Estimating Need, Demand and Supply in Primary Health Care Services: A Local Application in Argentina.

INTRODUCTION To provide equal access, health care provision should be distributed across geodemographic space based on need. In Argentina, the social security, publicly funded health care and private health care subsectors are responsible for delivering health services. In the public subsector, which is responsible for providing primary and secondary care mainly to the uninsured population, supply of services is not always associated with need. The lack of coordination between levels and subsectors makes it difficult to transform need into demand. OBJECTIVE Design a methodology to systematically estimate need, demand and supply of primary health care services based on secondary data sources in order to assess potential mismatches in any geographical area. METHODS An ecological analysis was conducted based on outpatient visits in primary care in Bahía Blanca, Buenos Aires Province, Argentina. A mathematical approach was proposed to systematize data collection by census tract regarding estimated need (number of outpatient visits needed, by specialty, according to age- and sex-specific care protocols and the area's demographics), demand (actual outpatient visits by specialty in each primary health care center), and supply (visit capacity or available appointment slots, taking into account number of personnel hours worked, by specialty). RESULTS Demand for outpatient visits exceeded need (299,731) by 24% while available visit capacity (993,903) could have covered more than twice the number demanded (370,881). Analysis of the three variables grouped by area found that supply correlated more closely with demand (ρ = 0.90) than with need (ρ = 0.68), while spatial analysis showed that supply distribution responded to need. Areas with greater need had a health facility relatively close by, although supply was often located in areas of lower need, and some areas struggle with relatively high need and insufficient supply. CONCLUSIONS Results suggest the need for some reconfiguration of primary health care in the study area. The proposed mechanism for estimating relationship among supply, demand and need is a useful tool to support decision-making. KEYWORDS Health services needs and demand, access to health care; health care accessibility, health care quality, access, and evaluation, health care inequalities, primary health care, Argentina.

[Total health expenditure efficiency: non-parametric analysis in a large sample of countries]. / Eficiencia del Gasto Total en Salud: Análisis no paramétrico en una muestra amplia de países.

OBJECTIVE: To measure the efficiency of 190 countries in producing health results and the factors that determine such efficiency. METHODOLOGY: A data envelopment analysis was conducted on worldwide data from the year 2009 in order to estimate the efficient frontier, based on total health expenditure per capita, as well on infant mortality rate and life expectancy at birth. At the same time, an analysis of the determinants of expenditure efficiency was performed through Tobit models. RESULTS: African nations have lower technical and allocative efficiency, but higher scale efficiency. The quality of institutions has a statistically significant impact on the levels of technical and allocative efficiency and on the levels of scale efficiency. The percentage of health expenditure financed by private insurers has an impact on technical and allocative efficiency, while urbanization rates affect the scale efficiency. DISCUSSION: the fact that more than 70 % of countries show decreasing returns suggest that, once certain minimal standards of life quality are achieved, the marginal effect of each additional dollar assigned to health is not substantial. Conversely, in poor countries, where the expenditure in health presents increasing returns, the health performance could be substantially better by marginally raising the expenditure. On the other hand, financing structures of health expenditures may influence technical-allocative efficiency, while urbanization levels may impact scale efficiency (source: MeSH, NLM).

OBJETIVO: Mostrar la utilidad de una herramienta estadística no paramétrica para medir la eficiencia de 190 países en la producción de status de salud, así como conocer los determinantes de dicha eficiencia. MÉTODOS: Con datos de 2009, y utilizando la técnica de Envolvente de Datos, se estima la frontera de eficiencia, utilizando como insumo al gasto total en salud per cápita y como productos la tasa de mortalidad infantil y la esperanza de vida al nacer. Se realiza un análisis de los determinantes de la eficiencia del gasto mediante el uso de modelos Tobit. RESULTADOS: Las naciones del continente africano presentan menor eficiencia técnico-asignativa, aunque mayor eficiencia de escala. La calidad de las instituciones muestra un impacto estadísticamente significativo sobre los niveles de eficiencia técnico-asignativa y de escala. El porcentaje de financiamiento del gasto por parte de aseguradoras privadas incide sobre la eficiencia técnico-asignativa mientras que el porcentaje de urbanización lo hace sobre la eficiencia de escala. DISCUSIÓN: El hecho de que más del 70 % de los países presente rendimientos decrecientes del gasto en salud sugeriría que, una vez alcanzados ciertos estándares mínimos de calidad de vida, el efecto marginal de cada dólar adicional destinado a salud no es sustancial. En países pobres donde el gasto en salud presenta rendimientos crecientes, el desempeño sanitario podría mejorar significativamente con incrementos marginales del gasto. Las estructuras de financiamiento del gasto en salud podrían estar influyendo sobre la eficiencia técnico-asignativa y el grado de urbanización podría hacerlo sobre la eficiencia de escala.

Differential induction of c-Jun NH2-terminal kinase and c-Abl kinase in DNA mismatch repair-proficient and -deficient cells exposed to cisplatin.

The c-Abl nonreceptor tyrosine kinase and the c-Jun NH2-terminal kinase (JNK/stress-activated protein kinase) are activated during the injury response to the DNA-damaging agent cisplatin. Loss of DNA mismatch repair activity results in resistance to cisplatin in human cancer cells, suggesting that the mismatch repair proteins function as a detector for cisplatin DNA adducts. To identify signaling pathways activated by this detector, we investigated the effect of the loss of DNA mismatch repair function on the ability of cisplatin to activate the JNK and c-Abl kinases. The results demonstrate that cisplatin activates JNK kinase 3.8 +/- 0.2-fold more efficiently in DNA mismatch repair-proficient than repair-deficient cells, and that activation of c-Abl is completely absent in the DNA mismatch repair-deficient cells. Furthermore, the results show that cisplatin-induced activation of JNK occurs through a stress-activated protein kinase/extracellular signal-regulated kinase kinase 1-independent mechanism. We conclude that activation of JNK and c-Abl by cisplatin is in part dependent upon the integrity of DNA mismatch repair function, suggesting that these kinases are part of the signal transduction pathway activated when mismatch repair proteins recognize cisplatin adducts in DNA.

In vitro and in vivo resistance to cisplatin in cells that have lost DNA mismatch repair.

In vitro studies have shown that loss of DNA mismatch repair due to lack of either hMSH2 or hMLH1 activity results in low-level resistance to cisplatin but not to oxaliplatin, an analogue that produces a different type of DNA adduct. No information is currently available on whether this low-level resistance is sufficient to result in enrichment of mismatch repair-deficient cells during drug exposure in vitro or to account for clinical failure of treatment in vivo. Mixed populations of cells containing a minority of DNA mismatch repair-deficient cells constitutively expressing green fluorescence protein were exposed repeatedly in vitro to cisplatin and oxaliplatin. Treatment with cisplatin resulted in a gradual enrichment for DNA mismatch repair-deficient cells, whereas treatment with oxaliplatin did not. MSH2-/- and MSH2+/+ embryonic stem cells were established as xenografts in athymic nude mice. Animals were treated 48 h after tumor implantation with a single LD10 dose of either cisplatin or oxaliplatin. MSH2-/- tumors were significantly less responsive to cisplatin than MSH2+/+ tumors, whereas there was no difference in sensitivity to oxaliplatin. These results demonstrate that the degree of cisplatin resistance conferred by loss of DNA mismatch repair is sufficient to produce both enrichment of mismatch repair-deficient cells during treatment in vitro and a large difference in clinical responsiveness in vivo. The results identify loss of DNA mismatch repair as a mechanism of resistance to cisplatin but not oxaliplatin.

The role of DNA mismatch repair in platinum drug resistance.

Loss of DNA mismatch repair occurs in many types of tumors. The effect of the loss of DNA mismatch repair activity on sensitivity to cisplatin and a panel of analogues was tested using two pairs of cell lines proficient or deficient in this function. HCT116+ch2, a human colon cancer cell line deficient in hMLH1, was 2.1-fold resistant to cisplatin and 1.3-fold resistant to carboplatin when compared to a subline complemented with chromosome 3 expressing a wild-type copy of hMLH1. Likewise, the human endometrial cancer cell line HEC59, which is deficient in hMSH2, was 1.8-fold resistant to cisplatin and 1.5-fold resistant to carboplatin when compared to a subline complemented with chromosome 2 with a wild-type hMSH2. In contrast to cisplatin and carboplatin, which form the same types of adducts in DNA, there was no difference in sensitivity between the DNA mismatch repair-proficient and -deficient cell lines for oxaliplatin, tetraplatin, transplatin, JM335, or JM216. The formation of protein-DNA complexes that contained hMSH2 and hMLH1 was documented by mobility shift assay when nuclear extracts were incubated with DNA platinated with cisplatin but not with oxaliplatin. These results demonstrate a correlation between failure of the DNA mismatch repair proteins to recognize the platinum adduct and low-level resistance, suggesting a role for the DNA mismatch repair system in generating signals that contribute to the generation of apoptotic activity. They also identify the use of drugs whose adducts are not recognized as a strategy for circumventing resistance due to loss of DNA mismatch repair.

Decreased cisplatin damage-dependent DNA synthesis in cellular extracts of mismatch repair deficient cells.

The proficiency of both nucleotide excision repair (NER) and DNA mismatch repair (MMR) influences cellular sensitivity to cisplatin (cis-diamminedichloroplatinum). To gain further insight into how MMR may influence platinum drug sensitivity, the effect of loss of MMR on repair synthesis was measured in vitro by a commonly used method that relies on whole-cell extracts to drive [alpha-32P]dATP incorporation into cisplatin-damaged plasmid DNA. Extracts evaluated include those from cells with or without functional hMLH1 (HCT116+ch2 versus HCT116+ch3, respectively) and hMSH2 (HEC59 versus HEC59+ch2, respectively). Loss of MMR in the HCT116 system was associated with a 2.8-fold reduction in cisplatin damage-specific DNA synthesis, whereas it was associated with a 3.0-fold reduction in the HEC59 system, suggesting that a decrease in the ability to repair cisplatin-damaged DNA accompanies loss of MMR. An in vitro DNA excision assay that utilized a substrate containing a site-specific cisplatin adduct was performed. Using this highly NER-specific assay, no significant difference was apparent between the extracts derived from NER-proficient versus -deficient cells. These and other data lead us to suggest that the increase in apparent repair synthesis in platinum-damaged plasmids by extracts from MMR-proficient versus -deficient cellular extracts may reflect a distinct and possibly adverse DNA synthetic process rather than productive NER.

Loss of DNA mismatch repair due to knockout of MSH2 or PMS2 results in resistance to cisplatin and carboplatin.

Loss of DNA mismatch repair is a common finding in hereditary nonpolyposis colon cancer as well as in many types of sporadic human tumors. The effect of loss of DNA mismatch repair activity on sensitivity to cisplatin and carboplatin was tested using MSH2 and PMS2 knockout cell lines. The knockout dMsh2 embryonic stem cell line was 2.1-fold more resistant to cisplatin and 1.7-fold more resistant to carboplatin when compared to the isogenic wild-type wt-2 cell line. Likewise, the PMS2(-/-) mouse fibroblasts were 1.9-fold more resistant to cisplatin and 1.5-fold more resistant to carboplatin when compared to the isogenic PMS2(+/+) fibroblasts. These findings demonstrate that loss of mismatch repair due to knockout of either MSH2 or PMS2 results in low-level resistance to cisplatin and carboplatin, drugs that form the same types of adducts in DNA. These data validate results previously obtained using non-isogenic mismatch repair-proficient and -deficient cell lines, and indicate that simple recognition of the cisplatin adduct by the MSH2/MSH6 heterodimer is not sufficient for full detector function, but that PMS2 is also required for the pro-apoptotic signal to be generated from this detector.

Eficiencia del Gasto Total en Salud: Análisis no paramétrico en una muestra amplia de países / Total health expenditure efficiency: non-parametric analysis in a large sample of countries

RESUMEN Objetivo Mostrar la utilidad de una herramienta estadística no paramétrica para medir la eficiencia de 190 países en la producción de status de salud, así como conocer los determinantes de dicha eficiencia. Métodos Con datos de 2009, y utilizando la técnica de Envolvente de Datos, se estima la frontera de eficiencia, utilizando como insumo al gasto total en salud per cápita y como productos la tasa de mortalidad infantil y la esperanza de vida al nacer. Se realiza un análisis de los determinantes de la eficiencia del gasto mediante el uso de modelos Tobit. Resultados Las naciones del continente africano presentan menor eficiencia técnico-asignativa, aunque mayor eficiencia de escala. La calidad de las instituciones muestra un impacto estadísticamente significativo sobre los niveles de eficiencia técnico-asignativa y de escala. El porcentaje de financiamiento del gasto por parte de aseguradoras privadas incide sobre la eficiencia técnico-asignativa mientras que el porcentaje de urbanización lo hace sobre la eficiencia de escala. Discusión El hecho de que más del 70 % de los países presente rendimientos decrecientes del gasto en salud sugeriría que, una vez alcanzados ciertos estándares mínimos de calidad de vida, el efecto marginal de cada dólar adicional destinado a salud no es sustancial. En países pobres donde el gasto en salud presenta rendimientos crecientes, el desempeño sanitario podría mejorar significativamente con incrementos marginales del gasto. Las estructuras de financiamiento del gasto en salud podrían estar influyendo sobre la eficiencia técnico-asignativa y el grado de urbanización podría hacerlo sobre la eficiencia de escala.(AU)

Objective To measure the efficiency of 190 countries in producing health results and the factors that determine such efficiency. Methodology A data envelopment analysis was conducted on worldwide data from the year 2009 in order to estimate the efficient frontier, based on total health expenditure per capita, as well on infant mortality rate and life expectancy at birth. At the same time, an analysis of the determinants of expenditure efficiency was performed through Tobit models. Results African nations have lower technical and allocative efficiency, but higher scale efficiency. The quality of institutions has a statistically significant impact on the levels of technical and allocative efficiency and on the levels of scale efficiency. The percentage of health expenditure financed by private insurers has an impact on technical and allocative efficiency, while urbanization rates affect the scale efficiency. Discussion the fact that more than 70 % of countries show decreasing returns suggest that, once certain minimal standards of life quality are achieved, the marginal effect of each additional dollar assigned to health is not substantial. Conversely, in poor countries, where the expenditure in health presents increasing returns, the health performance could be substantially better by marginally raising the expenditure. On the other hand, financing structures of health expenditures may influence technical-allocative efficiency, while urbanization levels may impact scale efficiency (source: MeSH, NLM).(AU)

"Local Food - Nutraceuticals": an example of a multidisciplinary research project on local knowledge.

Currently, dietary patterns change rapidly all over the world. Most notably, there is a fast increase in the convenience food market. Here we discuss the overall theoretical framework and strategy of an EU-funded project on local food, a common resource in many parts of the Mediterranean. Such food is often only available seasonally and is consumed either fresh (e.g. spring salads and vegetables, fruits in autumn) or in a conserved form (dried, fermented, pickled). There is an urgent need to document and analyse such local resources, which are today at the brink of disappearance. In this project, selected species were studied using a multidisciplinary approach, including strategies and methods from pharmacology, nutritional sciences and anthropology (i.e. ethnopharmacological or ethnonutritional ones). For example, all extracts were profiled using HPLC-MS, by determining their polyphenol content and using a variety of in vitro anti-oxidant assays (incl. guaiacol oxidation, xanthine oxidase inhibition, HOCl scavenging, eNOS activity). Such research also points to ways for ascertaining the intergenerational transmission of the knowledge and for sustainable development and management. Examples from field studies in southern Italy and from pharmacological studies using a variety of targets are used to illustrate the potential of such neglected resources. The wider implications of such an approach, for example, for the study of similar traditions in Central and Eastern Europe are also discussed.

Heat changes in lipid membranes under sudden osmotic stress.

Closed lipid vesicles act as osmometers increasing or decreasing their volume under the influence of osmotic gradients. The enthalpy changes accompanying membrane compression or expansion have not been measured yet, and first results obtained with high-sensitivity titration calorimetry are reported here. Phospholipid vesicles suspended in and in equilibrium with an electrolyte or nonelectrolyte with a defined initial concentration of c(i), were injected into a solution with a final concentration of c(f), and the heat changes were monitored with a titration microcalorimeter. Osmotic compression (delta c = c(f) - c(i) > 0) produced an exothermic heat change with deltaH approximately -500 +/- 100 cal/mol and osmotic expansion (delta c < 0) an endothermic heat change with deltaH approximately 1000 +/- 200 cal/mol; both results normalized to a concentration gradient of delta c = 1 M NaCl. The heats of compression and expansion varied linearly with the lipid content and the size of the osmotic gradient but were independent of the vesicle size. The cubic thermal expansion coefficient alpha(v) which equals (1/V)(deltaV/deltaT)p could be derived and was found to be 1.25 x 10(-3) and 2.5 x 10(-3) K(-1) for the compressed and expanded bilayer vesicles, respectively. The entropy changes associated with compression and expansion could be estimated. Compression of the membrane led to a negative entropy change and increased the hydrocarbon chain order. Expansion of the membrane was accompanied by a positive entropy change which can be explained, in part, by more disordered hydrocarbon chains. Vesicle expansion and compression thus appear to be asymmetric as far as the thermodynamic driving force is concerned.

Calorimetric detection of influenza virus induced membrane fusion.

Membrane fusion induced by the hemagglutinin glycoprotein of influenza virus has been extensively characterized, but the mechanism whereby the protein achieves the merger of the viral and target membrane lipids remains enigmatic. Various lipid intermediate structures have been proposed, and the energies required for their formation predicted. Here, we have analyzed the enthalpies of fusion of influenza with liposomes by titration calorimetry. If a small sample of virus in a weak neutral pH buffer was added to an excess of liposomes at low pH, a two-component reaction was seen, composed of an exothermic reaction and a slower endothermic reaction. The exothermic reaction was the result of acid-base reactions between the neutral pH virus sample and low pH buffer and low-pH-induced changes in the virus. The endothermic reaction was not observed in the absence of liposomes and much reduced if acid-inactivated virus, which had lost its fusion but not its binding activity, was added to liposomes. The endothermic reaction was more temperature dependent than the exothermic reaction; its pH dependence corresponded with that of fusion and its enthalpy was higher if fusion was more extensive. These data indicate that most of the endothermic reaction was due to membrane fusion. The experimentally determined enthalpy of fusion, 0.6-0.7 kcal per mol of viral phospholipids, is much higher than expected on the basis of current theories about the formation of lipid intermediates during membrane fusion.

Electrostatic and nonpolar peptide-membrane interactions. Lipid binding and functional properties of somatostatin analogues of charge z = +1 to z = +3.

The interaction of four structurally related somatostatin analogues (effective electric charge +0.4 < or = < or = +3) with lipid membranes was studied with titration calorimetry and was compared with the functional activity of the peptides. Surface activity measurements provided average cross-sections of 70 or 135 A2, indicating that the cyclic molecules orient at the air-water interface with their ring system either parallel (z = +3) or perpendicular (z = +1) to the surface or switching between the two orientations according to the surface density (z = +2). The nonspecific binding of the peptides to sonified lipid vesicles was enthalpy-driven with a delta H of -4 to -7.5 kcal/mol. A consistent quantitative analysis of the binding isotherms was achieved by combining electrostatic attractions, calculated via the Gouy-Chapman theory, with a nonspecific surface partition equilibrium for the nonpolar interactions. The electrostatic attraction of the cationic peptides varied strongly according to the peptide charge. Due to the flat ring structure of the cyclic peptides, their true physical charge was sensed at the membrane surface, and no "charge screening" was observed. Peptide binding to the negative charged membrane was accompanied by a proton-uptake of the N-terminal amino group of 0.23-0.38 H+/peptide. Deviations from the theoretical prediction of 0.39 H+/peptide can be explained by a preferential binding of the nonprotonated species. The nonpolar interactions, as described by the surface partition coefficients of the four peptides, fell into a narrow range of K congruent to 50-230 M-1 whereas the apparent overall binding constants were between 200 and 5000 M-1.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression of the DNA mismatch repair proteins hMLH1 and hPMS2 in normal human tissues.

hMLH1 and hPMS2 are part of the DNA mismatch repair complex. Mutations in these genes have been linked to hereditary non-polyposis colon cancer; they also occur in a variety of sporadic cancers. Western blot analysis and immunohistochemistry demonstrated that hMLH1 and hPMS2 are widely expressed nuclear proteins with a distribution pattern very similar to that previously described for hMSH2. These observations showing similar localization of hMLH1 and hPMS2 with hMSH2 are consistent with the biochemical function of these proteins in DNA mismatch repair.

Induction of JNK and c-Abl signalling by cisplatin and oxaliplatin in mismatch repair-proficient and -deficient cells.

Loss of DNA mismatch repair has been observed in a variety of human cancers. Recent studies have shown that loss of DNA mismatch repair results in resistance to cisplatin but not oxaliplatin, suggesting that the mismatch repair proteins serve as a detector for cisplatin but not oxaliplatin adducts. To identify the signal transduction pathways with which the detector communicates, we investigated the effect of loss of DNA mismatch repair on activation of known damage-responsive pathways, and recently reported that cisplatin differentially activates c-Jun NH2-terminal kinase (JNK) and c-Abl in repair-proficient vs.-deficient cells. In the current study, we directly compared differential activation of these pathways by cisplatin vs. oxaliplatin. The results confirm that cisplatin activates JNK kinase 5.7 +/- 1.5 (s.d.)-fold more efficiently in DNA mismatch repair-proficient than repair-deficient cells, and that the c-Abl response to cisplatin is completely absent in DNA mismatch repair-deficient cells. In contrast, there was no detectable activation of the JNK or c-Abl kinases in DNA mismatch repair-proficient or -deficient cells exposed to oxaliplatin. The present study demonstrates that, despite the similarity of the adducts produced by cisplatin and oxaliplatin, they appear to be recognized by different detectors. The DNA mismatch repair system plays an important part in the recognition of cisplatin adducts, and activation of both the JNK and c-Abl kinases in response to cisplatin damage is dependent on the detector function of the DNA mismatch repair proteins. In contrast, this detector does not respond to oxaliplatin adducts.

The effect of different chemotherapeutic agents on the enrichment of DNA mismatch repair-deficient tumour cells.

Loss of DNA mismatch repair is a common finding in hereditary non-polyposis colon cancer as well as in many types of sporadic human tumours. We compared the effect of loss of DNA mismatch repair on drug sensitivity as measured by a clonogenic assay with its effect on the ability of the same drug to enrich for mismatch repair-deficient cells in a proliferating tumour cell population. Mixed populations containing 50% DNA mismatch repair-deficient cells constitutively expressing green fluorescent protein and 50% mismatch repair-proficient cells were exposed to different chemotherapeutic agents. 6-Thioguanine, to which DNA mismatch repair-deficient cells are known to be resistant, was included as a control. The results in the cytotoxicity assays and in the enrichment experiments were concordant. Treatment with either carboplatin, cisplatin, doxorubicin, etoposide or 6-thioguanine resulted in enrichment for mismatch repair-deficient cells, and clonogenic assays demonstrated resistance to these agents, which varied from 1.3- to 4.8-fold. Treatment with melphalan, paclitaxel, perfosfamide or tamoxifen failed to enrich for mismatch repair-deficient cells, and no change in sensitivity to these agents was detected in the clonogenic assays. These results identify the topoisomerase II inhibitors etoposide and doxorubicin as additional agents for which loss of DNA mismatch repair causes drug resistance. The concordance of the results from the two assay systems validates the enrichment assay as a rapid and reliable method for screening for the effect of loss of DNA mismatch repair on sensitivity to additional drugs.

Enrichment for DNA mismatch repair-deficient cells during treatment with cisplatin.

In addition to playing a role in tumorigenesis, loss of DNA mismatch repair results in low-level intrinsic resistance to cisplatin and carboplatin. We used a mismatch repair-deficient (clone B) and -proficient (clone B/rev) pair of Chinese hamster ovary sublines to determine the ability of cisplatin to enrich for repair-deficient cells during growth in vitro and in vivo. Clone B cells were 1.8-fold resistant to cisplatin as measured by a clonogenic assay. These cells were molecularly engineered to express constitutively the green fluorescent protein, and changes in the fraction of these repair-deficient cells were monitored by flow cytometric analysis. A single 1-hr exposure to cisplatin at an IC50 concentration enriched populations initially containing either 5 or 10% clone B cells by 81 and 75%, respectively, when measured at 5 days. Enrichment increased as a function of drug concentration to 158 and 169%, respectively, following an IC90 exposure. When grown as a xenograft, a single LD10 dose of cisplatin enriched the tumors by 48% from 4.6 to 6.8% repair-deficient cells (p = 0.04). To determine whether similar enrichment occurs during the treatment of human ovarian cancer patients, paired tumor samples were obtained from 38 patients before and after treatment with a minimum of 3 cycles of platinum drug-based primary chemotherapy and analyzed immunohistochemically for changes in the fraction of tumor cells expressing hMHL1. Following treatment there was a reduction in hMLH1 staining in 66% of the cases (p = 0.0005). Our results demonstrate that, despite the fact that loss of mismatch repair yields only modest levels of cisplatin resistance, even a single exposure to cisplatin produces quite a marked enrichment for repair-deficient cells in vitro and in vivo. Our results are consistent with the concept that treatment with cisplatin or carboplatin selects for preexisting mismatch repair-deficient cells, and that this contributes to the frequent development of clinical resistance.

Mismatch repair proficiency and in vitro response to 5-fluorouracil.

BACKGROUND & AIMS: The DNA mismatch repair (MMR) system recognizes certain DNA adducts caused by alkylation damage in addition to its role in recognizing and directing repair of interstrand nucleotide mismatches and slippage mistakes at microsatellite sequences. Because defects in the MMR system can confer tolerance to acquired DNA damage and, by inference, the toxic effects of certain chemotherapeutic agents, we investigated the effect of 5-fluorouracil (5-FU) on colon cancer cell lines. METHODS: We determined growth selection by cell enrichment assay and cloning efficiency after treatment with 5 micromol/L 5-FU, assayed nucleic 3H-5-FU incorporation, and analyzed the cell cycle by flow cytometry. RESULTS: 5-FU treatment provided a growth advantage for MMR-deficient cell lines, indicating a relative degree of tolerance to 5-FU by the MMR-deficient cell lines. Enhanced survival was statistically significant after 5 days of growth, and a 28-fold reduction in survival was noted in the MMR-proficient cells by clonagenic assays after 10 days of growth. Differences in nucleotide uptake of 5-FU did not account for the observed growth differences, and specific cell cycle checkpoint arrest was not detected. CONCLUSIONS: Intact DNA MMR seems to recognize 5-FU incorporated into DNA but may do so in a different manner than other types of alkylation damage. Defective DNA MMR might be one mechanism for tumor resistance to 5-FU.