Effect of 2-methoxyestradiol on mammary tumor initiation and progression.

BACKGROUND: The anti-cancer agent 2-methoxyestradiol (2-ME) has been shown to have anti-proliferative and anti-angiogenic properties. Previously, the effect of 2-ME on early- and late-stage breast cancer (BC) was investigated in vivo using a transgenic mouse model (FVB/N-Tg(MMTV-PyVT)) of spontaneous mammary carcinoma. Anti-tumor effects were observed in late-stage BC with no effect on early-stage BC. Given the contrasting results obtained from the different BC stages, we have now investigated the effect of 2-ME when administered before the appearance of palpable tumors. METHODS: Each mouse received 100 mg/kg 2-ME on day 30 after birth, twice per week for 28 days, while control mice received vehicle only. Animals were terminated on day 59. Lung and mammary tissue were obtained for immunohistochemical analysis of CD163 and CD3 expression, and histological examination was performed to analyze tumor necrosis. Additionally, blood samples were collected to measure plasma cytokine levels. RESULTS: 2-ME increased tumor mass when compared to the untreated animals (p = .0139). The pro-tumorigenic activity of 2-ME was accompanied by lower CD3+ T-cell numbers in the tumor microenvironment (TME) and high levels of the pro-inflammatory cytokine interleukin (IL)-1ß. Conversely, 2-ME-treatment resulted in fewer CD163+ cells detectable in the TME, increased levels of tumor necrosis, increased IL-10 plasma levels, and low IL-6 and IL-27 plasma levels. CONCLUSION: Taken together, these findings suggest that 2-ME promotes early-stage BC development.

Ameloblastic carcinoma: A systematic review.

BACKGROUND: Ameloblastic carcinoma (AC) is the most common odontogenic malignancy, constituting approximately 30% of cases in this category. Literature is sparse on malignant odontogenic neoplasms, with a large proportion of current knowledge derived from case reports or small case series. METHODS: A systematic review of case series/case reports of AC was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) Statement guidelines. Demographic and clinical information, including duration of the lesion, location, clinical presentation and radiologic features, were analysed. Additionally, the origin of the lesion (primary/secondary), Ki-67 proliferation index, treatment performed, metastasis, tumour recurrence and prognosis were collected for analysis. RESULTS: A total of 126 studies, including 285 individual cases of AC, were included in this review. Patients presented with a near-equal distribution of painless and painful swellings. ACs presented at a median age of 45 years, with a male-to-female ratio of 1:2. The mandible was most frequently involved, with rare cases extending to involve more than one region, including crossing the midline. Although most lesions presented with poorly-demarcated borders (52.6%), unilocular lesions with well-demarcated borders (47.4%) comprised a substantial number in the sample. The proliferation index was only reported in 27 cases, with a mean score of 42% and a wide range. The probability of tumour recurrence increased, and the survival probability decreased with prolonged follow-up duration. CONCLUSION: This study provides more comprehensive, up-to-date descriptive data on these rare odontogenic malignancies, aiding clinicians and Pathologists with the diagnosis and surgeons in their management of cases.

Effect of 2-methoxyestradiol treatment on early- and late-stage breast cancer progression in a mouse model.

The prevalence of breast cancer (BC) continues to increase and is the leading cause of cancer deaths in many countries. Numerous in vitro and in vivo studies have demonstrated that 2-methoxyestradiol (2-ME) has antiproliferative and antiangiogenic effects in BC, thereby inhibiting tumour growth and metastasis. We compared the effect of 2-ME in early- and late-stage BC using a transgenic mouse model-FVB/N-Tg(MMTV-PyVT)-of spontaneously development of aggressive mammary carcinoma with lung metastasis. Mice received 100 mg/kg 2-ME treatment immediately when palpable mammary tumours were identified (early-stage BC; Experimental group 1) and 28 days after palpable mammary tumours were detected (late-stage BC; Experimental group 2). 2-ME was administered via oral gavage three times a week for 28 days after initiation of treatment, whereas control mice received the vehicle containing 10% dimethyl sulfoxide and 90% sunflower oil for the same duration as the treatment group. Mammary tumours were measured weekly over the 28 days and at termination, blood, mammary and lung tissue were collected for analysis. Mice with a tumour volume threshold of 4000 mm3 were killed before the treatment regime was completed. 2-ME treatment of early-stage BC led to lower levels of mammary tumour necrosis, whereas tumour mass and volume were increased. Additionally, necrotic lesions and anti-inflammatory CD163-expressing cells were more frequent in pulmonary metastatic tumours in this group. In contrast, 2-ME treatment of late-stage BC inhibited tumour growth over the 28-day period and resulted in increased CD3+ cell number and tumour necrosis. Furthermore, 2-ME treatment slowed down pulmonary metastasis but did not increase survival of late-stage BC mice. Besides late-stage tumour necrosis, none of the other results were statistically significant. This study demonstrates that 2-ME treatment has an antitumour effect on late-stage BC, however, with no increase in survival rate, whereas the treatment failed to demonstrate any benefit in early-stage BC.

Comparative Molecular Genetics of Odontogenic Keratocysts in Sporadic and Syndromic Patients.

Odontogenic keratocysts (OKCs) are common cysts of odontogenic origin that usually occur as a single nonsyndromic cyst in isolation (sporadic) or as syndromic multiple cysts as a manifestation of naevoid basal cell carcinoma syndrome. Alterations involving the PTCH gene are the most commonly identified factor associated with up to 85% and 84% of naevoid basal cell carcinoma syndrome and sporadic cases, respectively. Other Hedgehog pathway and non-Hedgehog pathway-associated genes have been implicated in the pathogenesis of OKCs. This pilot study used the Affymetrix OncoScan molecular assay to perform a comparative genomic analysis between 4 sporadic and 3 syndromic cases of OKC to identify molecular drivers that may be common and/or distinct in these 2 groups. The majority of alterations detected in both groups were copy number neutral loss of heterozygosity. Despite distinct molecular signatures observed in both groups, copy number neutral loss of heterozygosity alterations involving chromosome 9q affecting not only PTCH but also the NOTCH1 gene were detected in all syndromic and 3 sporadic cases. Loss of heterozygosity alterations involving 16p11.2 affecting genes not previously described in OKCs were also detected in all syndromic and 3 sporadic cases. Furthermore, alterations on 22q11.23 and 10q22.1 were also detected in both groups. Of note, alterations on 1p13.3, 2q22.1, and 6p21.33 detected in sporadic cases were absent in all syndromic cases. This study demonstrates that a more common group of genes may be affected in both groups of OKCs, whereas other alterations may be useful in distinguishing sporadic from syndromic cysts. These findings should be validated in larger OKC cohorts to improve molecular diagnosis and subsequent patient management.

Slc7a8 Deletion Is Protective against Diet-Induced Obesity and Attenuates Lipid Accumulation in Multiple Organs.

Adipogenesis, through adipocyte hyperplasia and/or hypertrophy, leads to increased adiposity, giving rise to obesity. A genome-wide transcriptome analysis of in vitro adipogenesis in human adipose-derived stromal/stem cells identified SLC7A8 (Solute Carrier Family 7 Member 8) as a potential novel mediator. The current study has investigated the role of SLC7A8 in adipose tissue biology using a mouse model of diet-induced obesity. slc7a8 knockout (KO) and wildtype (WT) C57BL/6J mice were fed either a control diet (CD) or a high-fat diet (HFD) for 14 weeks. On the HFD, both WT and KO mice (WTHFD and KOHFD) gained significantly more weight than their CD counterparts. However, KOHFD gained significantly less weight than WTHFD. KOHFD had significantly reduced levels of glucose intolerance compared with those observed in WTHFD. KOHFD also had significantly reduced adipocyte mass and hypertrophy in inguinal, mesenteric, perigonadal, and brown adipose depots, with a corresponding decrease in macrophage infiltration. Additionally, KOHFD had decreased lipid accumulation in the liver, heart, gastrocnemius muscle, lung, and kidney. This study demonstrates that targeting slc7a8 protects against diet-induced obesity by reducing lipid accumulation in multiple organs and suggests that if targeted, has the potential to mitigate the development of obesity-associated comorbidities.

Sexual Dimorphism in Changes That Occur in Tissues, Organs and Plasma during the Early Stages of Obesity Development.

Despite obesity being a major health concern, information on the early clinical changes that occur in plasma and tissues during obesity development and the influence of sexual dimorphism is lacking. This study investigated changes in tissue and organ histology, macrophage infiltration, plasma hormones, lipid, and chemokine and cytokine levels in mice fed on a high fat diet for 11-weeks. An increase in adiposity, accompanied by adipocyte hypertrophy and macrophage infiltration, was observed to be significantly greater in males than females. Important changes in cell morphology and histology were noted in the lungs, liver, kidney, spleen, and heart, which may indicate early signs for developing obesity associated comorbidities. Leptin, but not adiponectin, was significantly altered during weight gain. Additionally, leptin, but not adiposity, correlated with insulin levels. Interestingly, GM-CSF, TNFα, and IL-12 (p70) were not produced in the early stages of obesity development. Meanwhile, the production of MCP-1, IP-10, RANTES, IL-10, IL-6, KC, and IL-9 were greatly influenced by sexual dimorphism. Importantly, IL-6/IL-10 axis of anti-inflammatory cytokine regulation was observed only in females and may account for their significantly lower weight gain compared to males. This study provides new knowledge on how sexual dimorphism may influence the development of obesity and associated comorbidities.

Advancing global equity in cancer genomics - challenges and opportunities in Sub-Saharan Africa.

Developments in genomics in the last decade has improved our understanding of the role of genetics in health and disease. One area where the impact of genomics is very noticeable is in oncology, specifically in terms of diagnosis and elucidating genetic predisposition to rare and common cancers. Sub-Saharan Africa (SSA) stands to benefit from cancer genomics, given recent spikes in the incidence of various types of cancers in the region. This mini review presents, from a health and science equity perspective, how genomics could shape cancer research and clinical care in SSA. We highlight some pan-African genomics and cancer initiatives that are facilitating cancer genomics research in SSA. We conclude with recommendations on how the ideals of equity may be advanced in cancer genomics initiatives in SSA.

Adipogenesis: A Complex Interplay of Multiple Molecular Determinants and Pathways.

The formation of adipocytes during embryogenesis has been largely understudied. However, preadipocytes appear to originate from multipotent mesenchymal stromal/stem cells which migrate from the mesoderm to their anatomical localization. Most studies on adipocyte formation (adipogenesis) have used preadipocytes derived from adult stem/stromal cells. Adipogenesis consists of two phases, namely commitment and terminal differentiation. This review discusses the role of signalling pathways, epigenetic modifiers, and transcription factors in preadipocyte commitment and differentiation into mature adipocytes, as well as limitations in our understanding of these processes. To date, a limited number of transcription factors, genes and signalling pathways have been described to regulate preadipocyte commitment. One reason could be that most studies on adipogenesis have used preadipocytes already committed to the adipogenic lineage, which are therefore not suitable for studying preadipocyte commitment. Conversely, over a dozen molecular players including transcription factors, genes, signalling pathways, epigenetic regulators, and microRNAs have been described to be involved in the differentiation of preadipocytes to adipocytes; however, only peroxisome proliferator-activated receptor gamma has proven to be clinically relevant. A detailed understanding of how the molecular players underpinning adipogenesis relate to adipose tissue function could provide new therapeutic approaches for addressing obesity without compromising adipose tissue function.

The Role of Pref-1 during Adipogenic Differentiation: An Overview of Suggested Mechanisms.

Obesity contributes significantly to the global health burden. A better understanding of adipogenesis, the process of fat formation, may lead to the discovery of novel treatment strategies. However, it is of concern that the regulation of adipocyte differentiation has predominantly been studied using the murine 3T3-L1 preadipocyte cell line and murine experimental animal models. Translation of these findings to the human setting requires confirmation using experimental models of human origin. The ability of mesenchymal stromal/stem cells (MSCs) to differentiate into adipocytes is an attractive model to study adipogenesis in vitro. Differences in the ability of MSCs isolated from different sources to undergo adipogenic differentiation, may be useful in investigating elements responsible for regulating adipogenic differentiation potential. Genes involved may be divided into three broad categories: early, intermediate and late-stage regulators. Preadipocyte factor-1 (Pref-1) is an early negative regulator of adipogenic differentiation. In this review, we briefly discuss the adipogenic differentiation potential of MSCs derived from two different sources, namely adipose-derived stromal/stem cells (ASCs) and Wharton's Jelly derived stromal/stem cells (WJSCs). We then discuss the function and suggested mechanisms of action of Pref-1 in regulating adipogenesis, as well as current findings regarding Pref-1's role in human adipogenesis.

Amplification of 3q26.2, 5q14.3, 8q24.3, 8q22.3, and 14q32.33 Are Possible Common Genetic Alterations in Oral Cancer Patients.

The lack of clinical biomarkers for head and neck cancer subtypes limits early diagnosis and monitoring of disease progression. This study investigates genetic alterations in clinically identical tumor, tumor-adjacent dysplastic epithelium (TADE) and normal epithelium (NE) in five oral cancer patients to identify differences and commonalities between oral cancer, TADE and NE. A VELscope®Vx device was used to identify TADE and NE surrounding a clinical tumor for analysis of genetic alterations using the OncoScan® assay. One of the tumor samples examined was an "M" class tumor with a high confidence BRAF:p.G469A:c.1406G>C somatic mutation, which is the first to be reported in oral cancer. Another tumor showed mosaicism in genetic alterations, indicating the presence of multiple clones. Overall, each patient's tumor, TADE and NE showed a distinct genetic profile which indicates intertumoral clonal/genetic diversity. Interestingly, four tumors showed gain of 3q26.2, 5q14.3, 8q24.3, 8q22.3, 14q32.33 and loss/LOH in 9p21.3 while all TADE had LOH on 22q11.23. In addition, some genetic alterations progressed from NE through TADE into tumor in individual patients. Furthermore, no molecular event was identified that is common to all NE and/or TADE that progressed into tumor. This pilot study demonstrates the presence of genetic heterogeneity in oral tumorigenesis, and suggests that there might exist some common genetic alterations between tumors and TADE. However, this observation would need to be further investigated and validated in a larger cohort of oral cancer patients for its potential role in oral tumorigenesis.

The effect of medium supplementation and serial passaging on the transcriptome of human adipose-derived stromal cells expanded in vitro.

BACKGROUND: For adipose-derived stromal cells (ASCs) to be safe for use in the clinical setting, they need to be prepared using good manufacturing practices (GMPs). Fetal bovine serum (FBS), used to expand ASCs in vitro in some human clinical trials, runs the risk of xenoimmunization and zoonotic disease transmission. To ensure that GMP standards are maintained, pooled human platelet lysate (pHPL) has been used as an alternative to FBS. ASCs proliferate more rapidly in pHPL than in FBS, with no significant change in immunophenotype and differentiation capacity. However, not much is known about how pHPL affects the transcriptome of these cells. METHODS: This study investigated the effect of pHPL and FBS on the ASC transcriptome during in vitro serial expansion from passage 0 to passage 5 (P0 to P5). RNA was isolated from ASCs at each passage and hybridized to Affymetrix HuGene 2.0 ST arrays for gene expression analysis. RESULTS: We observed that the transcriptome of ASCs expanded in pHPL (pHPL-ASCs) and FBS (FBS-ASCs) had the greatest change in gene expression at P2. Gene ontology revealed that genes upregulated in pHPL-ASCs were enriched for cell cycle, migration, motility, and cell-cell interaction processes, while those in FBS-ASCs were enriched for immune response processes. ASC transcriptomes were most homogenous from P2 to P5 in FBS and from P3 to P5 in pHPL. FBS- and pHPL-gene-specific signatures were observed, which could be used as markers to identify cells previously grown in either FBS or pHPL for downstream clinical/research applications. The number of genes constituting the FBS-specific effect was 3 times greater than for pHPL, suggesting that pHPL may be a milder supplement for cell expansion. A set of genes were expressed in ASCs at all passages and in both media. This suggests that a unique ASC in vitro transcriptomic profile exists that is independent of the passage number or medium used. CONCLUSIONS: GO classification revealed that pHPL-ASCs are more involved in cell cycle processes and cellular proliferation when compared to FBS-ASCs, which are involved in more specialized or differentiation processes like cardiovascular and vascular development. This makes pHPL a potential superior supplement for expanding ASCs as they retain their proliferative capacity, remain untransformed and pHPL does not affect the genes involved in differentiation in specific developmental processes.

Molecular profile of tongue cancer in an 18-year-old female patient with no recognizable risk factor.

BACKGROUND: The occurrence of oral tongue squamous cell carcinoma (TSCC) in nonsmoking young adults, especially females, has increased. Yet, there is no clear evidence to support the existence of any single determinant. This case reports the presence of TSCC in an 18-year-old female with no recognizable risk factor for oral cancer development. METHODS: Histological examination and p16 immunohistochemistry were performed. Formalin-fixed paraffin-embedded sections were prepared from resected tissue and DNA was extracted for molecular OncoScan analysis. RESULTS: Histological and immunochemical analysis showed a p16-negative poorly differentiated keratinizing squamous cell carcinoma. OncoScan analysis of this tumor revealed a high confidence TP53:p.R213*:c.637C>T somatic mutation as well as copy number alterations of chromosomal regions including gains of 1p, 3q, 5p, 7p, 8p, 8q, 11q, 15q, 17q, and 20p, and losses on 1p, 3p, 18q, and 22q. CONCLUSION: The TP53:p.R213*:c.637C>T mutation detected is indicative of a genetic predisposition to cancer and it is the first to be reported in TSCC in a nonsmoking young adult. LEVEL OF EVIDENCE: Case report.

Heterozygosity of p16 expression in an oral squamous cell carcinoma with associated loss of heterozygosity and copy number alterations.

BACKGROUND: Oral field cancerization describes a multifocal development process involving many cells at once in response to prolong exposure to carcinogens. This case demonstrated differential p16 expression in different sections of the same HPV negative tumor on the floor of mouth in a patient with history of prolonged smoking. METHODS: Histological examination, presence of HPV infection and OncoScan analysis of DNA extracted from two well-defined areas with different p16 expression profiles were performed. RESULTS: Histological and immunochemical analysis revealed the presence of a dual architectural pattern squamous cell carcinoma with a p16 negative and a p16 positive component. OncoScan analysis showed genetic changes that define field cancerization of the p16 negative tumor as revealed by mosaicism in both loss of heterozygosity and copy number alterations in cancer-associated genes located on 3p, 7p, 9p 11q, and 17p. CONCLUSION: These changes were indicative of field cancerization in response to tobacco exposure.

The Role of Reactive Oxygen Species in Adipogenic Differentiation.

Interest in reactive oxygen species and adipocyte differentiation/adipose tissue function is steadily increasing. This is due in part to a search for alternative avenues for combating obesity, which results from the excess accumulation of adipose tissue. Obesity is a major risk factor for complex disorders such as cancer, type 2 diabetes, and cardiovascular diseases. The ability of mesenchymal stromal/stem cells (MSCs) to differentiate into adipocytes is often used as a model for studying adipogenesis in vitro. A key focus is the effect of both intra- and extracellular reactive oxygen species (ROS) on adipogenesis. The consensus from the majority of studies is that ROS, irrespective of the source, promote adipogenesis.The effect of ROS on adipogenesis is suppressed by antioxidants or ROS scavengers. Reactive oxygen species are generated during the process of adipocyte differentiation as well as by other cell metabolic processes. Despite many studies in this field, it is still not possible to state with certainty whether ROS measured during adipocyte differentiation are a cause or consequence of this process. In addition, it is still unclear what the exact sources are of the ROS that initiate and/or drive adipogenic differentiation in MSCs in vivo. This review provides an overview of our understanding of the role of ROS in adipocyte differentiation as well as how certain ROS scavengers and antioxidants might affect this process.

Synthetic Hemozoin (ß-Hematin) Crystals Nucleate at the Surface of Neutral Lipid Droplets that Control Their Sizes.

Emulsions of monopalmitoylglycerol (MPG) and of a neutral lipid blend (NLB), consisting of MPG, monostearoylglycerol, dipalmitoylglycerol, dioleoylglycerol and dilineoylglycerol (4:2:1:1:1), the composition associated with hemozoin from the malaria parasite Plasmodium falciparum, have been used to mediate the formation of ß-hematin microcrystals. Transmission electron microscopy (TEM), electron diffraction and electron spectroscopic imaging/electron energy loss spectroscopy (ESI/EELS) have been used to characterize both the lipid emulsion and ß-hematin crystals. The latter have been compared with ß-hematin formed at a pentanol/aqueous interface and with hemozoin both within P. falciparum parasites and extracted from the parasites. When lipid and ferriprotoporphyrin IX solutions in 1:9 v/v acetone/methanol were thoroughly pre-mixed either using an extruder or ultrasound, ß-hematin crystals were found formed in intimate association with the lipid droplets. These crystals resembled hemozoin crystals, with prominent {100} faces. Lattice fringes in TEM indicated that these faces made contact with the lipid surface. The average length of these crystals was 0.62 times the average diameter of NLB droplets and their size distributions were statistically equivalent after 10 min incubation, suggesting that the lipid droplets also controlled the sizes of the crystals. This most closely resembles hemozoin formation in the helminth worm Schistosoma mansoni, while in P. falciparum, crystal formation appears to be associated with the much more gently curved digestive vacuole membrane which apparently leads to formation of much larger hemozoin crystals, similar to those formed at the flat pentanol-water interface.

Insights into the role of heme in the mechanism of action of antimalarials.

By using cell fractionation and measurement of Fe(III)heme-pyridine, the antimalarial chloroquine (CQ) has been shown to cause a dose-dependent decrease in hemozoin and concomitant increase in toxic free heme in cultured Plasmodium falciparum that is directly correlated with parasite survival. Transmission electron microscopy techniques have further shown that heme is redistributed from the parasite digestive vacuole to the cytoplasm and that CQ disrupts hemozoin crystal growth, resulting in mosaic boundaries in the crystals formed in the parasite. Extension of the cell fractionation study to other drugs has shown that artesunate, amodiaquine, lumefantrine, mefloquine, and quinine, all clinically important antimalarials, also inhibit hemozoin formation in the parasite cell, while the antifolate pyrimethamine and its combination with sulfadoxine do not. This study finally provides direct evidence in support of the hemozoin inhibition hypothesis for the mechanism of action of CQ and shows that other quinoline and related antimalarials inhibit cellular hemozoin formation.

Neutral lipids associated with haemozoin mediate efficient and rapid ß-haematin formation at physiological pH, temperature and ionic composition.

BACKGROUND: The malaria parasite disposes of host-derived ferrihaem (iron(III)protoporphyrin IX, Fe(III)PPIX) by conversion to crystalline haemozoin in close association with neutral lipids. Lipids mediate synthetic haemozoin (ß-haematin) formation very efficiently. However, the effect on reaction rates of concentrations of lipid, Fe(III)PPIX and physiologically relevant ions and biomolecules are unknown. METHODS: Lipid emulsions containing Fe(III)PPIX were prepared in aqueous medium (pH 4.8, 37°C) to mediate ß-haematin formation. The reaction was quenched at various times and free Fe(III)PPIX measured colorimetrically as a pyridine complex and the kinetics and yields analysed. Products were also characterized by FTIR, TEM and electron diffraction. Autofluorescence was also used to monitor ß-haematin formation by confocal microscopy. RESULTS: At fixed Fe(III)PPIX concentration, ß-haematin yields remained constant with decreasing lipid concentration until a cut-off ratio was reached whereupon efficiency decreased dramatically. For the haemozoin-associated neutral lipid blend (NLB) and monopalmitoylglycerol (MPG), this occurred below a lipid/Fe(III)PPIX (L/H) ratio of 0.54. Rate constants were found to increase with L/H ratio above the cut-off. At 16 µM MPG, Fe(III)PPIX concentration could be raised until the L/H ratio reached the same ratio before a sudden decline in yield was observed. MPG-mediated ß-haematin formation was relatively insensitive to biologically relevant cations (Na(+), K(+), Mg(2+), Ca(2+)), or anions (H(2)PO(4)(-), HCO(3)(-), ATP, 2,3-diphosphoglycerate, glutathione). Confocal microscopy demonstrated ß-haematin formation occurs in association with the lipid particles. CONCLUSIONS: Kinetics of ß-haematin formation have shown that haemozoin-associated neutral lipids alone are capable of mediating ß-haematin formation at adequate rates under physiologically realistic conditions of ion concentrations to account for haemozoin formation.