Growth Inhibitory Activity of Extracted Material and Isolated Compounds from the Fruits of Kigelia pinnata

A study of the components of the fruits of Kigelia pinnata was undertaken to identify compounds with potential growth inhibitory activity against human melanoma cells, since extracts from the fruits of this plant have been described in traditional medicine to have application in the treatment of skin cancer and other skin ailments. A bioactivity-guided fractionation process yielded a number of crude fractions, which demonstrated cytotoxicity in vitro against human melanoma cells. Compounds isolated and identified included the isocoumarins, demethylkigelin (1) and kigelin 2), fatty acids, oleic (3) and heneicosanoic acids (4), the furonaphthoquinone, 2-(1-hydroxyethyl)-naphtho[2,3-b]furan-4,9-dione (5), and ferulic acid (6). A number of structurally related synthetic compounds were also tested using the MTT assay. The most potent series of these compounds, the furonaphthoquinones, also demonstrated a cytotoxic effect in two human breast cancer cell lines tested.

Synthesis and in Vitro and in Vivo Evaluation of a Series of Dihydroisocoumarin Derivatives Conjugated with Fatty Acids, Alcohols, and Amines as Potential Anticancer Agents

In this paper, we report the synthesis and biological activity of a series of dihydroisocoumarin analogues Conjugated with fatty acids, alcohols, or amines, of varying hydrocarbon chain length and degree of unsaturation, to (he dihydroisocoumarins, kigelin and mellein, at the C-7 and C-8 positions on the core dihydroisocoumarin structure. These compounds were evaluated for their antiproliferative activity against human breast cancer (MCF-7 and MDA-MB-468) and melanoma cells (SK-MEL-28 and Malme-3M) using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Two compounds Conjugated with gamma-linolenyl alcohol (18:3 n-6) demonstrated potent antiproliferative activity in vitro with one of these 4-hydroxy-3-oxo-1.3-dihydro-isobenzofuran-5-carboxylic acid octadeca-6,9,12-trienyl ester, demonstrating significant antitumor activity in vivo ill a number of human tumor xenograft models.

Development of a novel experimental model to investigate radiobiological implications of respiratory motion in advanced radiotherapy

Respiratory motion introduces complex spatio-temporal variations in the dosimetry of radiotherapy. There is a paucity of literature investigating the radiobiological consequences of intrafraction motion and concerns regarding the impact of movement when applied to cancer cell lines in vitro exist. We have addressed this by developing a novel model which accurately replicates respiratory motion under experimental conditions to allow clinically relevant irradiation of cell lines. A bespoke phantom and motor driven moving platform was adapted to accommodate flasks containing medium and cells in order to replicate respiratory motion using varying frequencies and amplitude settings. To study this effect on cell survival in vitro, dose response curves were determined for human lung cancer cell lines H1299 and H460 exposed to a uniform 6 MV radiation field under moving or stationary conditions. Cell survival curves showed no significant difference between irradiation at different dose points for these cell lines in the presence or absence of motion. These data indicate that motion of unshielded cells in vitro does not affect cell survival in the presence of uniform irradiation. This model provides a novel research platform to investigate the radiobiological consequences of respiratory motion in radiotherapy.

PsT-1: A new Tryptophyllin peptide from the skin secretion of Waxy Monkey Leaf Frog, Phyllomedusa sauvagei

The Waxy Monkey Leaf Frog, Phyllomedusa sauvagei, has been extensively-studied for many years, and a broad spectrum of bioactive peptides has been found in its skin secretions. Here we report the discovery of a novel tryptophyllin (TPH) peptide, named PsT-1, from this frog species. Skin secretions from specimens of P. sauvagei were collected by mild electrical stimulation. Peptides were identified and characterized by transcriptome cloning, and the structure was confirmed by MALDI-TOF mass spectrometry and automated Edman degradation. This novel peptide was encoded by a single precursor of 61 amino acid residues, whose primary structure was deduced from cloned skin cDNA. Analysis of different amphibian tryptophyllins revealed that PsT-1 exhibited a high degree of primary structural similarity to its homologues, PdT-1 and PdT-2, from the Mexican giant leaf frog, Pachymedusa dacnicolor. A synthetic replicate of PsT-1 was found to inhibit bradykinin-induced vasorelaxation of phenylephrine pre-constricted rat tail artery smooth muscle. It was also found that PsT-1 had an anti-proliferative effect on three different human prostate cancer cell lines (LNCaP/PC3/DU145), by use of an MTT assay coupled with direct cell counting as measures of cell growth. These data indicate that PsT-1 is a likely bradykinin receptor antagonist and its biological effects are probably mediated through bradykinin receptors. As a BK antagonist, PST-1, with antagonistic effects on BK in artery smooth muscle, inhibition of proliferation in prostate cancer cells and lack of undesirable side effects, may have potential in cardiovascular, inflammatory and anticancer therapy.

Achaete-scute homolog-1 linked to remodeling and preneoplasia of pulmonary epithelium

The basic helix-loop-helix protein achaete-scute homolog-1 (ASH1) is involved in lung neuroendocrine (NE) differentiation and tumor promotion in SV40 transgenic mice. Constitutive expression of human ASH-1 (hASH1) in mouse lung results in hyperplasia and remodeling that mimics bronchiolization of alveoli (BOA), a potentially premalignant lesion of human lung carcinomas. We now show that this is due to sustained cellular proliferation in terminal bronchioles and resistance to apoptosis. Throughout the airway epithelium the expression of anti-apoptotic Bcl-2 and c-Myb was increased and Akt/mTOR pathway activated. Moreover, the expression of matrix metalloproteases (MMPs) including MMP7 was specifically enhanced at the bronchiolo-alveolar duct junction and BOA suggesting that MMPs play a key role in this microenvironment during remodeling. We also detected MMP7 in 70% of human BOA lesions. Knockdown of hASH1 gene in human lung cancer cells in vitro suppressed growth by increasing apoptosis. We also show that forced expression of hASH1 in immortalized human bronchial epithelial cells decreases apoptosis. We conclude that the impact of hASH1 is not limited to cells with NE phenotype. Rather, constitutive expression of hASH1 in lung epithelium promotes remodeling through multiple pathways that are commonly activated during lung carcinogenesis. The collective results suggest a novel model of BOA formation via hASH1-induced suppression of the apoptotic pathway. Our study yields a promising new preclinical tool for chemoprevention of peripheral lung carcinomas. © 2007 USCAP, Inc All rights reserved.

Dose, dose-rate and field size effects on cell survival following exposure to non-uniform radiation fields

For the delivery of intensity-modulated radiation therapy (IMRT), highly modulated fields are used to achieve dose conformity across a target tumour volume. Recent in vitro evidence has demonstrated significant alterations in cell survival occurring out-of-field which cannot be accounted for on the basis of scattered dose. The radiobiological effect of area, dose and dose-rate on out-of-field cell survival responses following exposure to intensity-modulated radiation fields is presented in this study. Cell survival was determined by clonogenic assay in human prostate cancer (DU-145) and primary fibroblast (AG0-1522) cells following exposure to different modulated field configurations delivered using a X-Rad 225 kVp x-ray source. Uniform survival responses were compared to in- and out-of-field responses in which 25-99% of the cell population was shielded. Dose delivered to the out-of-field region was varied from 1.6-37.2% of that delivered to the in-field region using different levels of brass shielding. Dose rate effects were determined for 0.2-4 Gy min⁻¹ for uniform and modulated exposures with no effect seen in- or out-of-field. Survival responses showed little dependence on dose rate and area in- and out-of-field with a trend towards increased survival with decreased in-field area. Out-of-field survival responses were shown to scale in proportion to dose delivered to the in-field region and also local dose delivered out-of-field. Mathematical modelling of these findings has shown survival response to be highly dependent on dose delivered in- and out-of-field but not on area or dose rate. These data provide further insight into the radiobiological parameters impacting on cell survival following exposure to modulated irradiation fields highlighting the need for refinement of existing radiobiological models to incorporate non-targeted effects and modulated dose distributions.

Molecular Cloning of a Novel Tryptophyllin Peptide from the Skin of the Orange-Legged Monkey Frog, Phyllomedusa hypochondrialis

Tryptophyllins are a group of small (4–14 amino acids), heterogenous peptides, mostly from the skins of hylid frogs from the genera, Phyllomedusa and Litoria. To date, more than forty TPHs have been discovered in species from these two genera. Here, we describe the identification of a novel tryptophyllin type 3 peptide, PhT-3, from the extracts of skin of the orange-legged monkey frog, Phyllomedusa hypochondrialis, and molecular cloning of its precursor-encoding cDNA from a cDNA library constructed from the same skin sample. Full primary structural characterization was achieved using a combination of direct Edman degradation, mass spectrometry and deduction from cloned skin-derived cDNA. The open-reading frame of the precursor cDNA was found to consist of 63 amino acid residues. The mature peptide arising from this precursor contains a post-translationally modified N-terminal pyroglutamate (pGlu) residue, formed from acid-mediated cyclization of an N-terminal Gln (Q) residue, and with the structure: pGlu-Asp-Lys-Pro-Phe-Trp-Pro-Pro-Pro-Ile-Tyr-Pro-Met. Pharmacological assessment of a synthetic replicate of this peptide on phenylephrine preconstricted rat tail artery segments, revealed a reduction in relaxation induced by bradykinin. PhT-3 was also found to mediate antiproliferative effects on human prostate cancer cell lines.

Identification and Functional Analysis of a Novel Tryptophyllin Peptide from the Skin of the Red-eye Leaf Frog, Agalychnis callidryas

Amphibian skin has proved repeatedly to be a largely untapped source of bioactive peptides and this is especially true of members of the Phyllomedusinae subfamily of frogs native to South and Central America. Tryptophyllins are a group of peptides mainly found in the skin of members of this genus. In this study, a novel tryptophyllin (TPH) type 3 peptide, named AcT-3, has been isolated and structurally-characterised from the skin secretion and lyophilised skin extract of the red-eye leaf frog, Agalychnis callidryas. The peptide was identified in and purified from the skin secretion by reverse-phase HPLC. MALDI-TOF mass spectrometry and MS/MS fragmentation sequencing established its primary structure as: pGlu-Gly-Lys-Pro-Tyr-Trp-Pro-Pro-Pro-Phe-Leu-Pro-Glu, with a non-protonated molecular mass of 1538.19Da. The mature peptide possessed the canonical N-terminal pGlu residue that arises from post-translational modification of a Gln residue. The deduced open-reading frame consisted of 63 amino acid residues encoding a highly-conserved signal peptide of approximately 22 amino acid residues, an intervening acidic spacer peptide domain, a single AcT-3 encoding domain and a C terminal processing site. A synthetic replicate of AcT-3 was found to antagonise the effect of BK on rat tail artery smooth muscle and to contract the intestinal smooth muscle preparations. It was also found that AcT-3 could dose-dependently inhibit the proliferation of human prostate cancer cell lines after 72h incubation.

Oral Bisphosphonate Exposure and the Risk of Upper Gastrointestinal Cancers

The association between oral bisphosphonate use and upper gastrointestinal cancer has been controversial. Therefore, we examined the association with esophageal and gastric cancer within the Kaiser Permanente, Northern California population. A total of 1,011 cases of esophageal (squamous cell carcinoma and adenocarcinoma) and 1,923 cases of gastric adenocarcinoma (cardia, non-cardia and other) diagnosed between 1997 and 2011 from the Kaiser Permanente, Northern California cancer registry were matched to 49,886 and 93,747 controls, respectively. Oral bisphosphonate prescription fills at least one year prior to the index date were extracted. Conditional logistic regression was used to calculate odds ratios (OR) and 95% confidence intervals (95% CI) for the associations between prospectively evaluated oral bisphosphonate use with incident esophageal and gastric cancer diagnoses with adjustment for potential confounders. After adjustment for potential confounders, no significant associations were found for esophageal squamous cell carcinoma (OR 0.88; 95% CI: 0.51, 1.52), esophageal adenocarcinoma (OR 0.68; 95% CI: 0.37, 1.24), or gastric non-cardia adenocarcinoma (OR 0.83, 95% CI: 0.59, 1.18), but we observed an adverse association with gastric cardia adenocarcinoma (OR 1.64; 95% CI: 1.07, 2.50). In conclusion, we observed no association between oral bisphosphonate use and esophageal cancer risk within a large community-based population. A significant association was detected with gastric cardia and other adenocarcinoma risk, although this needs to be replicated.

Human Papillomavirus and Oral Cancer: the International Agency for Research on Cancer Multicenter Study

Background: Human papillomavirus (HPV), the causal agent of cervical cancer, appears to be involved in the etiology of cancer of the oral cavity and oropharynx. To investigate these associations, we conducted a multicenter case-control study of cancer of the oral cavity and oropharynx in nine countries. Methods: We recruited 1670 case patients (1415 with cancer of the oral cavity and 255 with cancer of the oropharynx) and 1732 control subjects and obtained an interview, oral exfoliated cells, and blood from all participants and fresh biopsy specimens from case patients. HPV DNA was detected by polymerase chain reaction (PCR). Antibodies against HPV16 L1, E6, and E7 proteins in plasma were detected with enzyme-linked immunosorbent assays. Multivariable models were used for case-control and case-case comparisons. Results: HPV DNA was detected in biopsy specimens of 3.9% (95% confidence interval [CI]=2.5% to 5.3%) of 766 cancers of the oral cavity with valid PCR results and 18.3% (95% CI=12.0% to 24.7%) of 142 cancers of the oropharynx (oropharynx and tonsil combined) with valid PCR results. HPV DNA in cancer biopsy specimens was detected less frequently among tobacco smokers and paan chewers and more frequently among subjects who reported more than one sexual partner or who practiced oral sex. HPV16 DNA was found in 94.7% of HPV DNA-positive case patients. HPV DNA in exfoliated cells was not associated with cancer risk or with HPV DNA detection in biopsy specimens. Antibodies against HPV16 L1 were associated with risk for cancers of the oral cavity (odds ratio [OR]=1.5, 95% CI=1.1 to 2.1) and the oropharynx (OR=3.5, 95% CI=2.1 to 5.9). Antibodies against HPV16 E6 or E7 were also associated with risk for cancers of the oral cavity (OR=2.9, 95% CI=1.7 to 4.8) and the oropharynx (OR=9.2, 95% CI=4.8 to 17.7). Conclusions: HPV appears to play an etiologic role in many cancers of the oropharynx and possibly a small subgroup of cancers of the oral cavity. The most common HPV type in genital cancers (HPV16) was also the most common in these tumors. The mechanism of transmission of HPV to the oral cavity warrants further investigation.

Molecular pathology of RUNX3 in human carcinogenesis

A major goal of molecular biology is to elucidate the mechanisms underlying cancer development and progression in order to achieve early detection, better diagnosis and staging and novel preventive and therapeutic strategies. We feel that an understanding of Runt-related transcription factor 3 (RUNX3)-regulated biological pathways will directly impact our knowledge of these areas of human carcinogenesis. The RUNX3 transcription factor is a downstream effector of the transforming growth factor-beta (TGF-beta) signaling pathway, and has a critical role in the regulation of cell proliferation and cell death by apoptosis, and in angiogenesis, cell adhesion and invasion. We previously identified RUNX3 as a major gastric tumor suppressor by establishing a causal relationship between loss of function and gastric carcinogenesis. More recently, we showed that RUNX3 functions as a bona fide initiator of colonic carcinogenesis by linking the Wnt oncogenic and TGF-beta tumor suppressive pathways. Apart from gastric and colorectal cancers. a multitude of epithelial cancers exhibit inactivation of RUNX3, thereby making it a putative tumor suppressor in human neoplasia. This review highlights our current understanding of the molecular mechanisms of RUNX3 inactivation in the context of cancer development and progression. (C) 2009 Elsevier B.V. All rights reserved.

Potential roles for the PIM1 kinase in human cancer - A molecular and therapeutic appraisal

In vitro experiments have shown the PIM1 kinase to have diverse biological roles in cell survival, proliferation and differentiation. In humans, PIM1 is often expressed in both normal and transformed cells. The PIM1 kinase is a true oncogene implicated in early transformation and tumour progression in haematopoietic malignancies and prostate carcinomas. it is associated with aggressive subgroups of lymphoma, is a marker of poor prognosis in prostate carcinomas and has been suggested to have a role in hormone insensitivity of prostate malignancies. PIM1 has a possible role in other carcinomas with 6p21 genomic alterations. On one hand, PIM1 (due to its role in malignancy) appears to be a promising target for drug development programmes but, on the other hand, the complexity of its molecular structure has posed challenges in the development of PIM1 inhibitors. In this review we discuss PIM1 expression in human tissues (including some new data from our laboratory), its role in human malignancies, as well as the possibilities and challenges in the development of target therapy for PIM1. (c) 2008 Elsevier Ltd. All rights reserved.

Human papillomavirus related head and neck cancer survival: A systematic review and meta-analysis

Human Papillomavirus (HPV) related oropharyngeal squamous cell carcinomas (OPSCCs) are reported to have improved prognosis and survival in comparison to other head and neck squamous cell cancers (HNSCCs). This systematic review and meta-analysis examines survival differences in HPV-positive HNSCC and OPSCC subtypes including tonsillar carcinoma in studies not previously investigated. Four electronic databases were searched from their inception till April 2011. A random effects meta-analysis was used to pool study estimates evaluating disease-specific (death from HNSCC), overall (all-cause mortality), progression-free and disease-free (recurrence free) survival outcomes in HPV-positive vs. HPV-negative HNSCCs. All statistical tests were two-sided. Forty-two studies were included. Patients with HPV-positive HNSCC had a 54% better overall survival compared to HPV-negative patients HR 0.46 (95% CI 0.37-0.57); the pooled HR for tonsillar cancer and OPSCC was 0.50 (95% CI 0.33-0.77) and HR 0.47 (95% CI 0.35-0.62) respectively. The pooled HR for disease specific survival was 0.28 (95% CI 0.19-0.40); similar effect sizes were found irrespective of the adjustment for confounders, HPV detection methods or study location. Both progression-free survival and disease-free survival were significantly improved in HPV-positive HNSCCs. HPV-positive HNSCCs and OPSCCs patients have a significantly lower disease specific mortality and are less likely to experience progression or recurrence of their cancer than HPV-negative patients; findings which have connotations for treatment selection in these patients.