Mamoun Ahram

Unraveling the role of proteases in cancer

Investigators have been studying the expression and activity of proteases in the final steps of tumor progression, invasion and metastasis, for the past 30 years. Recent studies, however, indicate that proteases are involved earlier in progression, e.g., in tumor growth both at the primary and metastatic sites. Extracellular proteases may co-operatively influence matrix degradation and tumor cell invasion through proteolytic cascades, with individual proteases having distinct roles in tumor growth, invasion, migration and angiogenesis. In this review, we use cathepsin B as an example to examine the involvement of proteases in tumor progression and metastasis. We discuss the effect of interactions among tumor cells, stromal cells, and the extracellular matrix on the regulation of protease expression. Further elucidation of the role of proteases in cancer will allow us to design more effective inhibitors and novel protease-based drugs for clinical use.

Post-analysis follow-up and validation of microarray experiments

Measurement of gene-expression profiles using microarray technology is becoming increasingly popular among the biomedical research community. Although there has been great progress in this field, investigators are still confronted with a difficult question after completing their experiments: how to validate the large data sets that are generated? This review summarizes current approaches to verifying global expression results, discusses the caveats that must be considered, and describes some methods that are being developed to address outstanding problems.

Evaluation of Non-Formalin Tissue Fixation for Molecular Profiling Studies

Using a general strategy for evaluating clinical tissue specimens, we found that 70% ethanol fixation and paraffin embedding is a useful method for molecular profiling studies. Human prostate and kidney were used as test tissues. The protein content of the samples was analyzed by one-dimensional gel electrophoresis, immunoblot, two-dimensional gel electrophoresis, and layered expression scanning. In each case, the fixed and embedded tissues produced results similar to that obtained from snap-frozen specimens, although the protein quantity was somewhat decreased. Recovery of mRNA was reduced in both quantity and quality in the ethanol-fixed samples, but was superior to that obtained from formalin-fixed samples and sufficient to perform reverse transcription polymerase chain reactions. Recovery of DNA from ethanol-fixed specimens was superior to formalin-fixed samples as determined by one-dimensional gel electrophoresis and polymerase chain reaction. In conclusion, specimens fixed in 70% ethanol and embedded in paraffin produce good histology and permit recovery of DNA, mRNA, and proteins sufficient for several downstream molecular analyses. Complete protocols and additional discussion of relevant issues are available on an accompanying website (http://cgap-mf.nih.gov/).

Proteomic analysis of human prostate cancer

Proteomics is a promising approach in the identification of proteins and biochemical pathways involved in tumorigenesis. In an effort to discover such proteins and pathways that are deregulated in prostate tumorigenesis, cellular proteomes of matched normal prostate epithelial cells and high‐grade prostate cancer cells were analyzed by tissue microdissection, two‐dimensional electrophoresis, and mass spectrometry. Forty protein alterations were detected in the tumors; however, the majority of these changes were not shared among the 12 neoplasms. In contrast, parallel cDNA microarray analysis identified a number of common gene expression changes. The marked heterogeneity of the observed protein alterations may have significance with regard to tumor biology and research strategies for molecular profiling analyses of human prostate cancer. Published 2002 Wiley‐Liss, Inc.

Public support and consent preference for biomedical research and biobanking in Jordan

The success of any biobank depends on a number of factors including public’s view of research and the extent to which it is willing to participate in research. As a prototype of surrounding countries, public interest in research and biobanking in addition to the influence and type of informed consent for biobanking were investigated in Jordan. Data were collected as part of a national survey of 3196 individuals representing the Jordanian population. The majority of respondents (88.6%) had a positive perception of the level of research in Jordan and they overwhelmingly (98.2%) agreed to the concept of investing as a country in research. When respondents were asked if the presence of an informed consent would influence their decision to participate in biobanking, more individuals (19.8%) considered having an informed consent mechanism as a positive factor than those who considered it to have negative connotations (13.1%). However, a substantial portion (65%) did not feel it affected their decision. The majority of survey participants (64%) expressed willingness to participate in biobanking and over 90% of them preferred an opt-in consent form whether general (75.2%) or specific for disease or treatment (16.9%). These results indicate a promising ground for research and biobanking in Jordan. Educational programs or mass awareness campaigns to promote participation in biobanking and increase awareness about informed consent and individual rights in research will benefit both the scientific community as well as the public.

Factors influencing public participation in biobanking

A diversity of public participation in biobanking is a highlight for the success of biobanks. It was previously reported that only two-thirds of Jordanians expressed interest in biobanking. To promote public involvement in a biobank, it is imperative to determine the aspects that influence the decision-making to participate. On the basis of a national survey involving 3196 respondents, the influence of 13 biobanking factors was assessed, including returning research results, privacy, freedom of choice, uncertainties about research, monetary and health considerations, and personal belief. Perception toward each factor was also correlated with willingness to participate in a biobank. A considerable number of respondents indicated returning research results as influential in their decision to become biobank donors. Interestingly, whereas the positive perception of availability of general results (39%) correlated with willingness to donate for biobanking, the negative view of unavailability of individualized results (47%) did not correlate with unwillingness. Religious permission of sample donation for research had the strongest positive influence (61%) and the highest correlation to participate among positively perceived factors. Unspecified research was highly indicated as a negative factor (45%), but did not correlate with unwillingness to become a biobank donor, whereas allowed withdrawal had a positive effect (31%) and correlation to contribute to biobanking. The negative perception of accessing medical information (9.5%) and re-contact (8.5%) had the strongest correlation with unwillingness to donate to a biobank. These results may provide an insight into how to formulate strategies to promote public participation in biomedical research and biobanking.

Large-scale proteomic analysis of membrane proteins.

Proteomic analysis of membrane proteins is a promising approach for the identification of novel drug targets and/or disease biomarkers. Despite notable technological developments, obstacles related to extraction and solublization of membrane proteins are encountered. A critical discussion of the different preparative methods of membrane proteins is offered in relation to downstream proteomic applications, mainly gel-based analyses and mass spectrometry. Frequently, unknown proteins are identified by high-throughput profiling of membrane proteins. In search for novel membrane proteins, analysis of protein sequences using computational tools is performed to predict the presence of transmembrane domains. This review also presents these bioinformatic tools with the human proteome as a case study. Along with technological innovations, advancements in the areas of sample preparation and computational prediction of membrane proteins will lead to exciting discoveries.

Association between human herpesvirus 6 and occurrence of multiple sclerosis among Jordanian patients.

Objectives –  Multiple sclerosis (MS) is hypothesized to be caused by an infectious agent that initiates an autoimmune reaction. Among the infectious agents linked to MS is human herpesvirus 6 (HHV‐6). Due to the high occurrence of MS among Jordanian population and the deficiency of MS studies in Jordan, the prevalence of HHV‐6 in sera and cerebrospinal fluids (CSFs) of 36 MS patients was investigated.

Exon 2 of human cathepsin B derives from an Alu element

Transcripts for the cysteine protease cathepsin B are alternatively spliced in the untranslated regions (UTRs). We show that a cathepsin B probe containing 5′‐UTR sequences hybridized to an RNA of ∼300 nt in addition to the typical 2.2 and 4.0 kbp mRNAs. Within this 5′‐UTR, exon 2 was found to be homologous to Alu repetitive elements. Specifically, exon 2 was part of an Alu element interspersed with the cathepsin B gene. The ∼300 nt band that hybridized to our cathepsin B probe likely corresponds to Alu transcripts, which are known to accumulate in human cells. Indeed, a similarly migrating band was detected with an authentic Alu probe. Thus, we suggest that primary transcripts for cathepsin B contain Alu sequences which are preserved as exon 2 in some fully spliced mRNAs.

Pattern of cigarette and waterpipe smoking in the adult population of Jordan.

An increasing prevalence of smoking in Jordan has been noted and this necessitates understanding the social behaviours that have lead to this habit. This cross-sectional, descriptive study determined the prevalence of smoking among Jordanians in relation to demographics with a focus on 4 types of tobacco products-cigarettes, waterpipes, cigars and pipes. A population sample of 3196 adults aged 18+ years answered an interview questionnaire about their smoking habit and types and amounts of tobacco consumed. Overall 32.3% of the respondents reported being current smokers (54.9% of males and 8.3% of females) and 2.9% were ex-smokers. Cigarette smoking was the most frequent type of tobacco smoking (93.0%) and one-third of cigarette smokers consumed more than 20 cigarettes per day. Waterpipe smoking was the second most common habit (8.6%) and was significantly associated with lower age in both males and females. Reducing the high rate of smoking in Jordan must be a priority.