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Biomedical subjects

G P Raaphorst

Publications and source records attributed to G P Raaphorst.

At least 37 records · Page 2Linked to original sources
PubMed1999

p53 status, cellular recovery and cell cycle arrest as prognosticators of in vitro radiosensitivity in human pancreatic adenocarcinoma cell lines.

PURPOSE: To investigate the factors contributing to the in vitro radiosensitivity of four human pancreatic adenocarcinoma cell lines differing in p53 status, and the basis for the lack of post-irradiation G1 arrest in the two cell lines that have retained a wild-type p53 allele. MATERIALS AND METHODS: Cells were X-irradiated and the parameters related to radiosensitivity, as well as the modulation of gene products linked to regulation of cell cycle transit (p53, p21/WAF1/CIP1, pRb) or DNA replication and repair (DNA topoisomerase I and II), were determined. RESULTS: Both cell lines expressing either mutant (mt) R248W or R273H p53 proteins were more radioresistant. All the cell lines arrested in G2. None of the cell lines arrested in G1 and this was linked to the inability to upregulate p21/WAF1/CIP1. There were no correlations between p53 status and the magnitude or time of maximum G2 arrest. However, there was a negative correlation between a protracted arrest in G2 and the ability to recover from potentially lethal damage (PLD). CONCLUSIONS: Variation in radiosensitivity is related to p53 status, but the survival advantage conferred by having mutant p53 status is not readily explained neither by recovery from PLD nor by cell cycle arrest kinetics. There is no p53-independent pathway for the recruitment of p21 in these cell lines following irradiation.

Adenocarcinoma↗
PubMed1999

Hypersensitivity to low single doses and split-dose recovery: two manifestations of induced resistance that might be related.

PURPOSE: To study retrospectively the relationship between intrinsic radiosensitivity (SF2), and both the low-dose inducible response (alpha(s)/alpha(r)) and the amount of split-dose recovery (betaRR). MATERIALS AND METHODS: A total of 53 sets of experimental data obtained with 44 human cell lines were collected from the literature and the above relationships were studied. RESULTS: Analysis showed a statistically significant correlation between alpha(s)/alpha(r) and SF2 (p = 0.0023, 10 sets of data), and a statistically significant inverse correlation between betaRR and SF2 (p = 0.0005, 36 sets of data, AT excluded). Furthermore, the analysis of the relationship between the challenge dose SF2 (after a clinical-sized priming dose) and that of the single-dose SF2 (27 sets of data, AT excluded) showed a statistically significant correlation (p<0.0001), which deviates from, and becomes higher than, the one-to-one relationship for single-dose SF2<0.30, suggesting that the final response to fractionated irradiation in radiosensitive cells might not be predictable on the basis of simple reconstitution of survival from the single-dose treatment. CONCLUSION: The comparison between the two relationships: SF2/(alpha(s)/alpha(r)) and SF2/betaRR, suggests some parallelism indicating that these two phenomena may be inversely correlated and could be attributed to induced resistance mechanisms that might be triggered differently in sensitive and resistant cell lines.

Cell Line↗
PubMed1999

Adaptive response and its variation in human normal and tumour cells.

PURPOSE: Adaptive responses in mammalian cells appear to be highly variable. Six human cell lines were used to evaluate whether adaptive response could be related to radiation sensitivity or tumour versus normal cell lines. MATERIALS AND METHODS: Six human cell lines (two fibroblasts, two melanoma, two breast carcinoma) were cultured under identical conditions to plateau phase. The cells were given inducing doses of 0, 0.2, 0.5, 1.0, 2.0 and 4.0 Gy followed after incubation at 37 degrees C by challenge doses to assess radiation response. RESULTS: An adaptive response (AR) was observed in some but not all the cell lines. The AR was observed at 2 h and persisted for up to 24 h. It was observed for doses as low as 0.2 and 0.5 Gy and declined with higher doses. In the resistant melanoma cell line there was no adaptive response; instead it showed sensitization for all inducing doses. CONCLUSIONS: Human cells can demonstrate an adaptive response. This response is variable among cell lines and appears not to be related to radiosensitivity nor differences between tumour and normal cell lines.

Adaptation, Physiological↗
PubMed1998

Comparison of high dose rate, low dose rate, and high dose rate fractionated radiation for optimizing differences in radiosensitivities in vitro.

Radiotherapy is administered with the assumption that all patients respond similarly to radiation although radiosensitivity does vary from patient to patient, resulting in different degrees of early and late effects. Because the dose given to a patient is limited by the response of normal tissue in the treatment field, it would be beneficial to determine the sensitivity of this normal tissue prior to therapy. Previous studies to predict radiosensitivity have used surviving fractions after a single dose given in vitro, however, differences in cell survival at this low level of kill are not easy to resolve. In this study, we set out to evaluate the use of alternative dose regimens which may better resolve differences in radiosensitivity. We have examined several radiation protocols for predictive value, including survival after high doses (6 Gy) at both high (112 cGy/min) and low (.882 cGy/min) dose rates and after fractionated doses of 2 Gy (6 fractions). A sensitive human fibroblast line (S11358) cultured from a patient showing severe effects after therapy is compared with a cell line (OMB1) cultured from an apparently normal subject. Differences between these cell lines have been compared with those between two human melanoma cell lines (SKMEL3 and HT144) which have shown resistant and sensitive response to radiation in vitro respectively. In both fibroblast and melanoma cell lines, the difference in the survival of normal and sensitive cells increased with increasing dose regardless of whether irradiation was delivered as low dose rate, high dose rate, or as fractionated doses. We propose that radiation doses which more closely mimic clinical treatment are more suitable than surviving fraction after 2 Gy (SF2) for in vitro evaluation of relative radiosensitivities of cell populations.

Adult↗
PubMed1998

Interactions of mild hyperthermia, cisplatin and split dose irradiation in human ovarian carcinoma cells.

PURPOSE: Human ovarian cancer cells were evaluated to determine whether combination treatment with mild hyperthermia and cisplatin could inhibit repair of sublethal radiation damage. MATERIALS AND METHODS: Human ovarian carcinoma cell lines A2780S (parental line) and A2780CP (cisplatin resistant variant) were used in this study. Cisplatin at concentrations of 1 or 3 microg/ml was given concomitantly with 1 h of heating at 40 degrees C either immediately before or after irradiation. Survival was determined using a colony-forming assay. RESULTS: Neither mild hyperthermia nor cisplatin treatments alone affected sublethal damage repair. The combined treatment showed an effect in both cell lines and was treatment sequence-dependent. The effect was greater in the parental cell line. CONCLUSIONS: The data show that combined treatment of cisplatin and hyperthermia may have clinical efficacy at cisplatin concentrations and hyperthermia temperatures which by themselves have little to no effect.

Animals↗
PubMed1998

Modulation of radiation-induced strand break repair by cisplatin in mammalian cells.

PURPOSE: To investigate the repair of ionizing radiation-induced DNA lesions in human skin fibroblasts in the presence of cisplatin-DNA adducts and to determine the persistence of DNA repair inhibition by cisplatin. MATERIALS AND METHODS: Normal human fibroblasts (AG 1522) treated with cisplatin were exposed to 4 Gy 60Co gamma-radiation and assayed for repair of radiation-induced damage under growth-permissive conditions. DNA damage was measured by the fluorescence analysis of DNA unwinding (FADU) and cytokinesis-blocked micronucleus assays. RESULTS: Rejoining of strand breaks caused by 4 Gy radiation in cells without cisplatin pre-treatment appeared to be biphasic with an initial fast component (up to 15 min of repair time) followed by a slower component, and was completed by 90 min. Cisplatin treatment (10 microg/ml, 30 min) immediately before irradiation had no effect on the fast rejoining component, but inhibited the slow component (p<0.01). The same cisplatin treatment 24 h prior to irradiation inhibited both slow and fast components (p<0.01). In contrast, decreasing the cisplatin exposure to 1.0 microg/ml for 30 min, 24h prior to irradiation, resulted in an increased amount of strand break repair at each time point measured compared with irradiated control cells. This mild cisplatin treatment (95% survival) also resulted in a reduction of radiation-generated micronuclei indicating an adaptive response. CONCLUSIONS: Cisplatin used in combination with ionizing radiation can produce differential cellular responses depending upon the severity of the cisplatin treatment and the time interval between cisplatin and radiation exposures.

Cell Nucleus↗
PubMed1997

Comparison of hyperthermia radiosensitization and DNA polymerase inactivation in human normal and melanoma cell lines of different radiosensitivities.

Two human melanoma cell lines (one radiosensitive, HT144 and one radioresistant, SK Mel-3) and one normal human fibroblast (AG1522) were evaluated for thermal radiosensitization and the thermal enhancement ratios (TERs) were calculated. These were compared with residual polymerase activity to determine if this activity could be used to predict TERs. In all three cell lines, there was a good correlation between TER and residual polymerase alpha or beta activity. Polymerase beta was more sensitive than polymerase alpha as an indicator for TER. There were small cell line-dependent differences (not related to radiosensitivity) among the correlation curves, indicating that for each cell/tumor-type polymerase activity, vs. TER may have to be calibrated.

Animals↗
PubMed1997

Classification of 1H MR spectra of biopsies from untreated and recurrent ovarian cancer using linear discriminant analysis.

Proton (1H) magnetic resonance (MR) spectra of ex vivo biopsy samples of ovarian cancers provided biochemical information that was used to discriminate cancer from normal ovarian tissue. Possible differences present in intrinsically resistant tumors or changes in biochemistry after the induction of resistance were identified. Using multivariate techniques, in particular linear discriminant analysis (LDA), ovarian cancer was distinguished from normal ovarian tissue with a sensitivity of 100%, a specificity of 95% and an accuracy of 98%. Moreover, LDA was able to distinguish untreated ovarian cancer from recurrent ovarian cancer with a sensitivity of 92%, a specificity of 100%, and an accuracy of 97%; removal of the single "fuzzy" specimen increased the accuracy to 100%. Applications of this knowledge to in vivo measurements could lead to noninvasive diagnosis of ovarian cancer.

Biopsy↗
PubMed1997

Cell cycle perturbations in cisplatin-sensitive and resistant human ovarian carcinoma cells following treatment with cisplatin and low dose rate irradiation.

PURPOSE: To investigate cell cycle pertubations in plateau-phase human ovarian carcinoma cells following treatment with cisplatin, low dose-rate irradiation (LDRI), or combined cisplatin and LDRI, in order to understand cell cycle mechanisms by which these two treatment modalities interact. METHODS: Human ovarian carcinoma cells sensitive (A2780) and resistant (2780CP) to cisplatin were grown to plateau phase and given protracted cisplatin treatments (A2780 0.7 and 2 microg/ml; 2780CP 5 and 15 microg/ml) and/or LDRI (0.41 cGy/min). Cell cycle distribution following treatment was determined by two-parameter flow cytometry, based on bromodeoxyuridine (BrdU) uptake and DNA content using propidium iodide staining. RESULTS: The cisplatin-sensitive A2780 cells exposed to cisplatin alone for up to 28 h showed depletion of the G1 fraction and accumulation in S-phase, although the percentage of S-phase cells actively incorporating BrdU dropped to almost zero. The cisplatin-resistant 2780CP cells exposed to cisplatin alone showed a G1 arrest when exposed to 15 microg/ml, but not when exposed to 5 microg/ml. LDRI alone caused little cell cycle redistribution different from controls in either cell line. When LDRI was combined with cisplatin, no significant cell cycle redistribution was observed, apart from a decline in the actively incorporating S-phase fraction. CONCLUSIONS: Cisplatin caused A2780 cells to accumulate in nonincorporating S-phase, with no evidence of G1 arrest. Cisplatin-resistant 2780CP cells showed a G1 block when exposed to a high enough cisplatin concentration. This could indicate a mechanism of cisplatin resistance in these cells. LDRI alone or in combination with cisplatin did not result in significant cell cycle redistribution.

Bromodeoxyuridine↗
PubMed1997

Rearrangement of human cell homologous chromosome domains in response to ionizing radiation.

Chromosomes are located within the interphase nucleus in regions called domains. Using fluorescence in situ hybridization with whole chromosome paints, a pain of homologous chromosomes can be visualized as two discrete domains and their relative spatial location determined. This study examines the effects of an ionizing radiation exposure on the relative spatial location of chromosome 7 and 21 domains in human skin fibroblasts and lung endothelial cells. The distance between homologous chromosome domains was assessed for each nucleus, before and after exposure to ionizing radiation, using conventional epifluorescence and confocal laser scanning microscopy. Results from conventional microscopy indicated that homologous chromosome domains were re-positioned closer to each other within interphase nuclei after exposure to radiation. Analysis of three-dimensional data obtained from confocal microscopy confirmed these results. In control cells, and in cells examined immediately after irradiation, 66.2% +/- 2.1% of the homologous chromosome 21 domains within endothelial cell nuclei were located greater than 4.0 microns apart (33.8% +/- 1.9% were less than 4.0 microns apart). However, when cells were examined 2 h after a 4.0 Gy gamma-ray exposure, only 30.5% +/- 2.1% of the homologous chromosome domains were greater than 4.0 microns apart (69.5% +/- 2.1% were less than 4.0 microns apart). Similar results were obtained for chromosomes 7 and 21 in skin fibroblast nuclei. The results indicate that homologous chromosome domains rearranged and became closer together within the interphase nuclei in response to ionizing radiation. The exact mechanism of this response is unknown, but it may be related to DNA repair processes. It is speculated that chromosome domains are re-positioned to permit repair of radiation-induced DNA damage.

Cell Nucleus↗
PubMed1996

Cisplatin and low dose rate irradiation in cisplatin resistant and sensitive human glioma cells.

PURPOSE: Human glioma cell lines resistant (U373MGCP) and sensitive (U373MG) to cisplatin were used to evaluate the effect of cisplatin as a sensitizer to low dose rate irradiation (LDRI). METHODS AND MATERIALS: A cisplatin resistant glioma cell line U373MGCP was developed by chronic exposure of parental U373MG cells to cisplatin. Plateau phase cells were treated with cisplatin, high dose rate (HDR) irradiation (1.12 Gy/min), LDRI (0.0088 Gy/min), or cisplatin concurrent with LDRI. Cell survival was determined by the colony forming assay. RESULTS: Both cell lines showed increased resistance to radiation at LDR compared with HDR, with Dose Modifying Factors (DMF at 10% survival level) of 1.7 for U373MG and 2.5 for U373MGCP. The increased LDR sparing effect in the cisplatin resistant U373MGCP cells indicates increased repair proficiency. The resistant cell line showed a fourfold increase in resistance to cisplatin cytotoxicity at the 10% survival level compared with the parental U373MG cells. Cisplatin enhanced the response of both cell lines to LDRI. The DMFs were 1.2, 1.2, and 1.7, respectively, for the sensitive U373MG cell line given 1 microgram/ml, and the resistant cell line given 3 or 6 micrograms/ml cisplatin treatments concurrent with LDRI. CONCLUSIONS: These data show that cisplatin can be an effective sensitizer to LDRI in both cisplatin resistant and sensitive glioma cell lines. However, in the resistant cell line, higher concentrations of cisplatin were necessary to achieve the same level of sensitization as in the sensitive cell line.

Astrocytoma↗
PubMed1996

Multiorgan fatal chronic complications following radiation treatment for cancer of the cervix--results of fibroblast assay.

We report a case of severe chronic normal tissue injury following conventional irradiation in a 33-year-old patient with a strong family history of cancer. Her cultured fibroblasts exhibited a response to low-dose-rate irradiation similar to fibroblasts taken from a normal individual. For acute irradiation her fibroblasts showed a slight increased sensitivity to single doses compared to fibroblasts from a normal individual. This difference was much larger when fractionated irradiation was given. It is concluded that the hypersensitive response of this patient was due to the fractionated radiotherapy and not to the low-dose-rate brachytherapy. We believe that this is the first report of this form of cellular radiosensitivity which is mainly expressed in fractionated radiotherapy.

Adult↗
PubMed1996

Cytotoxicity, accumulation, and efflux of cisplatin and its metabolites in human ovarian carcinoma cells.

Cisplatin (CP) is one of the most useful antineoplastic drugs. When CP is dissolved in human plasma, different metabolites are formed. Using the OV 2008 human ovarian cancer cell line, we examined the relative cytotoxicities of CP and its metabolites (aquated CP [AP], monomethionine CP [MP], bismethionine CP [BP], and a mixture of CP metabolites in ultrafiltrated human plasma [UP]) in vitro. By clonogenic assay, cell survival (percent of mean +/- SE) of OV 2008 cells exposed for 1 hr to 6.7 microM of CP was 9.8 +/- 0.7 and for its equal platinum contents of AP, MP, BP, and UP solutions were 3.3 +/- 0.7, 9.8 +/- 0.9, 15.9 +/- 1.1, 76.8 +/- 2.1, and 13.1 +/- 0.7, respectively. AP was the most cytotoxic species, and BP was the least cytotoxic species. Cellular platinum uptake (ng/10(6) cells) after addition of 0.33, 1.6, and 2.5 mM of each species for 1 hr was measured and a strong correlation was found between cytotoxicity of each CP metabolite and its corresponding cellular platinum (Pt) uptake (r = 0.997). There was a strong correlation between cytotoxicity of the CP metabolites and their DNA binding. With fractionation of these cells into DNA, nucleoplasm and cytoplasm, the highest platinum content was found on DNA. The most important factor that seems to be responsible for the cytotoxicities of the different CP metabolites is the amount of their associated intracellular accumulation, and particularly the degree of their binding to DNA.

Carcinoma↗
PubMed1996

Active vs. passive resistance, dose-response relationships, high dose chemotherapy, and resistance modulation: a hypothesis.

With chemotherapy, the in vitro and clinical dose-response curve is steep in some situations, but is relatively flat in others, possibly due to the mechanism by which tumors are resistant to chemotherapy. For tumors with resistance due to factors that actively decrease chemotherapy efficacy (e.g., p-glycoprotein, glutathione, etc.), one would predict that high dose chemotherapy and therapy with some resistance modulating agents would increase therapeutic efficacy. Such "active" resistance would most likely generally arise from gene amplification or over expression, and would be characterized by a shoulder on the log response vs. dose curve, with eventual saturation of the protective mechanism. On the other hand, one would expect that high dose chemotherapy and most resistance modulating agents would be of little value for tumors with resistance due to defective apoptosis or due to a deficiency in or decreased drug affinity for a drug target, drug activating enzyme, drug active uptake system, or essential cofactor. Such "passive" resistance would most likely generally arise from gene down regulation, deletion, or mutation, and would probably be characterized by a relatively flat log response vs. dose curve, or by a curve in which a steep initial section is followed by a plateau, as target, etc., is saturated. (If response were plotted vs. log dose, then compared to the curve for a sensitive cell line, the curve for active resistance would be analogous to the pharmacodynamic curve seen with competitive antagonism [i.e., a sigmoid curve shifted to the right], and the curve for most types of passive resistance would be analogous to the pharmacodynamic curve seen with noncompetitive antagonism [i.e., a sigmoid curve with reduced maximal efficacy]. As such, one might also refer to active vs. passive resistance as competitive vs. noncompetitive resistance, respectively.) Many tumor types probably possess a combination of active and passive mechanisms of resistance. New in vivo strategies could be helpful in defining dose-response relationships, mechanisms of resistance, and targets for resistance modulation. Such in vivo studies would be conducted initially in animals, but might also be tested clinically if animal studies demonstrated them to be feasible and useful. These in vivo studies would be conducted by randomizing 5-25 subjects to one of 10-20 dose levels over a potentially useful therapeutic range. Nonlinear regression analysis would then be used to define the characteristics of a curve generated by plotting against dose the log percent tumor remaining after the first course of therapy. While this might offer insight into the nature of resistance mechanisms present initially, plotting further tumor shrinkage vs. dose-intensity vs. course number for each later treatment course (or plotting dose-intensity vs. time to tumor progression) might provide information on how tumors become increasingly resistant to drugs following treatment.

Antineoplastic Agents↗
PubMed1996

A comparison of hyperthermia cisplatin sensitization in human ovarian carcinoma and glioma cell lines sensitive and resistant to cisplatin treatment.

Two pairs of human tumor cell lines (glioma and ovarian carcinoma (OvCa) each having a parental cell line and cisplatin-resistant variant, were evaluated for (a) cisplatin response, (b) hyperthermia response, and (c) combined hyperthermia and cisplatin response. The two resistant lines had comparable resistant responses while for the parental lines, the OvCa was more sensitive than the glioma to cisplatin doses up to 14 microgram/ml. For the hyperthermia response, the OvCa parental line was more resistant than the variant line at low-temperature hyperthermia (41 degrees C or 42 degrees C) but became more sensitive at high temperature (45 degree C). For the glioma, the parental line was more sensitive to hyperthermia at all temperatures tested. Hyperthermia caused sensitization to cisplatin in all cell lines but was generally greater in the glioma cell lines. In the OvCa system, hyperthermia had a slightly greater sensitizing effect on the resistant cell lines, while in the glioma the opposite was true. The degree of sensitization increased with hyperthermia temperature. In summary, the results showed that there is no cross- resistance for hyperthermia and cisplatin, that the degree of thermal sensitization is not reduced in cisplatin- resistant cell lines, and that cisplatin thermal sensitization is cell-line and temperature dependent. Thus, hyperthermia can effectively improve tumor cell response to cisplatin and may be useful in overcoming resistance to cisplatin.

Carcinoma↗
PubMed1996

Modelling of continuous low dose rate and accelerated fractionated high dose rate irradiation treatments in a human glioma cell line.

The Incomplete-Repair model of Thames in its original and in its two-repair processes forms is used to analyse continuous and fractionated irradiation of a glioma cell line treated in vitro. Furthermore, the Incomplete-Repair model is rederived based on the linear-quadratic model bearing a cubic term. Our results show a potentially enhanced but nonsignificantly improved fit to the data when comparing the cubic to the original form of this model. The repair half-time values showed a decrease, followed by an increase, as a function of small doses (< 1.5Gy) per fraction. This is observed using both the original and the cubic forms of the model. We propose this behaviour to be due to an induced resistance of the repair system followed by a saturation process. Two-repair processes were not seen directly due to the large scatter in the fast and the slow components of repair. Repair half-time values are estimated for various dose per fraction protocols using the original and extended forms of the Incomplete-Repair model. Two-repair processes that are consistent as a function of dose per fraction were not detectable in a glioma cell line treated in vitro.

Dose-Response Relationship, Radiation↗
PubMed1996

Concomitant low dose-rate irradiation and cisplatin treatment in ovarian carcinoma cell lines sensitive and resistant to cisplatin treatment.

Human ovarian carcinoma parental and cisplatin-resistant cells were evaluated for their radiation sensitivity to high and low dose-rate irradiation and for the effectiveness of cisplatin in radiosensitization. The cisplatin resistant variant A2780cp showed increased radiation resistance for both low dose-rate (LDRI) and high dose-rate irradiation. For cisplatin treatment for 1 h before and after HDRI there was radiosensitization in only the cisplatin-resistant variant. Concomitant cisplatin treatment during LDRI resulted in radiosensitization in both cell lines with dose-modifying factors ranging from 1.6 to 5.8. In this case greater radiosensitization was achieved in the parental cell line. In both cell lines the dose-modifying factors were larger when the cisplatin was refreshed every 6 h instead of 12 h during LDRI. These data show that cisplatin may be a very effective radiosensitizer when given during LDRI which is used in brachytherapy.

Antineoplastic Agents↗