Liquid biopsy assay for lung carcinoma using centrifuged supernatants from fine-needle aspiration specimens.

INTRODUCTION: Tumor mutation profiling is standard-of-care in lung carcinoma patients. However, comprehensive molecular profiling of small specimens, including core needle biopsy (CNB) and fine-needle aspiration (FNA) specimens, may often be inadequate due to limited tissue. Centrifuged FNA supernatants, which are typically discarded, have emerged recently as a novel liquid-based biopsy for molecular testing. In this study, we evaluate the use of lung carcinoma FNA supernatants for detecting clinically relevant mutations. METHODS: Supernatants from lung carcinoma FNA samples (n = 150) were evaluated. Samples were further analyzed using next-generation sequencing (NGS) and ultrasensitive droplet digital PCR (ddPCR). Mutation profiles in a subset of samples were compared with results derived from paired tissue samples from the same patient (n = 67) and available plasma liquid biopsy assay (n = 45). RESULTS: All 150 samples yielded adequate DNA and NGS were carried out successfully on 104 (90%) of 116 selected samples. Somatic mutations were detected in 82% of the samples and in 50% of these patients a clinically relevant mutation was identified that would qualify them for targeted therapy or a clinical trial. There was high overall concordance between the mutation profiles of supernatants and the corresponding tissue samples, with 100% concordance with concurrent FNA and 96% with concurrent CNB samples. Comparison of actionable driver mutations detected in supernatant versus plasma samples showed 84% concordance. CONCLUSIONS: FNA supernatants can provide a valuable specimen source for genotyping lung carcinoma especially in patients with insufficient tumor tissue, thereby reducing multigene mutation profiling failure rates, improving turnaround times, and avoiding repeat biopsies.

Functional impact of Galectin-3 and TRAIL expression in breast cancer cells.

OBJECTIVE: To examine the expression of Galectin-3 and TRAIL in breast cancer tissue and their effects on the proliferation and apoptosis of breast cancer cells. PATIENTS AND METHODS: Breast cancer and normal adjacent tissue were collected from 120 patients pathologically diagnosed with breast cancer who underwent a modified radical mastectomy. SP method of immunohistochemistry was used to detect the expression levels of Galectin-3 and TRAIL in breast cancer tissues and normal adjacent tissues. The correlation between the expressions of Galectin-3 and TRAIL, and clinical prognosis of breast cancer were analyzed. Breast cancer cells were transfected with Galectin-3 siRNA and TRAIL overexpression constructs. Cell proliferation was measured by XTT method, and apoptosis was detected by flow cytometry. RESULTS: Higher Galectin-3 level and lower TRAIL level were found in breast cancer tissues compared with those in normal adjacent tissues (p < 0.001). High expression level of Galectin-3 and low expression level of TRAIL were found to be positively correlated with the shorter median survival time and overall survival time. Galectin-3 silencing by siRNA interference and TRAIL overexpression significantly decreased cell viability of MDA-MB-231 and increased the number of apoptotic cells. CONCLUSIONS: The expression level of Galectin-3 in breast cancer tissues was significantly increased compared with that in normal tissues, while the level of TRAIL protein was significantly decreased in cancer tissue. The biological role of these two proteins seems to be synergistic in inhibiting apoptosis of cancer cells. Therefore, the evaluation method that combined both Galectin-3 and TRAIL is of great clinical value in the evaluation of clinical prognosis of patients with breast cancer.

Release of cytokines by human nasal epithelial cells and peripheral blood mononuclear cells infected with Mycoplasma pneumoniae.

Mycoplasma pneumoniae (Mp) infection is associated with asthma exacerbation in children. We hypothesized that Mp infection may cause airway inflammation by inducing the release of cytokines by respiratory epithelial cells. The levels of chemokines interleukin-8 (IL-8) and released upon activation, normal t cell expressed and secreted (RANTES) released by nasal epithelial cell (NEC) cultures established from asthmatic and nonasthmatic children were measured by ELISA at 4, 24, 48, and 72 hr after cells were inoculated with Mp, and were compared with baseline release of these factors. The presence of MP on apical membranes of NEC after infection was confirmed by transmission electron microscopy, and adherence was shown to be inhibited by erythromycin. Mp infection did not alter NEC release of IL-8 or RANTES at any time point. In contrast, tumor necrosis factor alpha (TNF-alpha) stimulated increased IL-8 at all time points, and respiratory syncytial virus (RSV) infection stimulated RANTES release at 48 and 72 hr by NEC. These results were not significantly different between NEC from asthmatic and nonasthmatic children. As a comparison, peripheral blood mononuclear cells from normal human volunteers were also incubated with Mp and had significantly increased release of IL-2, IL-6, and TNF-alpha. We conclude that Mp, unlike viral pathogens such as RSV, is unlikely to directly stimulate early airway surface cytokine responses via mechanisms involving epithelial cells. We speculate that the chronic presence of mononuclear cells at the airway surface of asthmatics provides a target for Mp-triggered cytokine production.

[A case-control study on the risk factors for preterm delivery].

The risk factors for preterm delivery were analyzed with 1:2 matched case-control study by conditional logistic regression analysis. The result showed that the main risk factors for preterm delivery were neuroticism scores, premature rupture of the membranes, husband's smoking frequency during the third trimester of pregnancy, pregnancy induced hypertension, working strength and first-trimester vaginal bleeding. It is indicated that an examination before delivery and gaining weight during pregnancy are helpful to decrease the occurrence of preterm delivery.

Efficient gene transfer to human endothelial cells using DNA complexed to adenovirus particles.

Human endothelial cells have been found to be relatively refractory to various methods of DNA transfection currently in common use. By using a transfection method involving DNA complexed with replication-deficient adenovirus particles, we have shown that 20% of a population of cultured endothelial cells can be transfected and high levels of transient expression achieved. Both early-passage human umbilical vein endothelial cells and the continuous differentiated line of human endothelium-derived EA.hy926 cells are responsive to this method of transfection. Efficient DNA transfection of endothelial cells is important for studies of endothelium-specific promoters and is a potentially useful route for transgenic therapy.

Fate of Schistosoma mansoni in Biomphalaria obstructa.

Histological sections of Biomphalaria obstructa snails exposed to miracidia of Schistosoma mansoni revealed that although viable sporocysts occurred in 6 of 9 snails at 3 days postexposure (DPE), all were dead by 7 DPE. Most dead sporocysts appeared to have degenerated slowly rather than having been killed by host hemocytic responses, which were minimal. What appeared to be amorphous remnants of sporocysts could still be found at 31 DPE. In 7 of 10 snails infected simultaneously with S. mansoni and Echinostoma paraensei, viable schistosome sporocysts occurred at 7 DPE, possibly as a result of interference with hemocyte function by the echinostome. However, in snails exposed to E. paraensei 48 hr prior to S. mansoni, no viable schistosome sporocysts were found at 7 DPE. Biomphalaria obstructa may be only temporarily susceptible to infection with E. paraensei, rediae of which undergo degeneration, hemocyte-mediated destruction, or both by 7-9 DPE.

Cytokine production by cultured human bronchial epithelial cells infected with a replication-deficient adenoviral gene transfer vector or wild-type adenovirus type 5.

Exposure of animals to adenoviral gene transfer vectors has been associated with respiratory tract inflammation. The pathogenesis of this inflammation is unclear. One hypothesis is that viral vectors directly induce production of inflammatory cytokines by host cells in the airways. We exposed cultured human lung cells to an adenovirus-5--based vector containing the cytomegalovirus promoter and lacZ reporter gene (Ad.CMV.lacZ) and to wild-type adenovirus 5 (wtAd5) and measured subsequent release of cytokines into cell culture supernatants. Inoculation of human bronchial epithelial (HBE) cells with Ad.CMV. lacZ at 10(1) to 10(4) plaque-forming units (pfu)/cell resulted in dose-related expression of lacZ by both X-gal staining and immunohistochemistry but did not increase release of interleukin (IL)-8 or IL-6 at 24, 48, or 96 h after inoculation. In the same cultures, tumor necrosis factor-alpha induced marked increases in release of both IL-8 and IL-6 at 24 and 48 h after stimulation. Similar data were observed in the BEAS-2B HBE cell line. HBE cells incubated with wtAd5 at doses of 10(1) to 10(3) pfu/cell did not release increased amounts of IL-6 or IL-8 up to 48 h after inoculation, though wild-type respiratory syncytial virus (3 pfu/HBE cell) infection resulted in increases in both cytokines. Human alveolar macrophages obtained by bronchoalveolar lavage also showed no increases in cytokine release after incubation with Ad.CMV.lacZ, though relatively little gene transfer occurred in macrophages. These data do not support a role for direct induction of airway epithelial or alveolar macrophage inflammatory cytokines in the pathogenesis of inflammation associated with exposure of airways to adenovirus or to adenoviral gene transfer vectors.

A replication-deficient adenovirus enhances liposome-mediated nucleic acid transfer into a stable cell line expressing T7 RNA polymerase.

Liposome-mediated transfer of nucleic acids into a cell line expressing bacteriophage T7 RNA polymerase was enhanced by addition of a replication-deficient adenovirus (Ad5-259A) to transfection mixtures. Increasing quantities of Ad5-259A resulted in a dose-related (up to 30-fold) enhancement of reporter gene activity expressed in BT7-H cells transfected with plasmid DNA containing the reporter sequence fused to the internal ribosome entry site of encephalomyocarditis virus. Similarly, Ad5-259A enhanced reporter gene expression 7-fold following transfection of DNA containing the reporter sequence under transcriptional control of the Rous sarcoma virus long terminal repeat. Addition of Ad5-295A to transfection mixtures increased the proportion of cells staining positively for reporter gene activity, from 2 to 25% when the reporter was expressed via the T7 polymerase and from 20 to 50% when the reporter was under the control of a eucaryotic promoter. Thus, Ad5-259A enhanced reporter protein activities expressed by cytoplasmic T7-directed transcription and cap-independent initiation of translation, or nuclear transcription and cap-dependent translation. Transfection enhancement was blocked by neutralizing antibody to Ad5, and is most likely related to the endosome-disrupting activities of the virus. Adenovirus enhancement of liposome-mediated transfection provides a useful method for efficient nucleic acid transfer into eucaryotic cells.

Transition mutations in the 23S rRNA of erythromycin-resistant isolates of Mycoplasma pneumoniae.

Erythromycin is the drug of choice for treatment of Mycoplasma pneumoniae infections due to its susceptibility to low levels of this antibiotic. After exposure of susceptible strains to erythromycin in vitro and in vivo, mutants resistant to erythromycin and other macrolides were isolated. Their phenotypes have been characterized, but the genetic basis for resistance has never been determined. We isolated two resistant mutants (M129-ER1 and M129-ER2) by growing M. pneumoniae M129 on agar containing different amounts of erythromycin. In broth dilution tests both strains displayed resistance to high levels of several macrolide-lincosamide-streptogramin B (MLS) antibiotics. In binding studies, ribosomes isolated from the resistant strains exhibited significantly lower affinity for [14C]erythromycin than did ribosomes from the M129 parent strain. Sequencing of DNA amplified from the region of the 2S rRNA gene encoding domain V revealed an A-to-G transition in the central loop at position 2063 of M129-ER1 and a similar A-to-G transition at position 2064 in M129-ER2. Transitions at homologous locations in the 23S rRNA from other organisms have been shown to result in resistance to MLS antibiotics. Thus, MLS-like resistance can occur in M. pneumoniae as the result of point mutations in the 23S rRNA gene which reduce the affinity of these antibiotics for the ribosome. Since they involve only single-base changes, development of resistance to erythromycin in vivo by these mechanisms could be relatively frequent event.

A method for immunoisolation of sporocysts of Schistosoma mansoni in nonsusceptible snails.

A technique is described for immunoisolation of sporocysts of Schistosoma mansoni in nonsusceptible Biomphalaria glabrata by microencapsulation in agarose. Based on histological evidence, all 11 microencapsulated sporocysts implanted into 3 schistosome-resistant 13-16-R1 snails were alive at 72 hr postimplantation, although in a developmentally retarded condition. However, among 146 sporocysts derived from miracidia that had penetrated 5 snails, 96% were dead by 72 hr. These results suggest that hemocyte contact is necessary for rapid sporocyst death in vivo.

The minimal gene complement of Mycoplasma genitalium.

The complete nucleotide sequence (580,070 base pairs) of the Mycoplasma genitalium genome, the smallest known genome of any free-living organism, has been determined by whole-genome random sequencing and assembly. A total of only 470 predicted coding regions were identified that include genes required for DNA replication, transcription and translation, DNA repair, cellular transport, and energy metabolism. Comparison of this genome to that of Haemophilus influenzae suggests that differences in genome content are reflected as profound differences in physiology and metabolic capacity between these two organisms.

A controlled study of adenoviral-vector-mediated gene transfer in the nasal epithelium of patients with cystic fibrosis.

BACKGROUND: Cystic fibrosis is a monogenic disease that deranges multiple systems of ion transport in the airways, culminating in chronic infection and destruction of the lung. The introduction of a normal copy of the cystic fibrosis transmembrane conductance regulator (CFTR) gene into the airway epithelium through gene transfer is an attractive approach to correcting the underlying defects in patients with cystic fibrosis. We tested the feasibility of gene therapy using adenoviral vectors in the nasal epithelium of such patients. METHODS: An adenoviral vector containing the normal CFTR complementary DNA in four logarithmically increasing doses (estimated multiplicity of infection, 1, 10, 100, and 1000), or vehicle alone, was administered in a randomized, blinded fashion to the nasal epithelium of 12 patients with cystic fibrosis. Gene transfer was quantitated by molecular techniques that detected the expression of CFTR messenger RNA and by functional measurements of transepithelial potential differences (PDs) to assess abnormalities of ion transport specific to cystic fibrosis. The safety of this treatment was monitored by nasal lavage and biopsy to assess inflammation and vector replication. RESULTS: The adenoviral vector was detected in nasal-lavage fluid by culture, the polymerase chain reaction (PCR), or both in a dose-dependent fashion for up to eight days after vector administration. There was molecular evidence of gene transfer by reverse-transcriptase PCR assays or in situ hybridization in five of six patients treated at the two highest doses. However, the percentage of epithelial cells transfected by the vector was very low (< 1 percent), and measurement of PD across the epithelium revealed no significant restoration of chloride transport or normalization of sodium transport. At the lower doses of vector, there were no toxic effects. However, at the highest dose there was mucosal inflammation in two of three patients. CONCLUSIONS: In patients with cystic fibrosis, adenoviral-vector-mediated transfer of the CFTR gene did not correct functional defects in nasal epithelium, and local inflammatory responses limited the dose of adenovirus that could be administered to overcome the inefficiency of gene transfer.

Gene therapy for hemophilia B: host immunosuppression prolongs the therapeutic effect of adenovirus-mediated factor IX expression.

Hemophilia B is caused by a deficiency of blood clotting factor IX (FIX). Previous studies have shown that the delivery of a recombinant adenoviral vector expressing canine FIX (cFIX) resulted in a complete correction of hemophilia B in FIX-deficient dogs, but that cFIX expression decreased to only about 1-2% of normal levels 3 weeks after treatment. In the present study, therapeutic levels of cFIX expression capable of producing a partial correction of hemophilia B were maintained for at least 6 months after the coadministration of the cFIX-expressing adenovirus and the immunosuppressive agent cyclosporin A (CsA). These findings support a recent report (Yang et al., 1994) that host T-cell-mediated immunity against virally transduced cells is a major contributing factor to the transient nature of adenovirus-mediated gene expression in immunocompetent animals. Although a second administration of the cFIX-expressing adenovirus 6 months after the first infusion had only a minimal effect on plasma FIX levels in a dog that had been continuously treated with CsA, the prolonged expression of the transgene indicates that immunosuppression may be applicable in attaining long-term treatment of clinically relevant disorders.

Genetic map of the Mycoplasma genitalium chromosome.

At 600 kb, the genome of Mycoplasma genitalium is among the smallest known for cellular organisms capable of independent replication. As such, elucidation of the genetic makeup and chromosome architecture of this organism is of considerable interest. We have located 631 markers on the physical map of M. genitalium. The clones have been mapped by hybridizing 20 overlapping cosmid and lambda clones which encompass the entire M. genitalium chromosome to replica filters containing 856 genomic DNA clones. Three hundred fifty-six of these clones represent sequence tag sites, which were previously characterized by database searches. The remaining markers represent clones with an average size of 2.5 kb derived from Sau3A1 partial digestion of genomic DNA. The hybridization data can be divided into three classes: clones which hybridized to only one cosmid; clones which hybridized to two adjacent and overlapping cosmids; and clones which hybridized to several cosmids, which represent repetitive DNA. This rapid approach for placing clones on the physical map has allowed useful comparisons to be made with other bacterial chromosomes, especially that of the closely related organism M. pneumoniae, and has provided insight to the types of events which may have led to the reduction in size of this genome. Future use of these data is discussed.

Differential effects of ozone on lung epithelial lining fluid volume and protein content.

Urea dilution has been used to estimate the volume of epithelial lining fluid (ELF) in the respiratory tract. However, ELF volume may be overestimated as the result of rapid net diffusion of urea from tissues into the bronchoalveolar lavage (BAL) fluid. This study established a protocol for rat BAL in a manner that minimizes this problem and then used this procedure to examine the edemagenic effects of ozone (O3) exposure on ELF volume and the concentrations of ELF protein and albumin. One passage lavage with variable dwell times up to 30 s showed no difference in recovered urea, protein, and albumin and ELF volume between 0 and 4 s, but a progressive increase of each thereafter. The calculated concentrations of protein and albumin in ELF did not vary significantly with dwell time. By increasing the number of lavage passages from one to three, the amounts of recovered urea, protein, and albumin and estimated ELF volume were increased with each passage. Again, the calculated concentrations of protein and albumin in ELF did not vary appreciably. When a single lavage passage and no added dwell time were used, it was observed that exposure of rats to 2 but not 0.5 and 1 ppm O3 increased urea, protein, and albumin in the BAL immediately after 6 h exposure. In addition, at 18 h postexposure to 1 ppm O3, ELF volume increased only 21%, but protein and albumin concentrations in ELF were 2.3- and 4.5-fold of control values, respectively. A higher O3 concentration (2 ppm) moderately increased ELF volume (+83%) and exerted even greater effects on concentrations of ELF protein (7.8-fold) and albumin (19-fold) while lower O3 dosage (0.5 ppm) had no significant effect. SDS-PAGE analysis showed that small serum proteins including albumin were greatly enriched in lung BAL fluid of 1 ppm O3-exposed rats. These results demonstrate that movement of water and protein into the airspaces after O3 exposure is not strictly coupled, and that protein recovery by BAL should cautiously be used to indicate airspace edema as a result of O3 injury.

Construction of an ordered genomic library of Mycoplasma genitalium.

As a first step towards sequencing the chromosome of the suspected human pathogen Mycoplasma genitalium, we attempted to clone its entire genome in a set of ordered cosmids. Cosmid libraries were established by partial digestion of M. genitalium genomic DNA with Sau3AI or EcoRI. A chromosome-walking strategy was used to identify 20 overlapping cosmid clones which contained over 99% of the genome. The final 5.1 kb could not be cloned in cosmids, and was eventually obtained from a genomic library established in a lambda vector. Correspondence of cloned and genomic EcoRI fragments indicated no detectable major deletions or rearrangements in the library. The library was oriented on established XhoI and SmaI physical maps of the chromosome with restriction sites present at the expected locations in the library. The genome contained 74 EcoRI fragments which added up to a total genome size of 578 kb. These were arranged in a partial EcoRI physical map, and those containing the MgPa major attachment protein-encoding operon and its repeat sequences were identified. The existence of this ordered genomic library, which accurately and completely encompasses the entire M. genitalium genome, should serve as a valuable tool for many future studies of this organism and facilitate our long-term goal of sequencing its genome.

Protection of immunized and previously infected chimpanzees challenged with Mycoplasma pneumoniae.

Following immunization, peak geometric mean serum metabolism inhibition antibody (MIT) titres were 1:13 and 1:16 for groups of three chimpanzees each that received either the formalin-inactivated OSU-1A or experimental acellular extract vaccine, respectively. Following challenge, the mean titres for chimpanzees given the acellular vaccine peaked at 1:256 in 4 weeks and was 1:48 at 10 weeks. Chimpanzees given the OSU-1A vaccine peaked at 1:80 in 4 weeks and remained at 1:80 at 10 weeks. There was no direct correlation between the serum MIT response and the severity of disease or colonization, and thus the MIT response was not a reliable measurement of protection. The two non-immunized chimpanzees showed significant signs of disease, including cough, pharyngitis, rhinitis, fever and abnormal X-ray findings, for about 5 weeks. The chimpanzees immunized with either vaccine were less colonized and showed far less disease than non-immunized controls. Protection afforded the chimpanzees was similar to that of vaccinees in the human clinical trial given the same OSU-1A vaccine (Wenzel et al., 1977). The two previously infected chimpanzees were most protected against colonization and disease on challenge.

Efficient transfection of primary cells in a canine hemophilia B model using adenovirus-polylysine-DNA complexes.

We have used molecular conjugates containing combinations of DNA, adenovirus, polylysine, and transferrin to transfect primary cells derived from canines with hemophilia B (factor IX deficiency), as well as a canine epithelial cell line. Transfection of canine hemophilia B fibroblasts with molecular conjugates resulted in efficient transfection and expression of luciferase DNA-adenovirus-polylysine (AdpL) conjugates or luciferase DNA-adenovirus-polylysine-transferrin (hTfpL/AdpL) conjugates. No expression in canine hemophilia B fibroblasts was evident after exposure to DNA alone, or DNA conjugated with polylysine and transferrin. Transfection efficiencies of 50% or more could be demonstrated in cells transfected with a beta-galactosidase reporter gene as part of an hTfpL/AdpL molecular conjugate. Transfection with canine factor IX AdpL conjugates or canine factor IX hTfpL/AdpL conjugates resulted in factor IX expression for more than 2 weeks in vitro in hemophilia B canine fibroblasts. Maximum levels of expression of over 700 ng of canine factor IX/10(6) cells/24 hr were demonstrated in fibroblasts after transfection with canine factor IX hTfpL/AdpL conjugates. Similar conjugates were used to transfect hemophilia B canine bone marrow stromal cells and Madin-Darby canine kidney cells that also expressed canine factor IX. The use of molecular conjugates to transfect primary cells may be feasible as a means of in vitro or in vivo gene therapy for hemophilia B, and can be tested in the canine hemophilia B model.

Immunoblot analyses of chimpanzee sera after infection and after immunization and challenge with Mycoplasma pneumoniae.

Consecutive weekly or biweekly serum specimens obtained during a 3- or 4-month study from 16 chimpanzees were examined by immunoblot analyses to identify the immunogenic components of Mycoplasma pneumoniae. Six experimentally infected chimpanzees showed significant signs of overt disease, including cough, pharyngitis, rhinitis, fever, and loss of appetite. The sera of these infected chimpanzees recognized from 17 to 20 protein bands. Two control chimpanzees that were not inoculated were included in the study. Three chimpanzees immunized with a formalin-inactivated OSU-1A vaccine and three chimpanzees immunized with an experimental acellular vaccine showed minimal signs of disease on challenge. After challenge, the serum immunoblot responses of the immunized chimpanzees were similar to those of the infected chimpanzees. Before challenge, the sera of two previously infected chimpanzees recognized protein bands of 169 (which comigrated with the P1 adhesin), 148, 130, 117, 86, 61, 44, 35, 30, and 29 kDa. After challenge, the previously infected chimpanzees showed the most intense serum immunoblot responses and were most protected against colonization and disease. The sera from each of the 16 chimpanzees examined recognized a large number of immunogenic components, and the serum immunoblot responses were virtually identical to those of patients. Sera from each chimpanzee and patient recognized 169-, 148-, 130-, 117-, 86-, 44-, and 35-kDa bands and many of them recognized 67-, 63-, 61-, 56-, 32-, 30-, and 29-kDa protein bands.

Strategy for achieving selective killing of carcinomas.

Carcinomas are malignancies derived from epithelial cells that frequently respond poorly to conventional chemotherapy. Selective expression or transduction of toxin genes to carcinomas, i.e. molecular chemotherapy, may offer important advantages over conventional chemotherapy. As one approach to developing a means of selectively expressing toxin genes, the transcriptional regulatory sequences of a gene expressed in multiple carcinomas were used to direct expression of the herpes simplex virus thymidine kinase (HSVtk) coding sequences. The secretory leukoprotease inhibitor (SLPI) gene was found to be expressed in lung, breast, oropharyngeal, bladder, endometrial, ovarian and colorectal carcinomas. The tissue-specific transcriptional regulatory sequences were isolated and used to construct a chimeric gene in which the SLPI sequences directed HSVtk expression. SLPI-expressing carcinomas were reduced in number by transduction of the SLPI-directed toxin plasmid plus ganciclovir, but the same treatment had no effect on a cell line that did not express SLPI. These results suggest that SLPI-directed therapeutic genes could be used for directing toxicity to carcinoma tissues, especially if combined with other targeting strategies.