Economic evaluation of arsenic trioxide for treatment of newly diagnosed acute promyelocytic leukaemia in Canada.

To assess, from a Canadian perspective, the economic impact of arsenic trioxide (ATO) + all-trans retinoic acid (ATRA) for treating newly diagnosed acute promyelocytic leukaemia (APL), the cost-effectiveness of ATO + ATRA compared to ATRA + idarubicin (IDA) was assessed over a lifetime horizon using a time-dependent Markov model. The model considers four health states: complete remission, treatment failure or relapse, post-failure, and death. Markov cycle length was 1 month for the first 48 months and 1 year thereafter. Efficacy outcomes in terms of event-free survival and overall survival were taken from a head-to-head clinical trial. Costs were associated with drug and administration, adverse events (AEs), treatment of relapses, follow-up visits, and productivity losses. Utilities and disutilities associated with health states and AEs were derived from the literature. Compared to ATRA + IDA, ATRA + ATO is associated with incremental cost-effectiveness ratios (ICERs) of $CAD50,193/quality-adjusted life years (QALY) and $CAD50,338/QALY from a Canadian Ministry of Health (MoH) and societal perspectives, respectively. Results of the one-way sensitivity analysis show that ICER varied from $CAD23,045 to $CAD60,759/QALY (MoH perspective) and from $CAD23,120 to $CAD60,905/QALY (societal perspective). ATO in the first-line therapy for patients with APL can be considered a more cost-effective strategy than standard treatment from a Canadian perspective.

Economic evaluation of arsenic trioxide compared to all-trans retinoic acid + conventional chemotherapy for treatment of relapsed acute promyelocytic leukemia in Canada.

OBJECTIVES: Acute promyelocytic leukemia (APL) is an uncommon type of acute leukemia characterized by high early mortality. Current first-line treatments include all-trans retinoic acid (ATRA), anthracyclines, and other conventional chemotherapies (CTs). Although APL is generally associated with a good prognosis, about 20% of patients who achieve remission subsequently relapse and are resistant to the previously administrated treatment. The objective of this study was to assess, from a Canadian perspective, the economic impact of arsenic trioxide (ATO) compared to ATRA+CT for treatment of patients with relapsed/refractory APL. METHODS: The cost-effectiveness of ATO compared to ATRA+CT for treating patients with relapsed/refractory APL was assessed over a lifetime horizon using a Markov model. The model considers five health states: induction, second remission, treatment failure or relapse, postfailure, and death. Markov cycle length was 1 month for the first 24 months and 1 yr thereafter. The model also takes into account the incidence of grade 3-4 adverse events reported in clinical trials. Analyses were conducted from a Canadian Ministry of Health (MoH) and a societal perspective. RESULTS: Compared to ATRA+CT, ATO was associated with incremental cost-effectiveness ratios of $ 20,551/quality-adjusted life year (QALY) from a MoH perspective and $ 22,219/QALY from a societal perspective. Results of the probabilistic sensitivity analysis indicated that ATO is a cost-effective strategy in 99.27% and 98.98% of the simulations from a MoH and a societal perspective, respectively. CONCLUSIONS: This economic evaluation demonstrates that ATO is a cost-effective strategy compared to ATRA+CT for treatment of patients with relapsed/refractory APL in Canada.

The Economic Impact of Treatment Sequencing in Chronic Lymphocytic Leukemia in Canada Using Venetoclax plus Obinutuzumab.

BACKGROUND: Bruton tyrosine kinase inhibitors (BTKis) represent an advancement in chronic lymphocytic leukemia; however, these agents are administered continuously until disease progression or unacceptable toxicity, raising concerns about their affordability. Venetoclax in combination with obinutuzumab (VO) is a fixed-duration (12-month) treatment, approved in Canada in 2020. This study estimated the total cumulative cost of different treatment sequences and evaluated the economic impact of introducing treatment sequences with/without VO, from a Canadian health care system perspective. METHODS: A 10-year partitioned survival model was developed, considering key clinical parameters and direct medical costs. Results were stratified by TP53 aberration. RESULTS: Treatment sequences starting with first-line (1L) VO resulted in lower 10-year cumulative costs compared to sequences starting with BTKis administered until disease progression, across both TP53 aberration subgroups. With a maximum of three lines of treatment over a 10-year period, cumulative costs were largely determined by the first two lines of treatment. When comparing sequences with the same 1L treatment, sequences with BTKis in second-line incurred greater costs compared to fixed-duration regimens. CONCLUSIONS: Overall, the economic impact of treating all patients with VO led to 10-year cumulative savings of CAD 169,341 and CAD 293,731 per patient, without and with TP53 aberration, respectively. These savings are mainly due to reductions in treatment costs associated with fixed treatment duration.

Early access for innovative oncology medicines: a different story in each nation.

BACKGROUND: As innovative oncology medicines are rapidly developed, there is increasing pressure on payers to offer patients timely access to life-saving therapies. The uncertainty surrounding these therapies when phase III clinical trials are pending has necessitated new, adapted pathways to market access, with timelines that greatly vary by country. Understanding differences between pathways may identify opportunities to expedite patient access universally. OBJECTIVES: To describe early access pathways for new oncology medicines among selected countries with established health technology assessment (HTA) frameworks and publicly funded health systems, with a special focus on real-world evidence (RWE). METHODS: We reviewed the HTA agency websites of the selected OECD countries: National Institute for Health and Care Excellence (NICE) for England and Wales; Haute Autorité de Santé (HAS) for France; IQWiG and G-BA for Germany; Agenzia Italiana del Farmaco (AIFA) for Italy; Pharmaceutical Benefits Advisory Committee (PBAC) for Australia; and CADTH and Institut National d'Excellence en Santé et Services Sociaux (INESSS) for Canada as the primary source of evidence. RESULTS: Processes for early patient access to innovative oncology therapies varied across selected countries; however, most countries have an established pathway for publicly funded early access (England and Wales, France, Germany, Italy, and Australia). The utilization of RWE to support earlier access (coverage with evidence) also varied by country, with some HTA organizations being actively engaged in these agreements (NICE, AIFA, and HAS) and others having no established processes in place (G-BA and CADTH/INESSS). CONCLUSIONS: This review of early access pathways for novel oncology medicines found substantial variability between countries of interest. Coverage with evidence frameworks may provide a unique opportunity for industry and payers to collaborate on earlier access to innovative cancer therapies with life-saving potential.

A Probabilistic Cost-Effectiveness Analysis of Venetoclax and Obinutuzumab as a First-Line Therapy in Chronic Lymphocytic Leukemia in Canada.

BACKGROUND: Venetoclax is a first-in-class targeted therapy option that is an inducer of apoptosis in chronic lymphocytic leukemia (CLL) cells. The open-label phase III CLL14 clinical trial showed that venetoclax combined with obinutuzumab (VEN+O) is superior to obinutuzumab combined with chlorambucil in newly diagnosed patients with CLL. The aim of this study was to assess the health economic value of VEN+O for the frontline treatment of CLL in Canada from a publicly funded healthcare system perspective. METHODS: A partitioned survival analyses model was developed including three health states: progression free, progressed, and death. A cycle length of 28 days and a time horizon of 10 years was assumed. VEN+O treatment for a fixed duration of 12 months was compared to obinutuzumab combined with chlorambucil, fludarabine plus cyclophosphamide plus rituximab, bendamustine plus rituximab, chlorambucil plus rituximab, ibrutinib, and acalabrutinib. The population in the model included both unfit and overall frontline CLL patients, two subgroups were also assessed (patients with del17p/TP53 mutations and patients without del17p/TP53 mutations). Survival data extrapolated from the CLL14 trial were used to populate the model. Uncertainty was assessed via one-way sensitivity analyses, probabilistic analyses, and scenario analyses. RESULTS: Based on the probabilistic analyses, unfit frontline CLL patients receiving VEN+O were estimated to incur costs of Canadian dollars ($) 217,727 [confidence interval (CI) $170,725, $300,761] (del17p/TP53: $209,102 [CI $159,698, $386,190], non-del17p/TP53: $217,732 [CI $171,232, $299,063]) and accrue 4.96 [CI 4.04, 5.82] quality-adjusted life-years (del17p/TP53: 3.11 [CI 2.00, 4.20], non-del17p/TP53: 5.04 [CI 4.05, 5.92]). Obinutuzumab combined with chlorambucil, bendamustine plus rituximab, chlorambucil plus rituximab, and ibrutinib accrued lower quality-adjusted life-years and higher costs and as such, VEN+O was the dominant treatment option. The full incremental analysis showed that acalabrutinib was more expensive and more efficacious compared with VEN+O with an incremental-cost-effectiveness-ratio of $2,139,180/quality-adjusted life-year versus VEN+O and not a cost-effective option in Canada. Probabilistic analyses show that at a willingness to pay of $50,000/quality-adjusted life-year gained, VEN+O has the greatest probability of being cost effective. CONCLUSIONS: VEN+O is a cost-effective treatment option for unfit frontline CLL patients and provides value for money to healthcare payers.

Scl regulates the quiescence and the long-term competence of hematopoietic stem cells.

The majority of long-term reconstituting hematopoietic stem cells (LT-HSCs) in the adult is in G(0), whereas a large proportion of progenitors are more cycling. We show here that the SCL/TAL1 transcription factor is highly expressed in LT-HSCs compared with short-term reconstituting HSCs and progenitors and that SCL negatively regulates the G(0)-G(1) transit of LT-HSCs. Furthermore, when SCL protein levels are decreased by gene targeting or by RNA interference, the reconstitution potential of HSCs is impaired in several transplantation assays. First, the mean stem cell activity of HSCs transplanted at approximately 1 competitive repopulating unit was 2-fold decreased when Scl gene dosage was decreased. Second, Scl(+/-) HSCs were at a marked competitive disadvantage with Scl(+/+) cells when transplanted at 4 competitive repopulating units equivalent. Third, reconstitution of the stem cell pool by adult HSCs expressing Scl-directed shRNAs was decreased compared with controls. At the molecular level, we found that SCL occupies the Cdkn1a and Id1 loci in primary hematopoietic cells and that the expression levels of these 2 regulators of HSC cell cycle and long-term functions are sensitive to Scl gene dosage. Together, our observations suggest that SCL impedes G(0)-G(1) transition in HSCs and regulates their long-term competence.

Quality of Life in Wilson's Disease: A Systematic Literature Review.

Background: Wilson's disease (WD) is a rare inherited genetic disorder characterized by the progressive accumulation of copper in the brain, liver, and other major organ systems. To date, there have been no comprehensive studies synthesizing evidence pertaining to the quality of life (QOL) in WD. Objective: We conducted a systematic literature review to identify and synthesize the evidence on QOL in patients with WD. Methods: To address this gap in the literature, we conducted a systematic literature review in MEDLINE and EMBASE to identify observational studies and clinical trials reporting QOL outcomes among people living with WD. Results: A total of 442 publications were identified, 41 publications were eligible for full-text screening, and 7 articles, representing 7 studies, met all inclusion criteria. QOL questionnaires used across studies included the 12-Item Short Form Health Survey Questionnaire (version 1) (SF-12) (n=2), the 36-Item Short Form Health Survey Questionnaire (version 1) (SF-36) (n=3), Global Assessment Scale (GAS) (n=1), and World Health Organization QOL brief questionnaire (WHO-QOL-BREF) (n=1). Overall, the pattern in QOL from most studies demonstrated a worse QOL in WD patients compared with the general population, a deterioration in QOL for patients presenting with neurologic symptoms, and more frequent psychiatric symptoms compared with the ones with hepatic symptoms. Discussion: Although our understanding of the underlying pathophysiology of WD has advanced, and novel therapeutics are on the horizon, our understanding of how WD affects overall QOL remains limited. Evidence from this review demonstrates the substantial heterogeneity in reporting outcomes pertaining to the QOL associated with WD. These differences may be attributable to the fact that QOL is not typically assessed and the lack of a standardized method for assessing QOL in WD. Conclusion: This review demonstrates a need for more up-to-date studies with larger sample sizes to further evaluate QOL in patients with WD. The study also demonstrates the need for a WD-specific instrument to measure the QOL in WD patients.

Can Standard Health Technology Assessment Approaches Help Guide the Price of Orphan Drugs in Canada? A Review of Submissions to the Canadian Agency for Drugs and Technologies in Health Common Drug Review.

Orphan drugs have high acquisition costs and when standard health technology assessment (HTA) approaches are used to assess their cost-effectiveness, they often appear not cost-effective. The Canadian Patented Medicine Review Board (PMPRB), through new regulations, will apply HTA assessment results from the Canadian Agency for Drugs and Technologies in Health (CADTH) and Institut national d'excellence en santé et en services sociaux (INESSS) when setting the maximum price that can be charged for Category I patented medicines (treatments with an annual cost exceeding 150% of GDP per capita of Canada or with expected annual market size >$50M). Through these regulations, PMPRB has also established a willingness-to-pay threshold of CAD$200,000 or CAD$150,000 per quality adjusted life year (QALY) for medications with a prevalence of no more than 1 in 2000 across all approved indications. We reviewed the orphan drug submissions made to CADTH's Common Drug Review (CDR) January 2015-May 2020 to understand how the methodology of assessing cost-effectiveness of orphan drugs has guided pricing in Canada. A total of 35 orphan drug submissions were assessed by CDR in this period, none of which met the willingness-to-pay threshold of CAD$50,000 per QALY. Only one drug met the CAD$200,000 per QALY for Therapeutic Criteria Level I, and two drugs met CAD$150,000 per QALY for other Therapeutic Criteria Levels proposed by PMPRB. Price reductions of 32-99% were recommended for treatments that were approved in order to be listed for reimbursement. This review showed that the new PMPRB regulations could be creating challenges for manufacturers of rare disease treatments to meet Canadian pricing regulations. These regulations may jeopardize the launch of new medicines and limit opportunities to add to the development of real-world evidence of orphan drugs, which can be used in reimbursement approaches such as pay-for-performance.

Reslizumab and mepolizumab for moderate-to-severe poorly controlled asthma: an indirect comparison meta-analysis.

Aim: A systematic literature review and network meta-analysis assessed the efficacy and safety of reslizumab 3.0 mg/kg and mepolizumab 100 mg. Materials & methods: Eligible studies evaluated reslizumab and mepolizumab in patients with inadequately-controlled severe eosinophilic asthma. Using a Bayesian network meta-analysis, 95% credible intervals and posterior probabilities were reported. Results: Of 19 indirect efficacy comparisons performed in base-case (Global Initiative for Asthma 4/5 patients with ≥2 exacerbations in the previous year) and overall populations, significant differences favoring reslizumab were observed for severe exacerbations, FEV1 at 4 weeks and eosinophil counts at 4, 16 and 24 weeks, with no other significant differences including risk of adverse events. Conclusion: Indirect comparison of reslizumab and mepolizumab largely showed no significant differences in efficacy or safety.

Regulation of brain endothelial cells migration and angiogenesis by P-glycoprotein/caveolin-1 interaction.

We have investigated the involvement of P-glycoprotein (P-gp)/caveolin-1 interaction in the regulation of brain endothelial cells (EC) migration and tubulogenesis. P-gp overexpression in MDCK-MDR cells was correlated with enhanced cell migration whereas treatment with P-gp inhibitors CsA or PSC833 reduced it. Transfection of RBE4 rat brain endothelial cells with mutated versions of MDR1, in the caveolin-1 interaction motif, decreased the interaction between P-gp and caveolin-1, enhanced P-gp transport activity and cell migration. Moreover, down-regulation of caveolin-1 in RBE4 cells by siRNA against caveolin-1 stimulated cell migration. Interestingly, the inhibition of P-gp/caveolin-1 interaction increased also EC tubulogenesis. Furthermore, decrease of P-gp expression by siRNA inhibited EC tubulogenesis. These data indicate that the level of P-gp/caveolin-1 interaction can modulate brain endothelial angiogenesis and P-gp dependent cell migration.

Multidrug-resistant cancer cells contain two populations of P-glycoprotein with differently stimulated P-gp ATPase activities: evidence from atomic force microscopy and biochemical analysis.

Considerable interest exists about the localization of P-gp (P-glycoprotein) in DRMs (detergent-resistant membranes) of multidrug resistant cancer cells, in particular concerning the potential modulating role of the closely related lipids and proteins on P-gp activity. Our observation of the opposite effect of verapamil on P-gp ATPase activity from DRM and solubilized-membrane fractions of CEM-resistant leukaemia cells, and results from Langmuir experiments on membrane monolayers from resistant CEM cells, strongly suggest that two functional populations of P-gp exist. The first is located in DRM regions: it displays its optimal P-gp ATPase activity, which is almost completely inhibited by orthovanadate and activated by verapamil. The second is located elsewhere in the membrane; it displays a lower P-gp ATPase activity that is less sensitive to orthovanadate and is inhibited by verapamil. A 40% cholesterol depletion of DRM caused the loss of 52% of the P-gp ATPase activity. Cholesterol repletion allowed recovery of the initial P-gp ATPase activity. In contrast, in the solubilized-membrane-containing fractions, cholesterol depletion and repletion had no effect on the P-gp ATPase activity whereas up to 100% saturation with cholesterol induced a 58% increased P-gp ATPase activity, while no significant modification was observed for the DRM-enriched fraction. DRMs were analysed by atomic force microscopy: 40-60% cholesterol depletion was necessary to remove P-gp from DRMs. In conclusion, P-gp in DRMs appears to contain closely surrounding cholesterol that can stimulate P-gp ATPase activity to its optimal value, whereas cholesterol in the second population seems deprived of this function.

Major cytogenetic aberrations and typical multidrug resistance phenotype of uveal melanoma: current views and new therapeutic prospects.

Uveal melanoma is the most frequent intra-ocular cancer. The recent development of new chromosome-related technologies have permitted the elucidation of both the cytogenetics and the natural history of this disease. Fifty to 60% of uveal melanomas are linked to a monosomy 3, which appears as an early and determinant event in tumor progression. Tumors with this anomaly have a very poor prognosis. Recent work suggests that this category of uveal melanoma represents a distinct pathologic entity from that associated with normal disomy 3. Chromosome 6 aberrations probably constitute a second entry point into the process of cancerogenesis, while gains in 8q seem to appear later in the natural history of uveal melanomas due to their higher frequency in larger tumors. Other anomalies will be reviewed. In spite of significant improvements in the local treatment of uveal melanoma, many patients die due to tumor metastasis. This disease is characterized by a constitutive chemoresistance whose typical multidrug resistance phenotype (MDR) is particularly complex since different combinations of several resistance proteins are simultaneously produced. Regulation of the expression of these proteins is a research priority, increasingly so as gene therapy-dependent chemosensitization strategies expand. Therefore, the development and improvement of methods to determine the chemoresistance profile become a crucial objective today in the therapeutic strategies against uveal melanoma.

Gene therapy of the typical multidrug resistance phenotype of cancers: a new hope?

The multidrug resistance (MDR) phenotype of cancers has generated a large amount of research, owing to its constant fatal clinical outcome. Many studies have focused on the discovery of chemomodulators; however, in spite of this huge effort, the side effects that these products induce, and their additive toxicity when used in the presence of anticancer drugs, have led to the disaffection of the pharmaceutical industry and possibly slowed down research in pharmacological modulation. New tools developed using molecular biology techniques have opened the way for gene therapy and given birth to new therapeutic hopes. However, these discoveries and especially their clinical applications have slowed due to a lack of knowledge of the systems that finely regulate the MDR genes. This weakness explains why, to date, no general review has focused on the possibilities of gene therapy of MDR derived form the strategic options now available. Based on molecular foundations and recent fundamental discoveries, we seek to inform clinicians of the therapeutic hopes for chemoresistant tumors brought about by potent and specific new tools such as transcriptional decoys, interfering RNAs, etc. After describing the causes and mechanisms of MDR, we critically review these new strategies and their corresponding clinical trials.

Characterization of the typical multidrug resistance profile in human uveal melanoma cell lines and in mouse liver metastasis derivatives.

Uveal melanoma is the most common intraocular malignancy. To study its biology, stable cell lines provide a useful tool, but these are very difficult to obtain. A stable and rapidly growing human choroidal melanoma cell line composed of pure epithelioid cells was established and maintained for at least 4 years. In vivo transplantation into BALB/cByJ nude mice induced vascularized tumours at the injection sites. Interestingly, two of three cases produced a liver metastasis. Other uveal melanoma cell lines displaying different morphological aspects were also obtained. To avoid the bias due to uncertain immunologically based staining approaches, several methods were juxtaposed to establish the multidrug resistance (MDR) profile. All the uveal melanomas studied expressed significant levels of the MDR-related MDR1, MRP1 (MDR-related protein 1) and LRP/MVP (lung resistance protein/major vault protein) messenger RNAs (mRNAs), produced their corresponding proteins and were able to functionally extrude daunomycin. When compared with the established MEWO skin melanoma cell line, our data showed that both primary and metastatic uveal melanomas intrinsically expressed the typical MDR phenotype, which precludes the use of any anticancer drugs known to be substrates of MDR-related proteins to treat the disease. Moreover, it appears that the metastasizing process does not change the status of the MDR phenotype.

Leucine-rich protein 130 contributes to apoptosis resistance of human hepatocarcinoma cells.

LRP130 is a ubiquitous protein involved in cellular homeostasis, microtubule alteration, and transactivation of a few multidrug resistance genes. Its role in resistance to apoptosis in HepG2 and HUH7 hepatocarcinoma cells was investigated. Using shRNA-producing lentiviruses to down-regulate the LRP130 gene, we showed that i) LRP130 did not affect the capacity of hepatocarcinoma cells to extrude drugs since LRP130 down-regulation was insufficient to significantly reduce P-glycoprotein production in these cells, and ii) the expression of 11 apoptosis-related genes measured by PCR-array was significantly reduced. Interestingly, six of these genes encode extrinsic pathway proapoptotic proteins whose expression was higher in LRP130-non producing than in LRP130-producing HepG2 cells. Fluorescence microscopy confirmed this new anti-apoptotic role of LRP130, which is strengthened by a significantly reduced cytochrome c oxidase activity in LRP130-down-regulated hepatocarcinoma cells.

Modulation of p-glycoprotein function by caveolin-1 phosphorylation.

p-glycoprotein (p-gp) is an ATP-binding cassette transporter and its overexpression is responsible for the acquisition of the multidrug resistance phenotype in human tumors. p-gp is localized at the blood-brain barrier and is involved in brain cytoprotection. Our previous work used immunoprecipitation to show that caveolin-1 can interact with p-gp. In this study, we provide evidence that caveolin-1 regulates p-gp transport activity in a rat brain endothelial cell line (RBE4). Down-regulation of caveolin-1 by siRNA reduced the interaction between p-gp and caveolin-1, followed by a decrease in [3H]-Taxol and [3H]-Vinblastine accumulation in RBE4 cells. The latter result showed that down-regulation of caveolin-1 enhanced p-gp transport activity. RBE4 cells were also transfected with Sarcoma in order to modulate caveolin-1 phosphorylation. Overexpression of Sarcoma, a protein tyrosine kinase, stimulated caveolin-1 phosphorylation and increased both [3H]-Taxol and [3H]-Vinblastine accumulation as well as Hoechst 33342 accumulation. Transfection of caveolin-1 inhibits p-gp transport activity. Conversely, transfection of the mutant cavY14F decreased the p-gp/caveolin-1 interaction and reduced accumulation of the two p-gp substrates. Thus, our data show that caveolin-1 regulates p-gp function through the phosphorylation state of caveolin-1 in endothelial cells from the blood-brain barrier.

Modulation of P-glycoprotein function by sphingosine kinase-1 in brain endothelial cells.

P-glycoprotein (P-gp), an ABC-transporter highly expressed in brain capillaries, protects the brain by extruding xenobiotics. However, its overexpression has also been associated with the multidrug resistance phenotype in tumors. Here, we have investigated the regulation of P-gp transport activity by sphingosine kinase 1 (SphK-1) in brain endothelial cells. We first demonstrated that SphK-1 is overexpressed in endothelial cells (EC) isolated from rat brain tumors compared with EC from normal brain. We also provide evidence that the overexpression of SphK-1 in the cerebral EC line RBE4 leads to the up-regulation of P-gp, both at the gene and protein levels, and that this modulation depends on the catalytic activity of SphK-1. Moreover, we determined the effect of sphingosine-1-phosphate (S1P), the product of SphK-1, on P-gp function. S1P strongly stimulates P-gp transport activity, without modulating its expression. Finally, we found that the S1P-mediated stimulation of P-gp activity is mediated by S1P-1 and S1P-3 receptors at the RBE4 cell surface. Altogether, these results indicate that SphK-1 and its product S1P are involved in the control of P-gp activity in RBE4 cells. Since SphK-1 is overexpressed in EC from brain tumors, these data also suggest that this kinase and its product could contribute to the acquisition and the maintenance of the multidrug resistance phenotype in brain tumor-derived endothelial cells.

Control of P-glycoprotein activity by membrane cholesterol amounts and their relation to multidrug resistance in human CEM leukemia cells.

P-glycoprotein (P-gp) is the most well-known ATP-binding cassette (ABC) transporter involved in unidirectional substrate translocation across the membrane lipid bilayer, thereby causing the typical multidrug resistance (MDR) phenotype expressed in many cancers. We observed that in human CEM acute lymphoblastic leukemia cells expressing various degrees of chemoresistance and where P-gp was the sole MDR-related ABC transporter detected, the amount of esterified cholesterol increased linearly with the level of resistance to vinblastine while the amounts of total and free cholesterol increased in a nonlinear way. Membrane cholesterol controlled the ATPase activity of P-gp in a linear manner, whereas the P-gp-induced daunomycin efflux decreased nonlinearly with the depletion of membrane cholesterol. All these elements suggest that cholesterol controls both the ATPase and the drug efflux activities of P-gp. In addition, in CEM cell lines that expressed increasing levels of elevated chemoresistance, the amount of P-gp increases to a plateau value of 40% of the total membrane proteins and remained unvaried while the amount of membrane cholesterol increased with the elevation of the MDR level, strongly suggesting that cholesterol may be directly involved in the typical MDR phenotype. Finally, we showed that the decreased daunomycin efflux by P-gp due to the partial depletion of membrane cholesterol was responsible for the efficient chemosensitization of resistant CEM cells, which could be totally reversed after cholesterol repletion.