Modulation of SLFN11 induces changes in DNA Damage response in breast cancer.

BACKGROUND: Lack of Schlafen family member 11 (SLFN11) expression has been recently identified as a dominant genomic determinant of response to DNA damaging agents in numerous cancer types. Thus, several strategies aimed at increasing SLFN11 are explored to restore chemosensitivity of refractory cancers. In this study, we examined various approaches to elevate SLFN11 expression in breast cancer cellular models and confirmed a corresponding increase in chemosensitivity with using the most successful efficient one. As oncogenic transcriptomic downregulation is often driven by methylation of the promotor region, we explore the demethylation effect of 5-aza-2'-deoxycytidine (decitabine), on the SLFN11 gene. Since SLFN11 has been reported as an interferon inducible gene, and interferon is secreted during an active anti-tumor immune response, we investigated the in vitro effect of IFN-γ on SLFN11 expression in breast cancer cell lines. As a secondary approach to pick up cross talk between immune cells and SLFN11 expression we used indirect co-culture of breast cancer cells with activated PBMCs and evaluated if this can drive SLFN11 upregulation. Finally, as a definitive and specific way to modulate SLFN11 expression we implemented SLFN11 dCas9 (dead CRISPR associated protein 9) systems to specifically increase or decrease SLFN11 expression. RESULTS: After confirming the previously reported correlation between methylation of SLFN11 promoter and its expression across multiple cell lines, we showed in-vitro that decitabine and IFN-γ could increase moderately the expression of SLFN11 in both BT-549 and T47D cell lines. The use of a CRISPR-dCas9 UNISAM and KRAB system could increase or decrease SLFN11 expression significantly (up to fivefold), stably and specifically in BT-549 and T47D cancer cell lines. We then used the modified cell lines to quantify the alteration in chemo sensitivity of those cells to treatment with DNA Damaging Agents (DDAs) such as Cisplatin and Epirubicin or DNA Damage Response (DDRs) drugs like Olaparib. RNAseq was used to elucidate the mechanisms of action affected by the alteration in SLFN11 expression. In cell lines with robust SLFN11 promoter methylation such as MDA-MB-231, no SLFN11 expression could be induced by any approach. CONCLUSION: To our knowledge this is the first report of the stable non-lethal increase of SLFN11 expression in a cancer cell line. Our results show that induction of SLFN11 expression can enhance DDA and DDR sensitivity in breast cancer cells and dCas9 systems may represent a novel approach to increase SLFN11 and achieve higher sensitivity to chemotherapeutic agents, improving outcome or decreasing required drug concentrations. SLFN11-targeting therapies might be explored pre-clinically to develop personalized approaches.

An integrated tumor, immune and microbiome atlas of colon cancer.

The lack of multi-omics cancer datasets with extensive follow-up information hinders the identification of accurate biomarkers of clinical outcome. In this cohort study, we performed comprehensive genomic analyses on fresh-frozen samples from 348 patients affected by primary colon cancer, encompassing RNA, whole-exome, deep T cell receptor and 16S bacterial rRNA gene sequencing on tumor and matched healthy colon tissue, complemented with tumor whole-genome sequencing for further microbiome characterization. A type 1 helper T cell, cytotoxic, gene expression signature, called Immunologic Constant of Rejection, captured the presence of clonally expanded, tumor-enriched T cell clones and outperformed conventional prognostic molecular biomarkers, such as the consensus molecular subtype and the microsatellite instability classifications. Quantification of genetic immunoediting, defined as a lower number of neoantigens than expected, further refined its prognostic value. We identified a microbiome signature, driven by Ruminococcus bromii, associated with a favorable outcome. By combining microbiome signature and Immunologic Constant of Rejection, we developed and validated a composite score (mICRoScore), which identifies a group of patients with excellent survival probability. The publicly available multi-omics dataset provides a resource for better understanding colon cancer biology that could facilitate the discovery of personalized therapeutic approaches.

Fabrication of polycaprolactone/calcium phosphates hybrid scaffolds impregnated with plant extracts using 3D printing for potential bone regeneration.

The increase in critical bone diseases and defects in the world's population increases the need for bone substitutes to restore form and function. Organic and inorganic scaffolds with antibacterial properties could provide advantages for bone regeneration. In this study, we obtained scaffolds of polycaprolactone (PCL) charged with calcium phosphates nanoparticles and impregnated with extracts of Colombian plants as an alternative for potential bone regeneration. Calcium phosphate nanoparticles were obtained via auto-combustion synthesis. The nanoparticles were incorporated into the PCL with a chemical dissolution-disperse process. The composite obtained was used to produce a filament to print Triply Periodic Minimal Surface (TPMS) based scaffolds. Such geometry facilitates cellular growth thanks to its interconnected porosity. The scaffolds were impregnated with extracts of Justicia cf colorifera (Acanthaceae), and Billia rosea (Sapindaceae) due to their ancestral medical applications. A physical and biological characterization was conducted. The process to print scaffolds with an enhanced geometry to facilitate the flux of biological fluids was successful. The scaffolds loaded with B. rosea showed strong antibacterial behavior, suggesting the presence of reported terpenoids with antibacterial properties. The approach used in this study evidenced promising prospects for bone defect repair.

Cytokine-chemokine network driven metastasis in esophageal cancer; promising avenue for targeted therapy.

Esophageal cancer (EC) is a disease often marked by aggressive growth and poor prognosis. Lack of targeted therapies, resistance to chemoradiation therapy, and distant metastases among patients with advanced disease account for the high mortality rate. The tumor microenvironment (TME) contains several cell types, including fibroblasts, immune cells, adipocytes, stromal proteins, and growth factors, which play a significant role in supporting the growth and aggressive behavior of cancer cells. The complex and dynamic interactions of the secreted cytokines, chemokines, growth factors, and their receptors mediate chronic inflammation and immunosuppressive TME favoring tumor progression, metastasis, and decreased response to therapy. The molecular changes in the TME are used as biological markers for diagnosis, prognosis, and response to treatment in patients. This review highlighted the novel insights into the understanding and functional impact of deregulated cytokines and chemokines in imparting aggressive EC, stressing the nature and therapeutic consequences of the cytokine-chemokine network. We also discuss cytokine-chemokine oncogenic potential by contributing to the Epithelial-Mesenchymal Transition (EMT), angiogenesis, immunosuppression, metastatic niche, and therapeutic resistance development. In addition, it discusses the wide range of changes and intracellular signaling pathways that occur in the TME. Overall, this is a relatively unexplored field that could provide crucial insights into tumor immunology and encourage the effective application of modulatory cytokine-chemokine therapy to EC.

IKKß Kinase Promotes Stemness, Migration, and Invasion in KRAS-Driven Lung Adenocarcinoma Cells.

KRAS oncogenic mutations are widespread in lung cancer and, because direct targeting of KRAS has proven to be challenging, KRAS-driven cancers lack effective therapies. One alternative strategy for developing KRAS targeted therapies is to identify downstream targets involved in promoting important malignant features, such as the acquisition of a cancer stem-like and metastatic phenotype. Based on previous studies showing that KRAS activates nuclear factor kappa-B (NF-κB) through inhibitor of nuclear factor kappa-B kinase ß (IKKß) to promote lung tumourigenesis, we hypothesized that inhibition of IKKß would reduce stemness, migration and invasion of KRAS-mutant human lung cancer cells. We show that KRAS-driven lung tumoursphere-derived cells exhibit stemness features and increased IKKß kinase activity. IKKß targeting by different approaches reduces the expression of stemness-associated genes, tumoursphere formation, and self-renewal, and preferentially impairs the proliferation of KRAS-driven lung tumoursphere-derived cells. Moreover, we show that IKKß targeting reduces tumour cell migration and invasion, potentially by regulating both expression and activity of matrix metalloproteinase 2 (MMP2). In conclusion, our results indicate that IKKß is an important mediator of KRAS-induced stemness and invasive features in lung cancer, and, therefore, might constitute a promising strategy to lower recurrence rates, reduce metastatic dissemination, and improve survival of lung cancer patients with KRAS-driven disease.

IKKß targeting reduces KRAS-induced lung cancer angiogenesis in vitro and in vivo: A potential anti-angiogenic therapeutic target.

OBJECTIVES: The ability of tumor cells to drive angiogenesis is an important cancer hallmark that positively correlates with metastatic potential and poor prognosis. Therefore, targeting angiogenesis is a rational therapeutic approach and dissecting proangiogenic pathways is important, particularly for malignancies driven by oncogenic KRAS, which are widespread and lack effective targeted therapies. Based on published studies showing that oncogenic RAS promotes angiogenesis by upregulating the proangiogenic NF-κB target genes IL-8 and VEGF, that NF-κB activation by KRAS requires the IKKß kinase, and that targeting IKKß reduces KRAS-induced lung tumor growth in vivo, but has limited effects on cell growth in vitro, we hypothesized that IKKß targeting would reduce lung tumor growth by inhibiting KRAS-induced angiogenesis. MATERIALS AND METHODS: To test this hypothesis, we targeted IKKß in KRAS-mutant lung cancer cell lines either by siRNA-mediated transfection or by treatment with Compound A (CmpdA), a highly specific IKKß inhibitor, and used in vitro and in vivo assays to evaluate angiogenesis. RESULTS AND CONCLUSIONS: Both pharmacological and siRNA-mediated IKKß targeting in lung cells reduced expression and secretion of NF-κB-regulated proangiogenic factors IL-8 and VEGF. Moreover, conditioned media from IKKß-targeted lung cells reduced human umbilical vein endothelial cell (HUVEC) migration, invasion and tube formation in vitro. Furthermore, siRNA-mediated IKKß inhibition reduced xenograft tumor growth and vascularity in vivo. Finally, IKKß inhibition also affects endothelial cell function in a cancer-independent manner, as IKKß inhibition reduced pathological retinal angiogenesis in a mouse model of oxygen-induced retinopathy. Taken together, these results provide a novel mechanistic understanding of how the IKKß pathway affects human lung tumorigenesis, indicating that IKKß promotes KRAS-induced angiogenesis both by cancer cell-intrinsic and cancer cell-independent mechanisms, which strongly suggests IKKß inhibition as a promising antiangiogenic approach to be explored for KRAS-induced lung cancer therapy.

Hypoxia regulates the expression of tissue factor pathway signaling elements in a rat glioma model.

Hypoxia and necrosis are fundamental features of glioma, and their emergence is critical for the rapid biological progression of this fatal tumor. The presence of vaso-occlusive thrombus is higher in grade IV tumors [glioblastoma multiforme (GBM)] compared with lower grade tumors, suggesting that the procoagulant properties of the tumor contribute to its aggressive behavior, as well as the establishment of tumor hypoxia and necrosis. Tissue factor (TF), the primary cellular initiator of coagulation, is overexpressed in GBMs and likely favors a thrombotic microenvironment. Phosphatase and tensin homolog (PTEN) loss and hypoxia are two common alterations observed in glioma that may be responsible for TF upregulation. In the present study, ST1 and P7 rat glioma lines, with different levels of aggressiveness, were comparatively analyzed with the aim of identifying differences in procoagulant mechanisms. The results indicated that P7 cells display potent procoagulant activity compared with ST1 cells. Flow cytometric analysis showed less pronounced levels of TF in ST1 cells compared with P7 cells. Notably, P7 cells supported factor X (FX) activation via factor VIIa, whereas no significant FXa generation was observed in ST1 cells. Furthermore, the exposure of phosphatidylserine on the surface of P7 and ST1 cells was investigated. The results supported the assembly of prothrombinase complexes, accounting for the production of thrombin. Furthermore, reverse transcription-quantitative polymerase chain reaction showed that CoCl2 (known to induce a hypoxic-like stress) led to an upregulation of TF levels in P7 and ST1 cells. Therefore, increased TF expression in P7 cells was accompanied by increased TF procoagulant activity. In addition, hypoxia increased the shedding of procoagulant TF-bearing microvesicles in both cell lines. Finally, hypoxic stress induced by treatment with CoCl2 upregulated the expression of the PAR1 receptor in both P7 and ST1 cells. In addition to PAR1, P7, but not ST1 cells, expressed higher levels of PAR2 under hypoxic stress. Thus, modulating these molecular interactions may provide additional insights for the development of more efficient therapeutic strategies against aggressive glioma.

Aurora kinase targeting in lung cancer reduces KRAS-induced transformation.

BACKGROUND: Activating mutations in KRAS are prevalent in lung cancer and have been causally linked to the oncogenic process. However, therapies targeted to oncogenic RAS have been ineffective to date and identification of KRAS targets that impinge on the oncogenic phenotype is warranted. Based on published studies showing that mitotic kinases Aurora A (AURKA) and B (AURKB) cooperate with oncogenic RAS to promote malignant transformation and that AURKA phosphorylates RAS effector pathway components, the aim of this study was to investigate whether AURKA and AURKB are KRAS targets in lung cancer and whether targeting these kinases might be therapeutically beneficial. METHODS: In order to determine whether oncogenic KRAS induces Aurora kinase expression, we used qPCR and western blotting in three different lung cell-based models of gain- or loss-of-function of KRAS. In order to determine the functional role of these kinases in KRAS-induced transformation, we generated KRAS-positive A549 and H358 cells with stable and inducible shRNA-mediated knockdown of AURKA or AURKB and evaluated transformation in vitro and tumor growth in vivo. In order to validate AURKA and/or AURKB as therapeutically relevant KRAS targets in lung cancer, we treated A549 and H358 cells, as well as two different lung cell based models of gain-of-function of KRAS with a dual Aurora kinase inhibitor and performed functional in vitro assays. RESULTS: We determined that KRAS positively regulates AURKA and AURKB expression. Furthermore, in KRAS-positive H358 and A549 cell lines, inducible knockdown of AURKA or AURKB, as well as treatment with a dual AURKA/AURKB inhibitor, decreased growth, viability, proliferation, transformation, and induced apoptosis in vitro. In addition, inducible shRNA-mediated knockdown of AURKA in A549 cells decreased tumor growth in vivo. More importantly, dual pharmacological inhibiton of AURKA and AURKB reduced growth, viability, transformation, and induced apoptosis in vitro in an oncogenic KRAS-dependent manner, indicating that Aurora kinase inhibition therapy can specifically target KRAS-transformed cells. CONCLUSIONS: Our results support our hypothesis that Aurora kinases are important KRAS targets in lung cancer and suggest Aurora kinase inhibition as a novel approach for KRAS-induced lung cancer therapy.

New environmentally-friendly antimicrobials and biocides from Andean and Mexican biodiversity.

Persistent application of pesticides often leads to accumulation in the environment and to the development of resistance in various organisms. These chemicals frequently degrade slowly and have the potential to bio-accumulate across the food chain and in top predators. Cancer and neuronal damage at genomic and proteomic levels have been linked to exposure to pesticides in humans. These negative effects encourage search for new sources of biopesticides that are more "environmentally-friendly" to the environment and human health. Many plant or fungal compounds have significant biological activity associated with the presence of secondary metabolites. Plant biotechnology and new molecular methods offer ways to understand regulation and to improve production of secondary metabolites of interest. Naturally occurring crop protection chemicals offer new approaches for pest management by providing new sources of biologically active natural products with biodegradability, low mammalian toxicity and environmentally-friendly qualities. Latin America is one of the world's most biodiverse regions and provide a previously unsuspected reservoir of new and potentially useful molecules. Phytochemicals from a number of families of plants and fungi from the southern Andes and from Mexico have now been evaluated. Andean basidiomycetes are also a great source of scientifically new compounds that are interesting and potentially useful. Use of biopesticides is an important component of integrated pest management (IPM) and can improve the risks and benefits of production of many crops all over the world.

Ecotin: Exploring a feasible antithrombotic profile.

Ecotin is an Escherichia coli-derived protein that can inhibit serine proteases. It has been suggested that this protein (ecotin-WT) and some of its variants could be used to develop a prototype to treat thrombosis. In this work, the effect of ecotin-WT and a variant of this protein harboring two mutations (Met84Arg and Met85Arg, ecotin-RR) were analyzed to determine their ability to prevent thrombus formation using in vivo models. Ecotins were analyzed in vitro using the coagulation tests. An in vivo venous thrombosis rat model and a pulmonary thromboembolism mouse model were used to investigate the antithrombotic activity. The bleeding time in rats using a tail-transection model was evaluated as a possible side effect caused by the administration of these proteins. Ecotin-RR was more effective in inhibiting thrombin than ecotin-WT. Both ecotins presented similar mechanisms of anticoagulation activity and were able to decrease thrombus formation. In contrast, only ecotin-RR increased survival rates in the in vivo pulmonary thromboembolism model, reinforcing the antithrombotic activity of ecotin-RR. Ecotin-WT and more so ecotin-RR showed potent antithrombotic effects, not associated with bleeding. The presented results indicate that ecotin-WT and ecotin-RR may be new scaffolds that could be used to develop anticoagulation molecules.

Expression of tissue factor signaling pathway elements correlates with the production of vascular endothelial growth factor and interleukin-8 in human astrocytoma patients.

The expression levels of tissue factor (TF), the clotting initiator protein, have been correlated with angiogenesis and the histological grade of malignancy in glioma patients. The pro-tumor function of TF is linked to a family of G protein-coupled receptors known as protease-activated receptors (PARs), which may be activated by blood coagulation proteases. Activation of PARs elicits a number of responses, including the expression of vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8). In the present study, we analyzed the expression of TF signaling pathway elements (TF, PAR1 and PAR2) and evaluated their correlation with the expression of downstream products (VEGF and IL-8) in human astrocytoma patients. Quantitative PCR (qPCR) showed a significant increase in TF expression in grade IV (glioblastoma) tumors, which was inversely correlated with the expression of the tumor-suppressor PTEN. Immunohistochemistry and qPCR analyses demonstrated a highly significant elevation in the expression of PAR1, but not PAR2, in tumor samples from high-grade astrocytoma patients. The elevated VEGF expression levels detected in the high-grade astrocytoma samples were positively correlated with TF, PAR1 and PAR2 expression. In addition, IL-8 was significantly increased in glioblastoma patients and positively correlated with TF and PAR2 expression. Further in vitro assays employing the human glioma cell lines U87-MG and HOG demonstrated that a synthetic peptide PAR2 agonist stimulated VEGF and IL-8 production. Our findings suggest a role for TF signaling pathway elements in astrocytoma progression, particularly in glioblastoma. Therefore, TF/PAR signaling elements may be suitable targets for the development of new therapies for the treatment of aggressive glioma.

Endothelial protein C receptor function in murine and human breast cancer development.

Several markers identify cancer stem cell-like populations, but little is known about the functional roles of stem cell surface receptors in tumor progression. Here, we show that the endothelial protein C receptor (EPCR), a stem cell marker in hematopoietic, neuronal and epithelial cells, is crucial for breast cancer growth in the orthotopic microenvironment of the mammary gland. Mice with a hypomorphic allele of EPCR show reduced tumor growth in the PyMT-model of spontaneous breast cancer development and deletion of EPCR in established PyMT tumor cells significantly attenuates transplanted tumor take and growth. We find expansion of EPCR(+) cancer stem cell-like populations in aggressive, mammary fat pad-enhanced human triple negative breast cancer cells. In this model, EPCR-expressing cells have markedly increased mammosphere- and tumor-cell initiating activity compared to another stable progenitor-like subpopulation present at comparable frequency. We show that receptor blocking antibodies to EPCR specifically attenuate in vivo tumor growth initiated by either EPCR(+) cells or the heterogenous mixture of EPCR(+) and EPCR(-) cells. Furthermore, we have identified tumor associated macrophages as a major source for recognized ligands of EPCR, suggesting a novel mechanism by which cancer stem cell-like populations are regulated by innate immune cells in the tumor microenvironment.

Increased expression of protease-activated receptor 1 (PAR-1) in human leukemias.

Protease-activated receptor 1 (PAR-1) is a G-protein-coupled receptor that is overexpressed in solid tumors, being associated with several pro-tumoral responses including primary growth, invasion, metastasis and angiogenesis. Expression of PAR-1 in human leukemic cell lines is reported but the status of its expression in human leukemic patients is currently unknown. In this study we evaluated the expression pattern of PAR-1 in patients with the four main types of leukemia - chronic lymphocytic leukemia subtype B (B-CLL), acute lymphoblastic leukemia subtype B (B-ALL), acute myeloid leukemia (AML) and chronic myeloid leukemia (CML). Flow cytometry analyses show that lymphocytes from B-CLL patients express this receptor at similar levels to healthy individuals. On the other hand, it was observed a significant increase in PAR-1 expression in B-ALL lymphocytes as compared to B-CLL and healthy donors. Flow cytometric and real-time PCR demonstrated a significant increase in PAR-1 expression in granulocytes from CML patients in blast phase (CML-BP) but not in chronic phase (CML-CP) as compared to healthy donors. Finally, a significant increase in PAR-1 expression has been also observed in blasts from AML (subtypes M4 and M5) patients, as compared to monocytes or granulocytes from healthy donors. We conclude that PAR-1 might play an important biological role in aggressive leukemias and might offer additional strategies for the development of new therapies.

Hospital-based surveillance of meningococcal meningitis in Salvador, Brazil.

This study aimed to describe the clinical, epidemiological and microbiological features of meningococcal meningitis in Salvador, Brazil. Between February 1996 and January 2001, a hospital-based surveillance prospectively identified cases of culture-positive meningococcal meningitis. Demographic and clinical data were collected through interview and medical chart review. Antisera and monoclonal antibodies were used to determine the serogroup and serotype:serosubtype of the isolates, respectively. Surveillance identified a total of 408 cases of meningococcal meningitis, with a case fatality rate of 8% (32/397). The mean annual incidence for the 304 culture-positive cases residing in metropolitan Salvador was 1.71 cases per 100,000 population. Infants <1 year old presented the highest incidence (14.7 cases per 100,000 population). Of the 377 serogrouped isolates, 82%, 16%, 2% and 0.3% were serogroups B, C, W135 and Y, respectively. A single serotype:serosubtype (4,7:P1.19,15) accounted for 64% of all cases. Continued surveillance is necessary to characterise strains and to define future prevention and control strategies.

Haemophilus influenzae meningitis 5 years after introduction of the Haemophilus influenzae type b conjugate vaccine in Brazil.

The long-term impact of Haemophilus influenzae type b (Hib) conjugate vaccine, introduced throughout Latin America in the late 1990s, has not been evaluated. Active surveillance for H. influenzae meningitis was performed from August 9, 1996 to August 8, 2004 in Metropolitan Salvador, Brazil. Five years after the introduction of Hib conjugate vaccine, Hib meningitis incidence decreased from 2.39 to 0.06 cases per 100,000 population (98%) overall, and from 60.9 to 3.1 cases per 100,000 population (95%) in children <1 year of age. A transient serotype replacement phenomenon was observed associated with a small increase of meningitis due to two H. influenzae type a clonal groups. These findings indicate that Hib immunization campaign has led to the virtual elimination of Hib disease in this region.

Trichomonas vaginalis: intrastrain polymorphisms within the ribosomal intergenic spacer do not correlate with clinical presentation.

Trichomoniasis presents a broad spectrum of clinical patterns ranging from asymptomatic to severe vaginitis and cervicitis. Despite its importance, very little is known about the genetic relatedness of its causative agent, Trichomonas vaginalis, and the clinical phenotypes. To address this question, analysis of restriction length polymorphism (RFLP) within the intergenic spacer of the ribosomal DNA (IGS) from 60 clinically defined isolates of T. vaginalis was performed. This is the first description of the IGS polymorphism of T. vaginalis. As expected, a considerable number of patients were asymptomatic (28%) while only 12% presented both leukorrhea and macular colpitis, the most evident symptoms of trichomoniasis. The IGS-RFLP with the use of eight restriction enzymes showed absence of correlation between the genetic relatedness of the isolates and symptomatology. Further studies are necessary to evaluate the importance of the IGS polymorphism to the parasite virulence and clinical phenotype.

A comparative evaluation of the Papanicolaou test for the diagnosis of trichomoniasis.

BACKGROUND: Trichomoniasis is the most prevalent nonviral sexually transmitted disease in humans worldwide. In addition to its pathologic implications, trichomoniasis is a risk factor for the transmission of the HIV and is associated with reproductive complications in females. Diagnosis of the disease is problematic due to inadequate accuracy of current diagnostic methods. Recently developed DNA-based techniques for detection of Trichomonas vaginalis seem to be promising alternatives. GOAL: The goal of this study was to evaluate the accuracy of the Papanicolaou test for the diagnosis of trichomoniasis by comparing polymerase chain reaction (PCR) with other current diagnostic methods. STUDY DESIGN: A total of 1008 cervicovaginal swab specimens from a randomized population attending a gynecological service were analyzed in this study. In addition to current diagnostic methods, two sets of specific primers were used for PCR detection of T vaginalis in the cervicovaginal DNA samples, with a PCR quality control. Different examiners conducted PCR and Papanicolaou analyses in a double-blind trial. RESULTS: The prevalence of trichomoniasis in this population was 6%. A considerable number of diagnostic results of the Papanicolaou test were false negative or false positive. Compared with PCR, specificity of the Papanicolaou test was 97.6%, whereas sensitivity was only 60.7%. The positive predictive value of the Papanicolaou smear was 61.7%. These results suggest that irregularly shaped parasites without clearly defined nuclei and flagella and bacteria-induced focal cytolysis limit the ability of the Papanicolaou test to detect T vaginalis. CONCLUSION: The Papanicolaou test, the most readily available cytologic method for screening sexually transmitted pathogens and cellular abnormalities in most developing countries, is inadequate for the diagnosis of trichomoniasis due to its inherent limitations. However, PCR is a highly sensitive and specific test for the diagnosis of trichomoniasis.