Targets For More Effective Therapies Research Articles

A comparison of the microbiome composition in lower respiratory tract at different sites in early lung cancer patients.

Lung microbiome dysbiosis has been associated with lung carcinogenesis. However, the differences in the microbiome composition at different lung sites of lung cancer patients remain little understood. Studying the whole lung microbiome in cancer patients could provide new insights for interpreting the complex interplay between the microbiome and lung cancer and finding new targets for more effective therapies and preventive measures. A total of 16 patients with non-small cell lung cancer (NSCLC) were recruited for this study. Samples were obtained from four sites, including lung tumor tissues (TT), para-tumor tissues (PT), distal normal lung tissues (DN), and bronchial tissues (BT). The DNA was isolated from the tissues, and the V3-V4 regions were amplified. Sequencing libraries were generated and sequenced on an Illumina NovaSeq6000 platform. The richness and evenness of the microbiome were generally consistent among the TT, PT, DN, and BT groups in lung cancer patients. Principal coordinate analysis (PCoA) and nonmetric multidimensional scaling (NMDS) based on Bray-Curtis, weighted and unweighted UniFrac distance showed no distinct separation trend among the four groups. Proteobacteria, Firmicutes, Bacteroidota, and Desulfobacterota were the most common phyla in all four groups, while TT showed the highest abundance of Proteobacteria and the lowest abundance of Firmicutes. At the genus level, Rubellimicrobium and Fictibacillus were higher in the TT group. In the predicted functional analysis by PICRUSt, there were no specifically discrepant pathways among the four groups. In addition, an inverse relationship between body mass index (BMI) and alpha diversity was observed in this study. A non-significant result was obtained from the microbiome diversity comparison between different tissues. However, we demonstrated that lung tumors were enriched with specific bacterial species, which might contribute to tumorigenesis. Moreover, we found an inverse relationship between BMI and alpha diversity in these tissues, providing a new clue for deciphering the mechanisms of lung carcinogenesis.

Protein kinases on carbon metabolism: potential targets for alternative chemotherapies against toxoplasmosis.

The apicomplexan parasite Toxoplasma gondii is the causative agent of toxoplasmosis, a global disease that significantly impacts human health. The clinical manifestations are mainly observed in immunocompromised patients, including ocular damage and neuronal alterations leading to psychiatric disorders. The congenital infection leads to miscarriage or severe alterations in the development of newborns. The conventional treatment is limited to the acute phase of illness, without effects in latent parasites; consequently, a cure is not available yet. Furthermore, considerable toxic effects and long-term therapy contribute to high treatment abandonment rates. The investigation of exclusive parasite pathways would provide new drug targets for more effective therapies, eliminating or reducing the side effects of conventional pharmacological approaches. Protein kinases (PKs) have emerged as promising targets for developing specific inhibitors with high selectivity and efficiency against diseases. Studies in T. gondii have indicated the presence of exclusive PKs without homologs in human cells, which could become important targets for developing new drugs. Knockout of specific kinases linked to energy metabolism have shown to impair the parasite development, reinforcing the essentiality of these enzymes in parasite metabolism. In addition, the specificities found in the PKs that regulate the energy metabolism in this parasite could bring new perspectives for safer and more efficient therapies for treating toxoplasmosis. Therefore, this review provides an overview of the limitations for reaching an efficient treatment and explores the role of PKs in regulating carbon metabolism in Toxoplasma, discussing their potential as targets for more applied and efficient pharmacological approaches.

The Role of Citrate Homeostasis in Merkel Cell Carcinoma Pathogenesis.

Simple SummaryMerkel cell carcinoma (MCC) is a rare but highly aggressive skin cancer. Despite important progress, overall understanding of the events that drive MCC carcinogenesis remains incomplete. We discovered that the plasma membrane citrate transporter (pmCiC) is upregulated in Merkel cell carcinoma cell lines. Cancer cells import extracellular citrate via pmCiC to support their metabolism, which is critical to support proliferation and metastatic spread. In this study, we show that inhibition of pmCiC can decrease the growth rate of Merkel cell carcinoma cell lines. Targeting pmCiC and thereby the tumor metabolism should be considered further as a potential anti-cancer therapy.Merkel cell carcinoma (MCC) is a rare but highly aggressive tumor of the skin with a poor prognosis. The factors driving this cancer must be better understood in order to discover novel targets for more effective therapies. In the search for targets, we followed our interest in citrate as a central and critical metabolite linked to fatty acid synthesis in cancer development. A key to citrate uptake in cancer cells is the high expression of the plasma membrane citrate transporter (pmCiC), which is upregulated in the different adenocarcinoma types tested so far. In this study, we show that the pmCiC is also highly expressed in Merkel cell carcinoma cell lines by western blot and human tissues by immunohistochemistry staining. In the presence of extracellular citrate, MCC cells show an increased proliferation rate in vitro; a specific pmCiC inhibitor (Na+-gluconate) blocks this citrate-induced proliferation. Furthermore, the 3D in vivo Chick Chorioallantoic Membrane (CAM) model showed that the application of Na+-gluconate also decreases Merkel cell carcinoma growth. Based on our results, we conclude that pmCiC and extracellular citrate uptake should be considered further as a potential novel target for the treatment of Merkel cell carcinoma.

The Gut-Brain Axis.

Preclinical evidence has firmly established bidirectional interactions among the brain, the gut, and the gut microbiome. Candidate signaling molecules and at least three communication channels have been identified. Communication within this system is nonlinear, is bidirectional with multiple feedback loops, and likely involves interactions between different channels. Alterations in gut-brain-microbiome interactions have been identified in rodent models of several digestive, psychiatric, and neurological disorders. While alterations in gut-brain interactions have clearly been established in irritable bowel syndrome, a causative role of the microbiome in irritable bowel syndrome remains to be determined. In the absence of specific microbial targets for more effective therapies, current approaches are limited to dietary interventions and centrally targeted pharmacological and behavioral approaches. A more comprehensive understanding of causative influences within the gut-brain-microbiome system and well-designed randomized controlled trials are needed to translate these exciting preclinical findings into effective therapies.

Impact of chronic wounds of various etiology on systemic profiles of key inflammatory cytokines, chemokines and growth factors, and their interplay.

Non-healing wounds are becoming a growing concern for public health as a result of their increasing prevalence in progressively aging societies. The aim of this article is to evaluate the effects of wound etiology on a panel of circulating cytokines in patients with non-healing wounds of the lower extremities. This prospective case-control study involved 104 individuals: healthy elderly people (n = 46) and patients with diabetes and/or cardiovascular disease (n = 58; among them 38 with chronic wounds of venous, ischemic or neurotrophic etiology). Selected serum cytokines - i.e. IL-1β, IL-4, IL-6, IL-8, FGF-2, G-CSF, GM-CSF, MCP-1, MIP-1α, TNF-α, VEGF-A, and PDGF-BB - were measured using the Luminex platform. Compared to healthy elderly people, presence of diabetes and/or cardiovascular disease was associated with elevated IL-6, IL-8, MCP-1 and G-CSF while non-healing wounds coexisted with the increase in the levels of all examined cytokines/growth factors except for G-CSF and GM-CSF. Among diseased elderly people, having wounds was associated with increased levels of IL-1β, IL-4, IL-6, IL-8, FGF-2, MIP-1α, PDGF-BB, and VEGF-A. Interleukin 1β elevation was a sole independent predictor of chronic wounds with an odds ratio (OR) of 6.3. Cytokines in healthy seniors were loosely interrelated, while the levels of cytokines in diseased patients with wounds displayed a tight pattern of association. When stratified by their etiology, the association pattern for IL-6, IL-8, MCP-1, and VEGF-A was disrupted in neurotrophic wounds. The results presented herein may improve our understanding of the pathomechanisms which lead to chronic wounds and of the effects they exert on a systemic level, as well as providing potential targets for more effective therapies.

Opposing roles of PGD2 in GBM

BackgroundThe World Health Organization classifies glioblastoma (GBM) as a grade IV astrocytoma. Despite the advances in chemotherapy, surgery, and radiation treatments that improve a patient’s length of survival, the overall trajectory of the disease remains unchanged. GBM cells produce significant levels of various types of bioactive lipids. Prostaglandin D2 (PGD2) influences both pro- and anti-tumorigenic activities in the cell; however, its role in GBM is unclear. Therefore, this study aimed to identify the impact of PGD2 on GBM cell activities in vitro. MethodsFirst we looked to identify the presence of the PGD2 synthesis pathway through RT-PCR, immunohistochemistry, and HPLC–MS/MS in three GBM cell lines. Then, to observe PGD2’s effects on cell count and apoptosis/mitosis (Hoechst 33342 stain), and migration (Transwell Assay), the cells were treated in vitro with physiological (<1μM) and/or supraphysiological (>1μM) concentrations of PGD2 over 72h. HPLC–MS/MS was used to identify the lipid composition of patients with either Grade II/III gliomas or GBM. ResultsWe identified the presence of endogenous PGD2 with its corresponding enzymes and receptors. Exogenous PGD2 both increased cell count (<1μM) and decreased cell count (10μM) in a concentration-dependent manner. There were no significant effects on apoptosis. A significant decrease in mitotic activity was seen only in U251MG, and a significant increase was seen in migration with 5μM PGD2 treatments. A very significant increase of PGD2 was seen from Grade II/III gliomas to GBM. ConclusionsOur study demonstrates that prostaglandin D2 possesses a dynamic, concentration-dependent effect in GBM cell activities. The increase of PGD2 production in GBM patients suggests a pro-tumorigenic role of PGD2 in glioma growth and invasion. Therefore, prostaglandin signaling in GBM requires further investigation to identify new targets for more effective therapies.

Arrived safely? Audit of documentation of successful transfer into adult care of young people attending paediatric rheumatology clinics

Arrived safely? Audit of documentation of successful transfer into adult care of young people attending paediatric rheumatology clinics

A118: Laboratory Investigation of the Role of Toll‐Like Receptors on Kidney Cells in Pathogenesis of Lupus Nephritis

Background/Purpose:Lupus nephritis (LN) is a serious complication of juvenile‐onset systemic lupus erythematosus (JSLE). Current treatments include longterm immunosuppressants with significant side effects. There is a need to identify targets for more effective therapies. Toll‐like receptors (TLRs) perform an important role in the innate immune response by recognising conserved molecules associated with pathogens. Previous studies have suggested a role for TLR7 and TLR9 in lymphocytes in the pathogenesis of SLE. Podocytes are specialised cells forming an important part of the glomerular filtration barrier. Biopsies from LN patients have demonstrated higher TLR7 and TLR9 expression in glomeruli compared with controls and suggest that these TLRs are localised to podocytes. We hypothesise that stimulation of TLR7 and/or TLR9 in podocytes, acting via nuclear factor kappa B (NFkB), leads to cellular damage and resultant kidney disease. This research aims to examine the role of TLR7 and TLR9 in podocytes to identify potential targets for more effective therapies of lupus nephritis.Methods:Conditionally‐immortalised human podocyte cell lines were cultured to examine expression of TLRs and the effects of their activation on the NFkB pathway and cell proliferation. Reverse transcriptionquantitative polymerase chain reaction (RT‐qPCR) and sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS‐PAGE) with Western blotting were used to detect TLR7 and TLR9 expression at the mRNA and protein level respectively. The effects of lipopolysaccharide (LPS), an inflammatory stimulus, on TLR expression were examined. Podocytes were stimulated with imiquimod, a specific TLR7 agonist and CpG oligodeoxynucleotide (ODN) 2216, a specific TLR9 agonist. Phosphorylation of NFkB pathway components was assessed with Western blotting and a cell proliferation assay used to estimate cell survival following treatments. The effects of treatment with TLR agonists in combination with dexamethasone were also examined.Results:Treatment of podocytes with LPS was associated with increased expression of TLR7 at both mRNA and protein levels whereas there was comparatively little change in TLR9. Exposure of the cells to imiquimod or CpG ODN 2216 increased phosphorylation of NFkB. Initial results from the cell proliferation assay suggested lower levels after imiquimod or CpG ODN 2216 treatment for 24 hours. Since podocytes are terminally differentiated this implies reduced cell numbers or lower metabolic activity. Exposure of the podocytes to dexamethasone was associated with lower expression of TLR7 and TLR9 at the mRNA level. There was preliminary evidence of less phosphorylation of NFkB when cells were treated with dexamethasone prior to TLR agonists compared with the latter alone.Conclusion:This study suggests that podocytes express TLR7 and TLR9. Agonists of these receptors have effects on intracellular signalling. If confirmed through ongoing work, the TLR‐NFkB pathway in kidney cells may be a potential target for novel therapies in lupus nephritis.

PReS-FINAL-2354: Laboratory investigation of the role of toll-like receptros on kidney cells in pathogenesis of lupus nephritis

Lupus nephritis (LN) is a serious complication of juvenile-onset systemic lupus erythematosus (JSLE). Current treatments include long-term immunosuppressants with significant side effects. There is a need to identify targets for more effective therapies. Toll-like receptors (TLRs) perform an important role in the innate immune response by recognising conserved molecules associated with pathogens. Previous studies have suggested a role for TLR7 and TLR9 in lymphocytes in the pathogenesis of SLE. Podocytes are specialised cells forming an important part of the glomerular filtration barrier. Biopsies from LN patients have demonstrated higher TLR7 and TLR9 expression in glomeruli compared with controls and suggest that these TLRs are localised to podocytes. We hypothesise that stimulation of TLR7 and/or TLR9 in podocytes, acting via nuclear factor kappa B (NFkB), leads to cellular damage and resultant kidney disease.

The frequency of KRAS and BRAF mutations in intrahepatic cholangiocarcinomas and their correlation with clinical outcome

The incidence of intrahepatic cholangiocarcinoma is increasing worldwide. The prognosis of intrahepatic cholangiocarcinoma is poor, and a better understanding of intrahepatic cholangiocarcinoma tumor biology is needed to more accurately predict clinical outcome and to suggest potential targets for more effective therapies. v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) and BRAF are frequently mutated oncogenes that promote carcinogenesis in a variety of tumor types. In this study, we analyze a large set of intrahepatic cholangiocarcinoma tumors (n = 54) for mutations in these genes and compare the clinical outcomes of wild type versus KRAS and BRAF mutant cases. Of 54 cases, 7.4% were mutant for KRAS, 7.4% were mutant for BRAF, and these were mutually exclusive. These mutant cases were associated with a higher tumor stage at time of resection and a greater likelihood of lymph node involvement. These cases were also associated with a worse long-term overall survival. Therefore, testing for KRAS and BRAF mutations could be a valuable adjunct in improving both prognosis and outcome stratification among patients with intrahepatic cholangiocarcinoma.

A measure of control for secondary cytokine-induced injury of articular cartilage: A computational study

In previous works, the author and collaborators establish a mathematical model for injury response in articular cartilage. In this paper we use mathematical software and computational techniques, applied to an existing model to explore in more detail how the behavior of cartilage cells is influenced by several of, what are believed to be, the most significant mechanisms underlying cartilage injury response at the cellular level. We introduce a control parameter, the radius of attenuation, and present some new simulations that shed light on how inflammation associated with cartilage injuries impacts the metabolic activity of cartilage cells. The details presented in the work can help to elucidate targets for more effective therapies in the preventative treatment of post-traumatic osteoarthritis.

The HUPO Human Proteome Project (HPP), a Global Health Research Collaboration.

The global Human Proteome Project (HPP) was announced by the Human Proteome Organization (HUPO) at the 2010 World Congress of Proteomics in Sydney, Australia, and launched at the 2011 World Congress of Proteomics in Geneva, Switzerland, with analogies to the highly successful Human Genome Project. Extensive progress was reported at the September 2012 World Congress in Boston, USA. The HPP is designed to map the entire human proteome using available and emerging technologies.The HPP aims to create a molecular and biological foundation for improving health globally through better understanding of disease processes, more accurate diagnoses, and targets for more effective therapies and preventive interventions against many diseases. There are opportunities for individual investigators everywhere to access advanced datasets and to join HPP research teams.

Abstract 5104: Identification of key tumorigenic pathways in pancreatic adenocarcinoma patients using massively parallel DNA and RNA sequencing

Abstract Purpose: Pancreatic adenocarcinoma is among the most lethal forms of all malignancies. While significant research has implicated multiple genes in disease pathogenesis, identification of therapeutic leads has been difficult. The majority of currently available therapies provide only marginal benefit. To address this issue, our goal was to assess global somatic events across multiple pancreatic tumors by using next-generation sequencing technologies (NGS). Experimental Design: We performed whole genome sequencing (WGS) on paired tumor and normal samples from 3 pancreatic adenocarcinoma patients. Additionally, we performed whole transcriptome sequencing (WTS) on 2 of these patients’ tumors for which tumor RNA was available. Results: We generated on average 132 billion mappable bases across all patients using WGS, and identified 140 somatic coding events including point mutations, insertion/deletions, and chromosomal copy number variants. Data from WGS and WTS were integrated and analyzed to identify the most heavily affected pancreatic cancer pathways across all patients. The KRAS signaling pathway was identified as the most significant map. Unbiased global pathway analysis was also performed to identify processes that may be impacted. Consistent with the previous analysis, the most significant pathway from this analysis is also KRAS signaling. This integrated analysis specifically identifies heavily affected pathways in pancreatic cancer and pinpoints genes and processes that may be considered as targets for more effective therapies. Conclusions: While sequencing of additional patients is necessary to strengthen selection of key pathways, we demonstrate the utility of using NGS to understand the processes driving pancreatic adenocarcinoma so that standardized treatments for patients may be improved. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5104. doi:1538-7445.AM2012-5104

Cyclooxygenase-2 expression is not associated with clinical outcome in synovial sarcoma

Several studies have identified cyclooxygenase-2 (COX-2) expression in a variety of sarcomas, including rhabdomyosarcoma, osteosarcoma and chondrosarcoma. Although overexpression of COX-2 has been associated with poor prognosis and decreased survival in chondrosarcoma and osteosarcoma, no relationship between COX-2 expression and patient outcome has been demonstrated in rhabdomyosarcoma or adult soft tissue sarcomas. Little is known concerning the expression of COX-2 in synovial sarcoma. Therefore, the aim of this study was to examine the expression of COX-2 in synovial sarcoma and if shown, to identify any association with tumor stage and oncologic outcome. Paraffin-embedded specimens from 27 patients with synovial sarcoma who were treated with surgical resection or biopsy were obtained. Specimens were evaluated for the degree of COX-2 expression after immunohistochemical staining. Specimens were assigned an immunoreactivity score (IS) based on the percent positivity of the specimen. A retrospective chart analysis was performed to determine the clinical stage at presentation, incidence of local recurrence, presence of metastatic disease and overall survival. Statistical analysis was then performed to determine whether there was a significant relationship between IS and stage at presentation or patient outcomes. COX-2 expression was detected in 18 of 27 (66.67%) of the pathological specimens. There was a statistically significant relationship between COX-2 expression and patient clinical stage at presentation; however, we were unable to identify a significant relationship between IS and patient survival. We also found no significant relationship between IS and development of metastases or local recurrence. COX-2 was expressed to some degree in 67% of the tumor specimens. There was a significant relationship between IS and patient stage at presentation, but no significant relationship between COX-2 expression and clinical outcome could be identified. The fact that these tumors do express COX-2, however, suggests the potential for an additional target for more effective therapy.

Reduced Stratifin Expression Can Serve As an Independent Prognostic Factor for Poor Survival in Patients with Esophageal Squamous Cell Carcinoma

Stratifin plays an important role in cancer biology by interfering with intracellular signalling pathways and cell-cycle checkpoints. Decreased expression of stratifin gene has been reported to be a poor prognostic indicator in a variety of human malignant tumors. To clarify the role and prognostic significance of stratifin in esophageal squamous cell carcinoma (ESCC). The alteration of stratifin messenger RNA (mRNA) and protein was analyzed by reverse-transcription and quantitative real-time polymerase chain reaction (QRT-PCR) and Western blotting in 20 paired ESCC and nonneoplastic esophageal mucosa tissues, respectively. Then, immunohistochemistry (IHC) was used to evaluate expression of stratifin in tissues of 148 ESCC patients (including the former 20 pairs of tissues) and correlate it with clinicopathological parameters and prognosis of ESCC patients. The stratifin level of mRNA and protein was markedly downregulated in ESCC tissue compared with in corresponding nonneoplastic esophageal epithelium (P<0.05). Similarly, the positive rate of stratifin protein expression was lower in the esophageal cancer than in paired nonneoplastic esophageal epithelium as detected by IHC (P=0.007). Statistically, the downregulation of stratifin expression was correlated with tumor infiltration depth (P=0.003), lymph node metastasis (P=0.008), distant metastasis (P=0.013), and lymphovascular invasion (P=0.007) of ESCC. Furthermore, the reduced stratifin expression was associated with shorter 5-year survival rate of ESCC patients after curative surgery (P<0.0001). On the basis of univariate and multivariate Cox regression analysis, we found that reduced stratifin expression, T4 stage, lymph node metastasis, and distant metastasis were independent risk factors for worse prognosis in ESCC patients. The present report indicates that stratifin could be a useful indicator for prognosis of this disease, as well as a potential target for more effective therapy.

Relationship of age and exercise performance in patients with heart failure: The HF-ACTION study

More than three fourths of patients with heart failure (HF) are 65 years and older, and older age is associated with worse symptoms and prognoses than is younger age. Reduced exercise capacity is a chief HF complaint and indicates poorer prognosis, especially among elderly persons, but the mechanisms underlying functional decline in older patients with HF are largely unknown. Baseline cardiopulmonary exercise testing data from the HF-ACTION trial were assessed to clarify age effects on peak oxygen consumption (VO(2)) and ventilation-carbon dioxide production (VE/VCO(2)) slope. Among 2,331 New York Heart Association class II-IV patients with HF, increased age corresponded to decreased peak VO(2) (-0.14 mL kg(-1) min(-1) per year >40 years; P < .0001) and increased VE/VCO(2) slope (0.30 U/y >70 years; P < .0001). In a multivariable model with 34 other potential determinants, age was the strongest independent predictor of peak VO(2) (partial R(2) 0.130, total R(2) 0.392; P < .001) and a significant but relatively weaker predictor of VE/VCO(2) slope (partial R(2) 0.037, total R(2) 0.199; P < .001). Blunted peak heart rate was also a strong predictor of peak VO(2). Although peak heart rate and age were strongly correlated, both were significant independent predictors of peak VO(2) when analyzed simultaneously in a model. Aggregate comorbidity increased significantly with age but did not account for age effects on peak VO(2). Age is the strongest predictor of peak VO(2) and a significant predictor of VE/VCO(2) slope in the HF-ACTION population. Age-dependent comorbidities do not explain changes in peak VO(2). Age-related changes in cardiovascular physiology, potentially magnified by the HF disease state, should be considered a contributor to the pathophysiology and a target for more effective therapy in older patients with HF.

The B cell in systemic lupus erythaematosus: a rational target for more effective therapy

Current treatment options for systemic lupus erythaematosus (SLE) are diverse and poorly defined, and aggressive therapy can be associated with serious toxicity and tolerability issues. There is, therefore, a need...

Androgen Receptor Remains Critical for Cell-Cycle Progression in Androgen-Independent CWR22 Prostate Cancer Cells

The majority of prostate cancers (PCa) that relapse after androgen deprivation therapy (androgen-independent PCa) continue to express androgen receptor (AR). To study the functional importance of AR in these tumors, we derived androgen-independent CWR22 PCa xenografts in castrated mice and generated a cell line from one of these xenografts (CWR22R3). Similarly to androgen-independent PCa in patients, the relapsed xenografts and cell line expressed AR and were resistant to treatment with bicalutamide. However, expression of the AR-regulated PSA gene in the CWR22R3 cell line was markedly decreased compared to the relapsed xenografts in vivo. Transfections with androgen-regulated reporter genes further indicated that the cells lacked androgen-independent AR transcriptional activity and were not hypersensitive to low androgen concentrations despite constitutive activation of the Erk/MAP kinases. Nonetheless, AR remained essential for androgen-independent growth because retroviral shRNA-mediated AR down-regulation resulted in marked long-term growth suppression. This was associated with increased levels of p27(kip1) and hypophosphorylation of retinoblastoma protein but not with decreases in D-type cyclin levels or MAP kinase activation. These results reveal a potentially critical function of AR in androgen-independent PCa that is distinct from its previously described transcriptional or nontranscriptional functions.

Predictive and prognostic markers in gastrointestinal cancers.

As the roles of specific genes and their functions in cancer etiology and progression become defined, the necessity of looking at multiple genes and their interaction becomes clearer. In gastrointestinal cancer, tumor suppressor and promoter genes, cell cycle control and angiogenic factors, repair genes, and metabolic genes have been examined as they relate to prognosis and therapeutic outcomes. To date, most correlative studies for specific molecular parameters have been done retrospectively. Methodologies, sample sizes, and definitions differ. The following analysis provides a brief review of the current state of knowledge and recent research on the most commonly studied molecular markers, and the newer areas of inquiry attempting to define how to use molecular-genetic features of individual tumors as targets for more effective therapies. Consideration is given to the design of trials employing molecular targets as outcome factors.