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Biomedical subjects

G B Mills

Publications and source records attributed to G B Mills.

At least 19 recordsLinked to original sources
PubMed2006

Identification of a novel splice variant of AML1b in ovarian cancer patients conferring loss of wild-type tumor suppressive functions.

Acute myeloid leukemia (AML) 1 is often disrupted by chromosomal translocations generating oncogenic fusions in human leukemias. However, its role in epithelial cancers has not been extensively investigated. Herein, we show a marked accumulation of AML1 transcripts including a high frequency of a novel alternatively spliced AML1b transcript lacking exon 6 (AML1b(Del179-242)) in ovarian cancer patients. The increases in RNA transcripts for total wild-type AML1 and AML1b(Del179-242) are associated with poor patient outcomes. We have shown that although both wild-type AML1b and AML1b(Del179-242) are localized to nuclear speckles, AML1b(Del179-242) was observed to have dramatically reduced transactivation potential with the plasminogen activator inhibitor-1 promoters and behaved as a weak dominant negative of wild-type AML1b. Wild-type AML1b was found to inhibit the growth of immortalized ovarian epithelial cells (T29) decreasing colony-forming ability. Moreover, we have identified a novel function of AML1b where it inhibits ovarian cell migration. In contrast, AML1b(Del179-242) has lost the ability to inhibit both ovarian cell proliferation and migration indicating that the functional effects observed with wild-type AML1b are dependent on amino acids 179-242. Collectively, these studies suggest that deregulated alternative splicing of AML1b transcripts may potentially contribute to the pathophysiology of ovarian cancers.

Alternative Splicing↗
PubMed2006

Silencing mammalian target of rapamycin signaling by small interfering RNA enhances rapamycin-induced autophagy in malignant glioma cells.

The mammalian target of rapamycin (mTOR) plays a central role in regulating the proliferation of malignant glioma cells, and mTOR-specific inhibitors such as rapamycin analogs are considered as promising therapy for malignant gliomas. However, the efficacy of mTOR inhibitors alone in the treatment of patients with malignant gliomas is only modest, potentially because these agents rather than acting as mTOR kinase inhibitors instead interfere with the function of only mTOR/raptor (regulatory-associated protein of mTOR) complex and thus do not perturb all mTOR functions. The purpose of this study was to determine whether global inhibition of the mTOR molecule enhances the antitumor effect of rapamycin on malignant glioma cells. We showed that rapamycin induced autophagy and that inhibition of autophagy by small interfering RNA (siRNA) directed against autophagy-related gene Beclin 1 attenuated the cytotoxicity of rapamycin in rapamycin-sensitive tumor cells, indicating that the autophagy was a primary mediator of rapamycin's antitumor effect rather than a protective response. Exogenous expression of an mTOR mutant interfering with its kinase activity markedly enhanced the incidence of rapamycin-induced autophagy. Moreover, silencing of mTOR with siRNA augmented the inhibitory effect of rapamycin on tumor cell viability by stimulating autophagy. Importantly, not only rapamycin-sensitive malignant glioma cells with PTEN mutations but also rapamycin-resistant malignant glioma cells with wild-type PTEN were sensitized to rapamycin by mTOR siRNA. These results indicate that rapamycin-induced autophagy is one of the agent's antitumor effects and that silencing or inhibiting mTOR kinase activity could enhance the effectiveness of rapamycin.

Autophagy↗
PubMed2006

Ovarian cancer: homeobox genes, autocrine/paracrine growth, and kinase signaling.

Epithelial ovarian cancer, the fourth leading cause of cancer deaths in American women, is currently classified by surgical and histologic appearance. However, the predictive value of this classification is limited. The risk of epithelial ovarian cancer increases with the number of ovulatory events. It is now thought that different ovarian tumors are derived from a single ovarian surface epithelial precursor cell with the degree and pattern of differentiation determined by combinatorial expression of homeobox genes normally involved in differentiation of the female genital tract. This aberrant differentiation occurs in association with histology-specific genomic aberrations, genomic instability, and resultant chromosomal changes, and may be triggered by prolonged abnormal or excessive exposure of surface epithelial cells to autocrine/paracrine stimulation by sex steroids and other growth factors. As the disease progresses, activation of kinase pathways and continued abnormal autocrine/paracrine stimulation contribute to genomic instability but also identify potential targets for novel therapeutic intervention.

ErbB Receptors↗
PubMed2005

The Akt inhibitor KP372-1 suppresses Akt activity and cell proliferation and induces apoptosis in thyroid cancer cells.

The phosphatidylinositol 3' kinase (PI3K)/phosphatase and tensin homologue deleted on chromosome ten/Akt pathway, which is a critical regulator of cell proliferation and survival, is mutated or activated in a wide variety of cancers. Akt appears to be a key central node in this pathway and thus is an attractive target for targeted molecular therapy. We demonstrated that Akt is highly phosphorylated in thyroid cancer cell lines and human thyroid cancer specimens, and hypothesised that KP372-1, an Akt inhibitor, would block signalling through the PI3K pathway and inhibit cell proliferation while inducing apoptosis of thyroid cancer cells. KP372-1 blocked signalling downstream of Akt in thyroid tumour cells, leading to inhibition of cell proliferation and increased apoptosis. As thyroid cancer consistently expresses phosphorylated Akt and KP372-1 effectively blocks Akt signalling, further preclinical evaluation of this compound for treatment of thyroid cancer is warranted.

Apoptosis↗
PubMed2004

Search for pi(0)-->nu(mu)nu(mu) decays in the LSND detector.

We observe a net beam excess of 8.7+/-6.3(stat)+/-2.4(syst) events, above 160 MeV, resulting from the charged-current reaction of nu(micro) and/or nu;(mu) on C and H in the LSND detector. No beam-related muon background is expected in this energy regime. Within an analysis framework of pi(0)-->nu(mu)nu;(mu), we set a direct upper limit for this branching ratio of Gamma(pi(0)-->nu(mu)nu;(mu))/Gamma(pi(0)-->all)<1.6 x 10(-6) at 90% confidence level.

Journal Article↗
PubMed2004

Inhibition of phosphatidylinositol 3-kinase dephosphorylates BAD and promotes apoptosis in myeloid leukemias.

The phosphatidylinositol 3-kinase (PI3K)/AKT protein kinase pathway is involved in cell growth, proliferation, and apoptosis. The functional activation of PI3K/AKT provides survival signals and blockade of this pathway may facilitate cell death. Downstream targets of PI3K-AKT include the proapoptotic protein BAD, caspase-9, NF-kappaB, and Forkhead. We have previously reported that BAD is constitutively phosphorylated in primary acute myeloid leukemia (AML) cells, a post-transcriptional modification, which inactivates its proapoptotic function. In this study, we tested the hypothesis that the inhibition of PI3K by LY294002 results in the dephosphorylation of AKT and BAD, and thus promote leukemia cell apoptosis. We investigated the effects of LY294002 in megakaryocytic leukemia-derived MO7E cells, primary AML and normal bone marrow progenitor cells. In MO7E cells, LY294002 reduced AKT kinase activity, induced dephosphorylation of AKT and BAD, and increased apoptosis. Concomitant inhibition of mitogen-activated protein kinase signaling or combination with all-trans retinoic acid further enhanced apoptosis of leukemic cells. In primary AML samples, clonogenic cell growth was significantly reduced. Normal hematopoietic progenitors were less affected, suggesting preferential targeting of leukemia cells. In conclusion, the data suggest that the inhibition of the PI3K/AKT signaling pathway restores apoptosis in AML and may be explored as a novel target for molecular therapeutics in AML.

Apoptosis↗
PubMed2003

Expression of endogenous granzyme B in a subset of human primary breast carcinomas.

Granzyme B (GrB) is the prototypic member of a serine protease family primarily used by cytotoxic lymphocytes to kill target cells. We report here that, by immunohistochemical staining of paraffin-embedded tumour sections, GrB protein was unexpectedly detected in malignant cells of a subset of breast cancers and their adjacent reactive endothelial and mesenchymal cells in which endogenous retinoblastoma protein (pRB) is overexpressed. The identity of the endogenous GrB was further confirmed experimentally in RB-deficient breast carcinoma cell culture upon overexpression of ectopic pRB. Our finding extends the recent paradigm-shifting trend for a more diverse biological role of granzyme B, and might provide a rational basis for exploring its potential prognostic value in a variety of human cancers.

Biopsy↗
PubMed2003

Effect of CD26/dipeptidyl peptidase IV on Jurkat sensitivity to G2/M arrest induced by topoisomerase II inhibitors.

CD26/dipeptidyl peptidase IV (DPPIV) is a surface antigen with multiple functions, including a role in T-cell activation and the development of certain human cancers. We previously demonstrated that CD26/DPPIV enhanced sensitivity of Jurkat cells to doxorubicin. We now show that expression of CD26/DPPIV enhanced sensitivity of CD26 Jurkat transfectants to G(2)-M arrest mediated by the antineoplastic agent etoposide. The increased sensitivity to etoposide-induced G(2)-M arrest was associated with disruption of cell cycle-related events, including hyperphosphorylation of p34(cdc2) kinase, change in cdc25C expression and phosphorylation, and alteration in cyclin B1 expression. CD26/DPPIV-associated enhancement of doxorubicin and etoposide-induced G(2)-M arrest was also observed in serum-free media, suggesting an effect of CD26 on cell-derived processes rather than serum-derived factors. Importantly, our work elucidated a potential mechanism for the enhanced susceptibility of CD26-expressing Jurkat cells to the topoisomerase II inhibitors by demonstrating that CD26/DPPIV surface expression was associated with increased topoisomerase II alpha levels and enhanced enzyme activity. Besides being the first to show a functional association between the multifaceted molecule CD26 and the key cellular protein topoisomerase II alpha, our studies provide additional evidence of a potential role for CD26 in the treatment of selected malignancies.

Antigens, Neoplasm↗
PubMed2003

Promising directions for the diagnosis and management of gynecological cancers.

Diagnosis and management of cancer requires tools with both high sensitivity and specificity. The minimally invasive cervical smear has demonstrated how a test, even one with low specificity, can change the public health profile of a cancer from a late stage deadly disease to early diagnosis with rare tumor-related deaths. The benefit of such a test is best demonstrated by the low frequency of cervix cancer and its good outcome in countries where this test is readily available and used with appropriate secondary follow up. Early and specific symptoms, and identification and prevention for high risk groups has had similar impact for endometrial cancer. Neither a robust test, nor reliable or specific early symptoms are available for ovarian cancer, making clinical and scientific advances in this area a critical world-wide need. Current approaches testing one protein or gene marker at a time will not address this crisis expeditiously. New sensitive, specific, accurate, and reliable technologies that can be implemented using high throughput mechanisms are needed at as low a cost as possible. Ideally, these technologies should be focused on readily available patient resources, such as blood or urine, or as in the case of cervix cancer, minimally invasive informative approaches such as cervical smears. Techniques that allow data mining from a large input database overcome the slow advances of one protein-one gene investigation, and further address the multi-faceted carcinogenesis process occurring even in germ line mutation-associated malignancy. Proteomics, the study of the cellular proteins and their activation states, has led the progress in biomarker development for ovarian and other cancers and is being applied to management assessment. Amenable to high throughput, internet interface, and representative of the proteome spectrum, proteomic technology is the newest and most promising direction for translational developments in gynecologic cancers.

Biomarkers, Tumor↗
PubMed2001

Heregulin-induced apoptosis is mediated by down-regulation of Bcl-2 and activation of caspase-7 and is potentiated by impairment of protein kinase C alpha activity.

Heregulins are a group of growth factors that play diverse and critical roles in the signaling network of the human epidermal growth factor receptor (HER or EGFR) superfamily. Our earlier studies have shown that recombinant heregulinbeta1 (HRG) induces apoptosis in SKBr3 breast cancer cells that overexpress HER2. Here we report molecular mechanisms of HRG-induced apoptosis. HRG treatment of SKBr3 cells for 72 h decreased the level of Bcl-2 protein. HRG treatment led to degradation of poly (ADP-ribose) polymerase (PARP) and activated both caspase-9 and caspase-7. No significant activation of caspase-3, -6, or -8 was detected. Expression of exogenous caspase-7 by adenovirus-caspase-7 (Ad-casp-7) in SKBr3 cells resulted in apoptosis, which mimicked the effect of HRG treatment. Expression of exogenous caspase-7 had no impact on Bcl-2 expression, but promoted PARP degradation. Two highly selective inhibitors of protein kinase C (PKC), GF109203X (GF) and Ro318425 (Ro), significantly enhanced HRG-induced apoptosis as determined by flow cytometric analysis and DNA fragmentation assay. Accordingly, the PKC inhibitor GF further decreased the level of Bcl-2 protein and further degraded PARP in HRG-treated cells. Assay of PKC activity indicated that HRG activated PKC in SKBr3 cells, predominantly affecting the PKCalpha isoform. To confirm which PKC isoform(s) mediated potentiation of HRG-induced apoptosis, the profile of PKC isoforms was measured in SKBr3 cells. Five PKC isoforms, PKCalpha, PKCiota, PKCzeta, PKClambda, and PKCdelta as well as their receptors (RACK1) were expressed in this cell line. Treatment with PKC inhibitors GF and Ro decreased protein levels of both PKCalpha and PKCdelta at 24 h. PKCalpha levels were still depressed at 72 h. GF and Ro had little effect on the expression of other PKC isoforms. An inhibitor of classical PKC isoforms (Go6976) enhanced HRG-induced apoptosis, whereas the PKCdelta selective inhibitor rottlerin did not. As PKCalpha was the only classical isoform expressed in SKBr3 cells, the effect of Go6976 on HRG-induced apoptosis largely related to inhibition of PKCalpha. Constitutive expression of wild-type PKCalpha attenuated the apoptosis produced by HRG and GF. Consequently, HRG-induced apoptosis in SKBr3 cells appeared to involve down-regulation of Bcl-2 protein, activation of caspase-9 and caspase-7, and degradation of PARP. Inhibition of PKC function enhanced HRG-induced apoptosis, leading to synergistic down-regulation of Bcl-2 expression. Impairment of the PKCalpha isoform alone was sufficient to potentiate HRG-induced apoptosis.

Apoptosis↗
PubMed2001

A single nucleotide polymorphism in the matrix metalloproteinase-1 promoter enhances lung cancer susceptibility.

Extracellular matrix-degrading matrix metalloproteinase-1 (MMP-1) is one of the interstitial collagenases likely to be involved in tumor invasion and metastasis. MMP-1 may also contribute to tumor initiation and development by altering the cellular microenvironment that facilitates tumor formation. Recent studies have found that overexpression of MMP-1 is associated with the initial stages of cancer development in addition to promoting cellular invasion; however, preexisting oncogenic mutations or chemical carcinogens are required to initiate tumorigenesis as well. There is a single nucleotide polymorphism located in the promoter region of MMP-1 that partially regulates gene expression. The 2G/2G genotype enhances transcriptional activity and may be associated with an increased lung cancer risk. Using a case-control study, we tested the hypotheses that (a) individuals with the 2G/2G genotype may be at an increased risk for lung cancer; and (b) the risk should be greatly elevated in smoking individuals. PCR-RFLP was used to determine the MMP-1 genotypes in 456 lung-cancer cases and 451 frequency-matched controls of Caucasian ethnicity. Overall, there was a significant association between the 2G/2G genotype and lung cancer risk [odds ratio (OR), 1.76; 95% confidence interval (CI), 1.29-2.39]. In current smokers, the lung cancer risk associated with the 2G/2G genotype was significantly elevated (OR, 3.16; 95% CI, 1.87-5.35). However, this association was less evident in former smokers (OR, 1.23; 95% CI, 0.81-1.87) and absent in never smokers (OR, 1.09; 95% CI, 0.31-3.91). Similarly, this risk was more evident in heavy smokers (OR, 2.55; 95% CI, 1.61-4.03) than in light smokers (OR, 1.40; 95% CI, 0.84-2.32). Interestingly, men were observed to have a 2.15-fold increased lung cancer risk (OR, 2.15; 95% CI, 1.42-3.26) compared with women (OR, 1.34; 95% CI, 0.84-2.15). Furthermore, subjects with 2G/2G genotype developed lung cancer earlier (60.94 +/- 0.64 years old) than patients with 1G/1G and 1G/2G genotypes (62.91 +/- 0.59 years old; P = 0.024). Our data demonstrate that the 2G/2G genotype enhances lung cancer susceptibility especially in current smokers. To our knowledge, these results report the first molecular epidemiological evidence of the MMP-1 promoter polymorphism associated with the development of lung cancer in the presence of continuing carcinogenic exposure.

Age Factors↗
PubMed2001

Expression of CD26 and its associated dipeptidyl peptidase IV enzyme activity enhances sensitivity to doxorubicin-induced cell cycle arrest at the G(2)/M checkpoint.

CD26, a M(r) 110,000 surface-bound ectopeptidase with dipeptidyl peptidase IV (DPPIV) activity, has an array of diverse functional properties, with a role in T-cell physiology and the development of certain human cancers. In this study, we report that surface expression of CD26, through its associated DPPIV enzyme activity, enhanced sensitivity of Jurkat T-cell transfectants to G(2)-M arrest induced by the chemotherapeutic drug, doxorubicin. This was associated with disruption of cell cycle-related events, including hyperphosphorylation and inhibition of p34(cdc2) kinase activity, phosphorylation of cdc25C, and alteration in cyclin B1 expression. In addition, we demonstrate that the addition of exogenous soluble DPPIV enhanced sensitivity of lymphoid tumor cell lines to doxorubicin, suggesting a potentially useful clinical role for CD26/DPPIV in the treatment of selected human hematological malignancies.

Antibiotics, Antineoplastic↗
PubMed2001

Genetic analysis of early- versus late-stage ovarian tumors.

In the United States, ovarian cancer is the fourth most common cause of cancer-related deaths among women. The most important prognostic factor for this cancer is tumor stage, or extent of disease at diagnosis. Although women with low-stage tumors have a relatively good prognosis, most women diagnosed with late-stage disease eventually succumb to their cancer. In an attempt to understand early events in ovarian carcinogenesis, and to explore steps in its progression, we have applied multiple molecular genetic techniques to the analysis of 21 early-stage (stage I/II) and 17 advanced-stage (stage III/IV) ovarian tumors. These techniques included expression profiling with cDNA microarrays containing approximately 18,000 expressed sequences, and comparative genomic hybridization to address the chromosomal locations of copy number gains as well as losses. Results from the analysis indicate that early-stage ovarian cancers exhibit profound alterations in gene expression, many of which are similar to those identified in late-stage tumors. However, differences observed at the genomic level suggest differences between the early- and late-stage tumors and provide support for a progression model for ovarian cancer development.

Disease Progression↗
PubMed2001

Involvement of JNK-mediated pathway in EGF-mediated protection against paclitaxel-induced apoptosis in SiHa human cervical cancer cells.

We investigated the signalling pathways by which epidermal growth factor (EGF) modulates paclitaxel-induced apoptosis in SiHa human cervical cancer cells. SiHa cells exposed to paclitaxel underwent apoptosis, which was strongly inhibited by EGF. This inhibition of apoptosis by EGF was not altered by pharmacological blockade of phosphatidylinositol 3'-OH kinase (PI-3K) with the PI-3K specific inhibitor LY294002 or blockade of the mitogen-activated protein kinase (MAPK) kinase (MEK) with the MEK specific inhibitor PD98059, or by transfection of the cells with PI-3K or MEK dominant-negative expression vectors. EGF did not stimulate PI-3K/Akt, MEK/MAPK, or p38 MAPK activity in SiHa cells but did transiently activate the c-Jun NH2-terminal kinase (JNK). Co-exposure of SiHa cells to SB202190 at concentrations that inhibit JNK abolished the protective effect of EGF on SiHa cells against paclitaxel-induced apoptosis. Our findings indicate that the JNK signaling pathway plays an important role in EGF-mediated protection from paclitaxel-induced apoptosis in SiHa cells.

Antineoplastic Agents, Phytogenic↗
PubMed2001

Triterpenoid saponins from Acacia victoriae (Bentham) decrease tumor cell proliferation and induce apoptosis.

This report describes the isolation and partial purification of novel triterpenoid saponins [Fraction 35 (F035)] and two pure biologically active derivatives (termed avicins D and G) from Acacia victoriae, an Australian desert tree of the Leguminosae family. F035 and the avicins markedly inhibited the growth of several tumor cell lines with minimum growth inhibition in human foreskin fibroblasts, mouse fibroblasts, and immortalized breast epithelial cells at similar concentrations. F035 and the avicins induced cell cycle (G1) arrest of the human MDA-MB-453 breast cancer cell line and apoptosis of the Jurkat (T-cell leukemia) and the MDA-MB-435 breast cancer cell line. The triterpenoid saponins also partially inhibited phosphatidylinositol 3-kinase activity in Jurkat T cells in a time-dependent manner and phosphorylation in the downstream protein Akt, whereas no affect was seen on the Ras/mitogen-activated protein kinase cascade. These observations as well as other work from our laboratory demonstrating mitochondrial perturbation, chemoprevention, and inhibition of nuclear factor kappaB suggest that triterpenoid saponins from A. victoriae have potential as novel anticancer agents. Recent work linking Akt signaling with glucose metabolism, stress resistance, and longevity suggests other potential applications of these compounds.

Acacia↗