The drug resistance-related protein LRP is the human major vault protein.

Multidrug-resistant cancer cells frequently overexpress the 110-kD LRP protein (originally named Lung Resistance-related Protein). LRP overexpression has been found to predict a poor response to chemotherapy in acute myeloid leukaemia and ovarian carcinoma. We describe the cloning and chromosome localization of the gene coding for this novel protein. The deduced LRP amino acid sequence shows 87.7% identity with the 104-kD rat major vault protein. Vaults are multi-subunit structures that may be involved in nucleo-cytoplasmic transport. The LRP gene is located on chromosome 16, close to the genes coding for multidrug resistance-associated protein and protein kinase C-beta, and may mediate drug resistance, perhaps via a transport process.

Extramural Venous Invasion as Prognostic Factor of Recurrence in Stage 1 and 2 Colon Cancer.

AIM: Extramural venous invasion (EMVI) is a prognostic indicator in patients with colorectal cancer. However, its additional value in patients with stage 1 and 2 colorectal cancer is uncertain. In the present study, the incidence of EMVI and the hazard ratio for recurrence in patients with stage 1 and 2 colon cancer were studied. METHODS: 184 patients treated for stage 1 and 2 colon cancer were included with a follow-up of at least 5 years. Chart review was performed and EMVI was assessed by two separate pathologists. EMVI was scored with additional caldesmon staining on the resection specimen. Primary outcomes were recurrence-free survival (RFS) measured through the Cox regression analysis and prevalence of EMVI. RESULTS: There were 10 cases of EMVI and 3 cases of intramural venous invasion (IMVI) all occurring in patients with stage 2 disease corresponding to a prevalence of 9%. Thirty-one percent of the patients with venous invasion experienced recurrence versus 14% in patients without, corresponding with a hazard ratio of 2.39 (p = 0.11). CONCLUSION: The present study demonstrates a trend towards an increased risk of recurrence in patients with stage 2 colon cancer with venous invasion. This warrants consideration of adjuvant chemotherapy despite the lack of lymph node metastases.

Drug resistance-associated marker Lrp for prediction of response to chemotherapy and prognoses in advanced ovarian carcinoma.

BACKGROUND: Drug resistance is a major impediment to the successful treatment of ovarian carcinoma. None of the earlier-identified resistance mechanisms, such as overexpression of the MDR1 gene product, P-glycoprotein (Pgp), has been shown to be a major determinant of clinical response to chemotherapy and survival for ovarian cancer patients. The multidrug resistance-associated protein (Mrp) and the lung resistance protein (Lrp, also called the p110 major vault protein), are newly described proteins associated with multidrug resistance in vitro. PURPOSE: The aim of this retrospective study was to investigate the expression of Mrp and Lrp, in addition to Pgp, in advanced ovarian carcinoma and to determine whether such expression was predictive of response to chemotherapy and survival. METHODS: Fifty-seven banked frozen specimens, previously collected and frozen at the time of diagnosis from an equal number of patients with International Federation of Gynecology and Obstetrics (FIGO) stage III or IV ovarian carcinoma, were immunostained for Pgp (with monoclonal antibodies [MAbs] MRK-16 and JSB-1), Mrp (with MAb MRPrl), and Lrp (with MAb LRP-56). All patients had received platinum- or alkylating-based chemotherapy after debulking surgery. Clinicopathologic parameters determined at diagnosis were retrospectively assessed for their relationship with Pgp, Mrp, and Lrp expression. Response to treatment and survival were compared between Pgp, Mrp, and Lrp expression groups. Qualitative variables were analyzed using Fisher's exact test or the chi-squared test. All reported P values are two-tailed. RESULTS: Nine (16%), 39 (68%), and 44 (77%) of the 57 tumor specimens examined showed positive immunostaining for Pgp, Mrp, and Lrp, respectively. Positive immunostaining for these proteins was not associated with any other prognostic factor examined. No association was found between Pgp and Mrp expression and response to chemotherapy and survival. In contrast, patients with Lrp-positive tumors had poorer response to chemotherapy (P = .004) and shorter progression-free (P = .003) and overall (P = .007) survival than Lrp-negative patients. Multivariate analysis of Lrp expression, FIGO stage, residual tumor after initial surgery, tumor grade, and presence or absence of ascites showed that only Lrp status was independently related to both progression-free survival and overall survival. CONCLUSIONS: Positive Lrp immunostaining in advanced ovarian carcinoma appears to be an indicator of poor response to standard chemotherapy (platinum or alkylating agents) and of adverse prognoses. IMPLICATIONS: The functional characterization of Lrp and related proteins may reveal new approaches to modulate Lrp-associated drug resistance. A large prospective study is warranted to confirm the prognostic value of Lrp.

Immunochemical detection of the multidrug resistance-associated protein MRP in human multidrug-resistant tumor cells by monoclonal antibodies.

We have generated rat and murine monoclonal antibodies against multidrug resistance-associated protein (MRP), a M(r) 180,000-195,000 membrane glycoprotein involved in a non-P-glycoprotein multidrug resistance of human tumor cells. The antibodies were raised against two different segments of MRP and found to be suitable for protein blot analyses, immunohistochemical and cytochemical studies, as well as flow cytometry of permeabilized cells. The antibodies do not cross-react with the human P-glycoproteins. Immunocytochemistry using MRP-overexpressing tumor cells of different histogenetic origins showed that MRP is predominantly located in the plasma membrane. Immunoelectron microscopy confirmed the plasma membrane location of MRP. The MRP antibodies provide a sensitive and specific tool for studies on MRP-mediated multidrug resistance.

Cytokeratin phenotyping does not help in distinguishing oesophageal adenocarcinoma from cancer of the gastric cardia.

BACKGROUND: It is sometimes difficult to distinguish between cardia cancer and oesophageal cancer. AIMS: To evaluate whether cytokeratin (CK) expression of the tumour can be of value in differentiating between the two tumour types. METHODS: Consecutive patients with a malignant tumour in the oesophagus or stomach were recruited. Biopsy specimens were taken for routine haematoxylin and eosin staining. One tissue block with representative tissue was selected for immunohistochemical staining (CK7 and CK20). RESULTS: Endoscopically located adenocarcinoma of the oesophagus was present in 84 patients (64 men, 20 women; mean age, 68 years; range, 44-91). Cancer located primarily in the gastric cardia was present in 63 patients (42 men, 21 women; mean age, 68 years; range, 42-88). The histological diagnosis was metastasis from a primary tumour outside the oesophagus or stomach in 19 patients. The patients were divided into three groups for the immunohistochemical analysis. Patients in group A had definite oesophageal cancer, group B patients had a definite carcinoma located in the gastric cardia, and group C patients had an obstructing tumour distal in the oesophagus at the level of the diaphragm, which could not be passed with the endoscope. Paraffin wax embedded material was available from 122 patients for immunostaining and CK analysis. There was no significant difference in expression or distribution of CK7 or CK20 in the three groups of patients. CONCLUSION: CK phenotyping cannot distinguish between cancer arising from a Barrett's oesophagus and carcinoma originating in the gastric cardia.

Optimal immunocytochemical and flow cytometric detection of P-gp, MRP and LRP in childhood acute lymphoblastic leukemia.

The clinical relevance of multidrug resistance (MDR)-related proteins in childhood acute lymphoblastic leukemia (ALL) is largely unknown. The diversity of techniques, fixation methods, storage of cells (fresh or cryopreserved) etc, may contribute to discrepancies observed between several studies. We therefore optimized the detection of P-glycoprotein (P-gp), MDR-associated protein (MRP) and lung resistance-related protein (LRP) by immunocytochemistry and flow cytometry in childhood ALL cells. Thirteen fixation methods were compared using six antibodies in both immunocytochemistry and flow cytometry. The optimal fixation for P-gp (C219, MRK16), MRP (MRPr1) and LRP (LRP56) was a mixture of 2% (v/v) formaldehyde solution and acetone incubated for only 10 s at room temperature (FAc). For MRP recognized by MRPm6, the optimal fixation condition was acetone for 5 min at room temperature in immunocytochemistry, and methanol for 15 min at -20 degrees C in flow cytometry. P-gp staining by 4E3 was strongly antibody batch-dependent; on cytospins FAc fixation was optimal, but inconclusive data were obtained by flow cytometry. The optimized fixation conditions on fresh samples revealed a day-to-day variation in staining (both increasing and decreasing) in one third of the immunocytochemical tests. In flow cytometry the day-to-day variation in the fluorescence index was -1 +/- 22%. In both techniques, staining was comparable between fresh and cryopreserved cells. We recommend the use of the above mentioned fixation methods in order to study the clinical relevance of P-gp, MRP and LRP in childhood ALL.

Expression of the multidrug resistance-associated protein (MRP) gene in human cancers.

We determined the expression of a newly recognized drug resistance gene, the multidrug resistance-associated protein (MRP) gene, [Cole et al., Science (Washington DC), 258: 1650-1654, 1992], in normal human tissues and in >370 human tumor biopsies using a quantitative RNase protection assay and immunohistochemistry. MRP mRNA appeared to be ubiquitously expressed at low levels in all normal tissues, including peripheral blood, the endocrine glands (adrenal and thyroid), striated muscle, the lymphoreticular system (spleen and tonsil), the digestive tract (salivary gland, esophagus, liver, gall bladder, pancreas, and colon), the respiratory tract (lung), and the urogenital tract (kidney, bladder, testis, and ovary). The human cancers analyzed could be divided into three groups with regard to MRP expression. Group 1 consists of tumors that often exhibit high to very high MRP mRNA levels (e.g., chronic lymphocytic leukemia). Group 2 comprises the tumors that often exhibit low, but occasionally exhibit high MRP mRNA expression (e.g., esophagus squamous cell carcinoma, non-small cell lung cancer, and acute myelocytic leukemia). Group 3 comprises the tumors with predominantly low levels of MRP mRNA, comparable to the levels found in normal tissues (e.g., other hematological malignancies, soft tissue sarcomas, melanoma, and cancers of the prostate, breast, kidney, bladder, testis, ovary, and colon). Using the MRP-specific mAbs MRPr1 and MRPm6, we confirmed the elevated MRP mRNA levels in tumor tissues by immunohistochemistry. We conclude that hyperexpression of MRP is observed in several human cancers, and that additional studies are needed to assess the clinical relevance of MRP.

Multidrug resistance-related proteins in primary choroidal melanomas and in vitro cell lines.

PURPOSE: Metastatic uveal melanoma is strongly resistant to chemotherapy, and multidrug resistance (MDR) may be involved. To investigate the role of MDR, the presence of the MDR-associated proteins P-glycoprotein (Pgp), MRP, and lung resistance protein (LRP) was determined on primary choroidal melanomas and cell lines. METHODS: A panel of primary choroidal melanomas was examined for the presence of MDR-associated proteins by immunohistochemical analysis. In cell lines established from four primary choroidal melanomas and one metastatic choroidal melanoma, the expression of MDR-associated proteins was determined with monoclonal antibodies in cytospin preparations and flow cytometry. In addition, the functional capacities of transporter proteins Pgp and MRP as adenosine triphosphate-driven efflux pumps were determined by measuring the cellular accumulation and efflux of the fluorescent dyes rhodamine 123 and calcein-AM, with and without the presence of specific pump inhibitors PSC833 and probenecid. RESULTS: Low levels of Pgp and MRP were detected in most primary tumors and in some cell lines. Measurable transporter function of Pgp could be determined in cell line OCM-1. Lung-resistance protein was present in all primary tumors and cell lines and showed high expression levels. CONCLUSIONS: This study revealed the involvement of LRP and at least a minor role of Pgp and MRP in chemoresistance of choroidal melanoma. Compared with cutaneous melanomas, uveal melanomas appear to express slightly higher levels of Pgp. These findings provide insights into the drug-resistant phenotype of this disease and can aid in the design of therapeutic protocols.

Increased expression of multidrug resistance related proteins Pgp, MRP1, and LRP/MVP occurs early in colorectal carcinogenesis.

AIM: To analyse the expression of multidrug resistance (MDR) related proteins at different steps in colorectal carcinogenesis. METHODS: The presence of three MDR related proteins (Pgp, MRP1, and LRP/MVP) was studied by means of immunohistochemistry in normal, adenomatous, and malignant colorectal epithelium. Formaldehyde fixed, paraffin embedded tissue sections of 17 samples of colorectal tissue were used (normal mucosa, n = 4; adjacent mucosa, n = 5; adenoma, n = 5; carcinoma, n = 3). RESULTS: For all three proteins, expression was found in the surface epithelium and the upper parts of the crypts in normal colon. In the adenomas, staining was seen along the complete length of the crypts. In the carcinomas analysed, all epithelium showed positive staining. Mucosa adjacent to either carcinoma or adenoma showed staining patterns mostly resembling those of normal mucosa, but sometimes some extension of staining was seen along the crypt. CONCLUSIONS: These proteins already show increased expression in the adenoma stage. In the absence of adequate mucin production in adenomas, MDR related proteins could be an important factor in protecting the epithelium against further environmentally induced genetic damage. This could be one of the reasons why only about 5% of colorectal adenomas will actually progress to carcinomas.

Immunohistochemical analysis of drug resistance-associated proteins in ovarian carcinomas.

Loss of function of the tumor suppressor gene p53, increased expression of glutathione-S-transferase pi (GST7pi) and the major vault protein are involved in drug resistance of ovarian carcinomas. However, a study comparing these factors has not yet been performed. Therefore, paraffin-embedded material of 213 ovarian tumors with well-documented follow-up was used for immunohistochemical analysis of p53 protein, GSTpi, and major vault protein (antibodies LRP-56, LMR-5). Forty-six percent of the cases showed nuclear p53 accumulation. Strong immunoreactivity for GSTpi, LRP-56, and LMR-5 was seen in 50%, 36%, and 47%, respectively. p53 positivity was most often found in serous carcinomas (p < 0.05). Strong GSTpi expression was the only factor that correlated with clinical resistance to chemotherapy (p = 0.04). In the whole group, as well as in FIGO III cases stratified for residual disease < or = and >2 cm, p53 and GSTpi correlated with an adverse outcome (p = 0.01 for p53 and p = 0.04 for GSTpi). Strong LRP-56 or LMR-5 staining was associated with a tendency towards poorer prognosis, without reaching statistical significance. In multivariate analysis for FIGO III, only residual disease and p53 proved to be independent prognostic factors. Our observations confirm the prognostic significance of p53 accumulation in ovarian carcinomas. Only GSTpi immunoreactivity was significantly correlated with drug resistance.

Multidrug resistance related proteins in primary cutaneous lymphomas.

BACKGROUND: In patients with primary cutaneous B-cell lymphomas (CBCL) and primary cutaneous T-cell lymphomas (CTCL), extracutaneous sites may become involved and then polychemotherapy is indicated. Multi-agent chemotherapy may induce long lasting complete remissions in CBCL's. Most CTCL's, especially mycosis fungoides (MF), and CD30 negative primary cutaneous large T-cell lymphoma (PCLTCL) respond poorly or partially to Multi-agent Chemotherapy. PURPOSE: We have studied whether cutaneous lymphomas express the following multidrug resistance (MDR) related proteins: multidrug resistance protein (MRP), lung resistance protein (LRP) and P-glycoprotein (Pgp). METHODS: From the files of the Dutch Cutaneous Lymphoma Working Group we selected pretreatment punch biopsy specimens of the skin from 14 patients with MF, 10 patients with a PCLTCL and 8 patients with a CBCL. In several patients with a clinical relapse of their disease after multi-agent chemotherapy, punch biopsy specimens of cutaneous lesions were available (6 MF, 3 PCLTCL, 1 CBCL). Benign dermatoses with a dense lymphoid infiltrate were included as a control. Immunohistochemistry was done on formalin fixed, paraffin-embedded punch biopsy specimens with monoclonal antibodies MRPrl (anti-MRP); LRP-56 (anti-LRP); C219 (anti-Pgp). Staining was performed by the biotin-streptavidin immunoperoxidase method. RESULTS: MRPrl staining was found in the cytoplasm of > or = 5%-50% of lymphoid cells in 13 out of 14 cases of MF and in 6 out of 10 patients with a PCLTCL. In 2 out of 8 cases of CBCL > or = 5%-50% positive tumorcells were found. Strong staining (> or = 50% of the cells positive) was found in 10 out of the total of 24 CTCL cases. LRP56 staining of lymphoid cells was found in 1 out of 14 cases of MF and in 1 out of 10 cases of PCLTCL and in 1 out of 8 cases of CBCL. C219 expression was found in 4 out of 10 cases of PCLTCL and in 2 out of 8 cases of CBCL. After chemotherapy both a higher staining intensity and a higher number of positive cells were found with MRPrl especially in patients with MF. CONCLUSION: The present study shows that lymphoid cells in both primary cutaneous lymphomas and benign skin disorders may express MDR related proteins and that the expression profile of these proteins is roughly related to the tumor cell phenotype. However, the functional role of these proteins in clinical drug resistance in primary cutaneous lymphomas has to be proven.

The incidence of colorectal cancer is decreasing in the older age cohorts in the zaanstreek region in the Netherlands: an age-cohort effect.

Introduction. Colorectal cancer (CRC) has a high incidence. Removal of adenomas, the precursor lesion, could be helpful in the prevention of cancer. Aim. To investigate the yearly incidence of CRC in consecutive years. Patients and Methods. All patients diagnosed with CRC in the years 1990 till 2010 were studied. Date of diagnosis, age at time of diagnosis, gender, and localisation of the tumour were assessed. Results. A total of 1575 incident CRC were diagnosed, 865 men (55%) and 710 women (45%). CRC occurred more often in men. In the course of the years, the occurrence of CRC increased. After exclusion of rectal cancer, the percentage of proximal cancer in the colon shows a trend towards increase in the consecutive years. In the twenty consecutive years, the population of the Zaanstreek region increased from 130.000 to 145.330. There was a significant increase of CRC in the age cohort 51-70 in the period of twenty years, while a significant decreasing incidence of cancer was seen in patients above 71 years. Conclusion. The decreasing incidence of colorectal cancer in the age cohorts above 71 years possibly reflects indirect evidence of an age-cohort effect due to removal of adenomas in these age cohorts earlier in life.

What is the consequence of hyperplastic polyps? Do adenomas and colorectal cancer develop in these patients? A clinical study.

AIM: Do patients with hyperplastic polyps (HP) have an increased risk for developing adenomas and colorectal cancer (CRC)? A study was done to detect the number of patients developing adenomas and CRC. MATERIAL AND METHODS: From 1990-1995 all patients with a HP diagnosed via endoscopy and significant follow-up were studied. The patients were separated in three groups; Group 1 HP in patients with previous adenoma and/or CRC. Group 2 HP with a concurrent adenoma and/or CRC. Group 3 patients with only HP. RESULTS: Group 1 consisted of 20 patients, group 2 of 39 patients, and group 3 of 136 patients The follow-up was 12.5, 12.6, and 13.4 years respectively. In group 1 there was one patient with an adenoma in the index investigation.In group 2 adenomas were seen coinciding to hyperplastic polyp(s) in 29 patients, while in 11 cases there was a colorectal cancer. Obviously patients in group 3 only had hyperplastic polyps. In group 1 7 patients previously had an adenoma and 12 previously had colorectal cancer. Four patients in group 3 developed cancer : 13, 14, 15, and 15 years after the detection of a hyperplastic polyp. In group 2 only one patient developed cancer in the cecum 9 years after the index investigation. In the four patients of group 3 who developed cancer no one previously had a serrated adenoma at revision of the original histology. Twelve patients developed an adenoma. CONCLUSION: Only five CRC's developed in patients with HP. Thus, at the best the risk for developing CRC in patients with HP is not very high and equals that of adenomas.

Collagenous colitis: description of a single centre series of 83 patients.

INTRODUCTION: Collagenous colitis can be the cause of chronic diarrhoea. AIM: Assess the prevalence of collagenous colitis in a single centre. PATIENTS: All records of patients with diarrhoea in whom histological examination showed collagenous colitis, were retrieved. Demographic data, clinical presentation, associated diseases and treatment were studied. RESULTS: In a period of 15 years 83 patients were identified with collagenous colitis. These were 16 men and 67 women, mean age 60 years (range 20-87) at time of diagnosis. Thirty four patients (38%) complained of mushy stools and 49 (62%) of watery diarrhoea. Eight patients had rectal bleeding. Mucous discharge was noted by 18 patients. There was no weight loss in 55 patients. Six patients complained of loss of appetite, 9 had nausea, and 2 complained of vomitus. A macroscopically normal colon was present in 63 patients. Associated diseases, like celiac disease and hypothyroidism, only were seen in women. Twenty eight patients did not receive any treatment, ten patients received mesalazine. One patient was treated with steroids. Fourteen patients were treated for accompanying bacterial overgrowth. Fourteen patients used loperamide. Budesonide was applied with success in 17 patients. During follow-up 58 patients had no complaints anymore, 21 had mild diarrhoea, 3 moderate, while it was unknown in one patient. CONCLUSION: Collagenous colitis has a higher prevalence as usually reported. There is an association with auto-immune disorders and dysbacteriosis.

Calciphylaxis in chronic renal failure: An approach to risk factors.

We present a case of calcifying panniculitis due to calciphylaxis in a nontherapy compliance 65-year-old man suffering from chronic renal failure. Calciphylaxis, a life threatening condition, is characterized by high calcium x phosphate product, presence of calcium crystals in the skin and secondary hyperparathyroidism. The clinical presentation includes painful firm plaques, which could progress to nonhealing ulcers. A review of literature is given with emphasis on identification of risk factors and early diagnosis.

Relationship of LRP-human major vault protein to in vitro and clinical resistance to anticancer drugs.

Multidrug resistance (MDR) has been related to two members of the ABC-superfamily of transporters, P-glycoprotein (Pgp) and Multidrug Resistance-associated Protein (MRP). We have described a 110 kD protein termed the Lung Resistance-related Protein (LRP) that is overexpressed in several non-Pgp MDR cells lines of different histogenetic origin. Reversal of MDR parallels a decrease in LRP expression. In a panel of 61 cancer cell lines which have not been subjected to laboratory drug selection, LRP was a superior predictor for in vitro resistance to MDR-related drugs when compared to Pgp and MRP, and LRP's predictive value extended to MDR unrelated drugs, such as platinum compounds. LRP is widely distributed in clinical cancer specimens, but the frequency of LRP expression inversely correlates with the known chemosensitivity of different tumour types. Furthermore, LRP expression at diagnosis has been shown to be a strong and independent prognostic factor for response to chemotherapy and outcome in acute myeloid leukemia and ovarian carcinoma (platinum-based treatment) patients. Recently, LRP has been identified as the human major protein. Vaults are novel cellular organelles broadly distributed and highly conserved among diverse eukaryotic cells, suggesting that they play a role in fundamental cell processes. Vaults localise to nuclear pore complexes and may be the central plug of the nuclear pore complexes. Vaults structure and localisation support a transport function for this particle which could involve a variety of substrates. Vaults may therefore play a role in drug resistance by regulating the nucleocytoplasmic transport of drugs.

Overexpression of the ABC transporter TAP in multidrug-resistant human cancer cell lines.

Multidrug resistance (MDR) to anti-cancer drugs has been associated with the overexpression of P-glycoprotein (P-gp) and the multidrug resistance-associated protein (MRP), both being members of the ATP-binding cassette (ABC) superfamily of transporters. We investigated whether in addition to P-gp and MRP, another ABC transporter, the transporter associated with antigen processing (TAP), is associated with MDR. TAP plays a major role in MHC class I-restricted antigen presentation by mediating peptide translocation over the endoplasmic reticulum membrane. TAP1 and P-gp share a significant degree of homology among their transmembrane domains, which are thought to be the primary determinants of substrate specificity, and both can apparently mediate the translocation of peptides. Using immunocytochemistry and Western blot, TAP was overexpressed in parallel with MHC class I in several MDR human cancer cell lines. TAP was overexpressed more frequently in MRP-positive MDR cell lines (three out of three) than in P-gp positive MDR cells (two out of five). Reversal of resistance resulted in a decrease in TAP levels. Transfection of the TAP genes into TAP-deficient lymphoblastoid T2 cells conferred mild resistance to etoposide, vincristine and doxorubicin (2- to 2.5-fold). Furthermore, etoposide and vincristine inhibited TAP-dependent peptide translocation to the endoplasmic reticulum. Collectively, our results suggest that TAP may modestly contribute to the MDR phenotype, in particular in MRP- overexpressing MDR cells. Further insight into the role of TAP in MDR will require the study of other transfectants, as well as the investigation of TAP expression in P-gp and MRP-negative MDR cancer cell lines.

Characterization of a clonal human colon adenocarcinoma line intrinsically resistant to doxorubicin.

Intrinsic low-level resistance to anti-cancer drugs is a major problem in the treatment of gastrointestinal malignancies. To address the problem presented by intrinsically resistant tumours, we have isolated two monoclonal lines from LoVo human colon adenocarcinoma cells: LoVo/C7, which is intrinsically resistant to doxorubicin (DOX); and LoVo/C5, which shows the same resistance index for DOX as the mixed parental cell population. For comparison, we have included in the study a LoVo-resistant line selected by continuous exposure to DOX and expressing a typical multidrug resistant (MDR) phenotype. In these cell lines we have studied the expression and/or activity of a number of proteins, including P-glycoprotein 170 (P-gp), multidrug resistance-associated protein (MRP), lung resistance-related protein (LRP), glutathione (GSH)-dependent enzymes and protein kinase C (PKC) isoforms, which have been implicated in anti-cancer drug resistance. Intracellular DOX distribution has been assessed by confocal microscopy. The results of the present study indicate that resistance in LoVo/C7 cells cannot be attributed to alterations in P-gp, LRP or GSH/GSH-dependent enzyme levels. Increased expression of MRP, accompanied by alterations in the subcellular distribution of DOX, has been observed in LoVo/C7 cells; changes in PKC isoform pattern have been detected in both intrinsically and pharmacologically resistant cells.