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Biomedical subjects

W Arap

Publications and source records attributed to W Arap.

33 records · Page 2Linked to original sources
PubMed1999

Anti-cancer activity of targeted pro-apoptotic peptides.

We have designed short peptides composed of two functional domains, one a tumor blood vessel 'homing' motif and the other a programmed cell death-inducing sequence, and synthesized them by simple peptide chemistry. The 'homing' domain was designed to guide the peptide to targeted cells and allow its internalization. The pro-apoptotic domain was designed to be nontoxic outside cells, but toxic when internalized into targeted cells by the disruption of mitochondrial membranes. Although our prototypes contain only 21 and 26 residues, they were selectively toxic to angiogenic endothelial cells and showed anti-cancer activity in mice. This approach may yield new therapeutic agents.

Amino Acid Sequence↗
PubMed1999

Identification of receptor ligands with phage display peptide libraries.

With the development and maturation of the technology of displaying peptides on bacteriophage, it has become possible to isolate peptide ligands to various targets. In the phage display strategy, up to 10(9) peptides of different permutations are expressed on the surface of filamentous phage. Thus, peptides capable of binding target molecules in vitro and even target tissues in vivo can be identified. In recent years, a series of libraries that display degenerate peptides of different lengths have been constructed, and specific ligands to cell surface receptors, such as integrins, have been isolated. In the in vivo biopanning, peptides targeting distinct organs or tumors have been rescued after intravenous administration of phage libraries into mice. In one application, the isolated peptide ligands have been used to direct a cytotoxic drug to tumor vasculature in mice. Further applications in radioimaging and radiotherapy are being investigated.

Animals↗
PubMed1998

Molecular heterogeneity of the vascular endothelium revealed by in vivo phage display.

Vascular beds are known to differ in structure and metabolic function, but less is known about their molecular diversity. We have studied organ-specific molecular differences of the endothelium in various tissues by using in vivo screening of peptide libraries expressed on the surface of a bacteriophage. We report here that targeting of a large number of tissues with this method yielded, in each case, phage that homed selectively to the targeted organ. Different peptide motifs were recovered from each of these tissues. The enrichment in homing to the target organs relative to an unselected phage was 3-35-fold. Peptide sequences that conferred selective phage homing to the vasculature of lung, skin, and pancreas were characterized in detail. Immunohistochemistry showed that the phage localized in the blood vessels of their target organ. When tested, the phage homing was blocked in the presence of the cognate peptide. By targeting several tissues and by showing that specific homing could be achieved in each case, we provide evidence that organ- and tissue-specific molecular heterogeneity of the vasculature is a general, perhaps even universal, phenomenon. Our results also show that these molecular differences can serve as molecular addresses.

Animals↗
PubMed1998

Cancer treatment by targeted drug delivery to tumor vasculature in a mouse model.

In vivo selection of phage display libraries was used to isolate peptides that home specifically to tumor blood vessels. When coupled to the anticancer drug doxorubicin, two of these peptides-one containing an alphav integrin-binding Arg-Gly-Asp motif and the other an Asn-Gly-Arg motif-enhanced the efficacy of the drug against human breast cancer xenografts in nude mice and also reduced its toxicity. These results indicate that it may be possible to develop targeted chemotherapy strategies that are based on selective expression of receptors in tumor vasculature.

Animals↗
PubMed1998

Chemotherapy targeted to tumor vasculature.

Vasculature-targeted chemotherapy--the destruction of tumor blood vessels with cytotoxic agents--makes use of biochemical differences between angiogenic and resting blood vessels. This approach may minimize or eliminate some of the problems associated with conventional solid-tumor targeting, such as poor tissue penetration and drug resistance. Experiments with antiangiogenic and thrombotic factors have shown that eliminating tumor blood supply has dramatic antitumor effects in mice. Targeting chemotherapeutic agents to the tumor vasculature combines the blood vessel destruction with the usual antitumor activities of the drug, resulting in increased efficacy and reduced toxicity in experiments with tumor-bearing mice. Human clinical trials, which are soon to follow, will determine the final value of this approach.

Antineoplastic Agents↗
PubMed1997

Functional analysis of wild-type and malignant glioma derived CDKN2Abeta alleles: evidence for an RB-independent growth suppressive pathway.

The tumor suppressor gene CDKN2A (p16/MTS1/INK4A), which encodes the cyclin-dependent kinase inhibitor p16(INK4a), is a target of 9p21 deletions during the malignant progression of human gliomas. This gene also encodes a second protein product (human p16beta, murine p19ARF), which originates from an unrelated exon of CDKN2A (exon 1beta) spliced onto exon 2 in an alternate reading frame. Cell cycle arrest by p16beta is caused by an as yet unidentified pathway. In order to test the candidacy of p16beta as a glioma suppressor, we replaced p16(INK4a), p15(INK4b) and p16beta wild-type as well as a series of seven glioma-derived p16beta alleles (R87H, A112V, R120H, A121V, G125R, A128A and A128V), into glioma cell lines that had either CDKN2A-/RB+ (U-87MG and U-251MG) or CDKN2A+/RB- (LN-319) endogenous backgrounds and demonstrated that p16beta can act as a functional glioma cell growth suppressor. Moreover, p16beta, but not p16(INK4a) or p15(INK4b) inhibited the growth of RB-negative LN-319 cells, indicating that p16beta likely exerts its effects through an RB-independent pathway. In vitro and in vivo assays of pRB phosphorylation were consistent with this interpretation. Since none of the glioma-derived p16beta mutations inactivated their growth suppressive activities, it appears that mutations in CDKN2A exon 2 (which is shared in the coding sequences of p16(INK4a) and p16beta) likely exclusively target p16(INK4a).

Alleles↗
PubMed1997

Cyclin-dependent kinase 6 (CDK6) amplification in human gliomas identified using two-dimensional separation of genomic DNA.

DNA amplification is a common mechanism invoked by many human tumors to elicit overexpression of genes whose products are involved in drug resistance or cell proliferation. Although amplified regions in tumor DNA may exceed several megabases in size, segments of amplicons with a high probability of containing gene sequences may be amenable to detection by restriction landmark genomic scanning (RLGS), a high-resolution DNA analysis that separates labeled NotI fragments in two dimensions. Here, we tested this by applying RLGS to matched samples of glioma and normal brain DNA and found tumor-specific amplification of the gene encoding cyclin-dependent kinase 6 (CDK6), an observation not previously reported in human tumors. The CDK6 gene has been localized to chromosome 7q21-22, but in the gliomas studied here, it was not coamplified with either the syntenic MET (7q31) or epidermal growth factor receptor (7p11-p12) genes, suggesting that this may be part of a novel amplicon in gliomas. We then corroborated this finding by identifying both amplification-associated and amplification-independent increases in CDK6 protein levels in gliomas relative to matched normal brain samples. These data implicate the CDK6 gene in genomic amplification and illustrate the potential of RLGS for the more general identification and cloning of novel genes that are amplified in human cancer.

Base Sequence↗
PubMed1997

Point mutations can inactivate in vitro and in vivo activities of p16(INK4a)/CDKN2A in human glioma.

Deletions of chromosomal region 9p21 are among the most common genetic alterations observed during the clonal evolution of high grade malignant gliomas. Structural and functional evidence has suggested that homozygous deletion involving CDKN2A (the genetic locus encoding the cyclin-dependent kinase inhibitor p16(NK4a)) is a mechanism of inactivation of this gene and that it can be a growth suppressor in human gliomas. However, the presence of other potential suppressor genes in the 9p21 region and the relatively large sizes of the deletions has made it difficult to be certain that the CDKN2A gene is their actual target. Here, we tested this hypothesis by determining the growth suppressive effects, cell cycle inhibitions, and the activities of seven naturally occurring glioma-derived CDKN2A alleles carrying point mutations and found that two of them were functionally compromised. To resolve discrepancies among the different existing functional assays, we developed an assay for p16(INK4a) function that allowed us to demonstrate that the expression of wild-type CDKN2A, but not alleles with inactivating mutations, prevents pRB phosphorylation in vivo in human glioma cells. These data suggest that CDKN2A is a critical target for mutational inactivation in human malignant gliomas.

Alleles↗
PubMed1996

Suppression of glioblastoma angiogenicity and tumorigenicity by inhibition of endogenous expression of vascular endothelial growth factor.

The development of new capillary networks from the normal microvasculature of the host appears to be required for growth of solid tumors. Tumor cells influence this process by producing both inhibitors and positive effectors of angiogenesis. Among the latter, the vascular endothelial growth factor (VEGF) has assumed prime candidacy as a major positive physiological effector. Here, we have directly tested this hypothesis in the brain tumor, glioblastoma multiforme, one of the most highly vascularized human cancers. We introduced an antisense VEGF expression construct into glioblastoma cells and found that (i) VEGF mRNA and protein levels were markedly reduced, (ii) the modified cells did not secrete sufficient factors so as to be chemoattractive for primary human microvascular endothelial cells, (iii) the modified cells were not able to sustain tumor growth in immunodeficient animals, and (iv) the density of in vivo blood vessel formation was reduced in direct relation to the reduction of VEGF secretion and tumor formation. Moreover, revertant cells that recovered the ability to secrete VEGF regained each of these tumorigenic properties. These results suggest that VEGF plays a major angiogenic role in glioblastoma.

Animals↗
PubMed1995

Loss of P16INK4 expression is frequent in high grade gliomas.

P16INK4 is a cell cycle regulator that specifically binds to and inactivates cyclin-dependent kinase 4 (CDK4). Its encoding gene (p16/CDKN2) maps to chromosome 9p21, a region that undergoes frequent loss of heterozygosity in a variety of human tumors. We have analyzed the p16/CDKN2 gene and its expression in a series of primary glioma samples. Although homozygous deletion or mutation of the p16/CDKN2 gene was uncommon in this series and P16INK4 protein was detectable in all grade II tumors, it was present in only 50% of grade III and grade IV samples. Conversely, in some grade IV tumors that level of P16INK4 protein was elevated; in these cases, its target, CDK4, was amplified and overexpressed. These results suggest: (a) the involvement of P16INK4 in glioma progression; (b) that mechanisms other than mutation or deletion can down-regulate expression of the p16/CDKN2 gene; and (c) that the balance between CDK4 and its cognate inhibitor, P16INK4, may confer a cell growth advantage and facilitate tumor progression.

Astrocytoma↗
PubMed1995

Replacement of the p16/CDKN2 gene suppresses human glioma cell growth.

The p16/CDKN2 gene has many features of a growth suppressor gene: it maps to 9p21, a frequent region of loss of heterozygozity in a variety of tumor types; it encodes an inhibitor of cyclin-dependent kinase 4; and its homozygous deletion is common in tumor-derived cell lines. However, the lower frequency of alteration of the gene in primary tumor tissue as compared to the cognate tumor cell lines has brought this interpretation into question. We have assessed the growth suppressive function of p16/CDKN2 by gene transfer. The introduction of full-length p16/CDKN2 cDNA caused marked growth suppression in p16/CDKN2-null human glioma cells, but was without significant effect in those cells with endogenous wild-type p16/CDKN2 alleles. These results provide functional evidence in support of the hypothesis that the p16/CDKN2 gene is a functional growth suppressor gene, at least in gliomas.

Base Sequence↗
PubMed1989

Tumor and serum beta-2-microglobulin expression in women with breast cancer.

To investigate whether the tumor expression of beta-2-microglobulin (beta 2-M) could serve as a marker of tumor biologic behavior, the authors studied specimens of breast carcinomas from 60 consecutive female patients. Presence of beta 2-M was analyzed by immunohistochemistry. No significant correlations were found between tumor beta 2-M expression and several histologic attributes such as type, histologic and nuclear grades, mitotic index, necrosis, vascular invasion, and lymphocytic infiltration. Likewise, beta 2-M was not associated with markers of disease extension such as TNM, (UICC, classification of malignant tumors) staging and axillary lymph node involvement or with estrogen, progesterone, and glucocorticoid receptor levels. However, there was a significantly positive association between tumor beta 2-M expression and the degree of lymphocytic infiltration in the tumor tissue. Beta 2-M serum levels were determined by an enzyme-linked immunosorbent assay in samples from 22 of the above women. Although some of the highest values had been obtained in women with larger (T4) primary tumors, the authors failed to detect any statistical relationship between beta 2-M expression in the tumor with serum levels or between serum beta 2-M and the above histologic, laboratory, and clinical factors.

Adult↗