families. We analyzed the haplotypes of families with recurrent MEN1 mutations with seven polymorphic markers in the 11q13 region surrounding the MEN1 gene (from D11S1883 to D11S4908). Disease haplotypes were inferred from germline DNA and also from tumors with 11q13 loss of heterozygosity. Two different disease haplotype cores were shared by apparently unrelated families for two mutations in exon 2 (five families with 416delC and six families with 512delC). These two repeat mutations were associated with the two founder effects that we reported in a prior haplotype analysis. The disease haplotypes for each of the other six repeat mutations (seen twice each) were discordant, suggesting independent origins of these recurrent mutations. Most of the MEN1 germline mutations including all of those recurring independently occur in regions of CpG/CpNpG, short DNA repeats or single nucleotide repeat motifs. In conclusion, recurring germline mutations account for about half of the mutations in North American MEN1 families. They result from either founder effects or independent occurrence of one mutation more than one time. Hum Mutat 12:75–82, 1998. Published 1998 Wiley‐Liss, Inc. This article is a US Government work and, as such, is in the public domain in the United States of America.