Understanding and circumventing resistance to anticancer monoclonal antibodies

With the widespread use of therapeutic monoclonal antibodies in the treatment of patients with cancer, resistance to these agents has become a major issue. Preclinical models of drug action or resistance have contributed to unravel the main mechanisms of resistance, involving both tumor-associated and host related factors. However our understanding of how a monoclonal antibody destroys cancer cells in a patient and why it one day stops being effective are still far from being complete. This review focuses on the available data on mechanisms of action and resistance to rituximab, and includes some additional information for other monoclonal antibodies. Innovative approaches designed to overcome resistance, such as combination immunotherapy, costimulation with cytokines or growth factors are presented.     

Understanding and circumventing resistance to anticancer monoclonal antibodies

With the widespread use of therapeutic monoclonal antibodies in the treatment of patients with cancer, resistance to these agents has become a major issue. Preclinical models of drug action or resistance have contributed to unravel the main mechanisms of resistance, involving both tumor-associated and host related factors. However our understanding of how a monoclonal antibody destroys cancer cells in a patient and why it one day stops being effective are still far from being complete. This review focuses on the available data on mechanisms of action and resistance to rituximab and includes some additional information for other monoclonal antibodies. Innovative approaches designed to overcome resistance, such as combination immunotherapy, costimulation with cytokines or growth factors are presented.Key words: monoclonal antibodies, resistance, rituximab, cetuximab, trastuzumab     

Cytokine-mediated reversal of multidrug resistance

The occurrence of the multidrug resistance phenotype still represents a limiting factor for successful cancer chemotherapy. Numerous efforts have been made to develop strategies for reversal and/or modulation of this major therapy obstacle through targeting at different levels of intervention. The phenomenon of MDR is often associated with overexpression of resistance-associated genes. Since the classical type of MDR in human cancers is mainly mediated by the P-glycoprotein encoded by the multidrug resistance gene 1, mdr1, the majority of reversal approaches target the expression and/or function of the mdr1 gene/P-glycoprotein. Due to the fact that the multidrug phenotype always represents the net effect of a panel of resistance-associated genes/gene products, other resistance genes, e.g. those encoding the multidrug resistance-associated protein MRP or the lung resistance protein LRP, were included in the studies. Cytokines such as tumor necrosis factor α and interleukin-2 have been shown to modulate the MDR phenotype in different experimental settings in vitro and in vivo. Several studies have been performed to evaluate their potential as chemosensitizers of tumor cells in the context of a combined application of MDR-associated anticancer drugs like doxorubicin and vincristine with cytokines. Moreover, the capability of cytokines to modulate the expression of MDR-associated genes was demonstrated, either by external addition or by transduction of the respective cytokine gene. Knowledge of the combination effects of cytokines and cytostatics and its link to their MDR-modulating capacity may contribute to a more efficient and to a more individualized immuno-chemotherapy of human malignancies. This revised version was published online in August 2006 with corrections to the Cover Date.     

Pharmacologic circumvention of multidrug resistance

The ability of malignant cells to develop resistance to chemotherapeutic drugs is a major obstacle to the successful treatment of clinical tumors. The phenomenon multidrug resistance (MDR) in cancer cells results in cross-resistance to a broad range of structurally diverse antineoplastic agents, due to outward efflux of cytotoxic substrates by themdr1 gene product, P-glycoprotein (P-gp). Numerous pharmacologic agents have been identified which inhibit the efflux pump and modulate MDR. The biochemical, cellular and clinical pharmacology of agents used to circumvent MDR is analyzed in terms of their mechanism of action and potential clinical utility. MDR antagonists, termed chemosensitizers, may be grouped into several classes, and include calcium channel blockers, calmodulin antagonists, anthracycline andVinca alkaloid analogs, cyclosporines, dipyridamole, and other hydrophobic, cationic compounds. Structural features important for chemosensitizer activity have been identified, and a model for the interaction of these drugs with P-gp is proposed. Other possible cellular targets for the reversal of MDR are also discussed, such as protein kinase C. Strategies for the clinical modulation of MDR and trials combining chemosensitizers with chemotherapeutic drugs in humans are reviewed. Several novel approaches for the modulation of MDR are examined.Abbreviations ALL acute lymphocytic leukemia - AML acute myelogenous leukemia - CaM calmodulin - CsA cyclosporin A - MDR multidrug resistance - P-gp P-glycoprotein - PMA phorbol 12-myristate 13-acetate - PKC protein kinase C     

Euphorbia and Momordica metabolites for overcoming multidrug resistance

Multidrug resistance (MDR) is the major obstacle for cancer chemotherapy. MDR is a multifactorial phenomenon that can result from several mechanisms, including an increased drug efflux, due to overexpression of P-glycoprotein (P-gp) that transports anticancer drugs out of the cells. Thus, the role of this transporter has made it a therapeutic target and the development of P-gp modulators considered among the most realistic approaches for overcoming P-gp-mediated MDR. Many other strategies have been proposed. One of them is the identification of compounds that selectively kill multidrug resistant cells. In our search for MDR modulators from plants, the P-gp inhibition ability of a large number of compounds on resistant cancer cells was evaluated. These compounds, presented in this review, comprise mainly diterpenes, triterpenes and phenolic derivatives. The most relevant results were obtained from two sets of compounds: macrocyclic diterpenes with the jatrophane and lathyrane scaffold, and triterpenes of the cucurbitane-type isolated from Euphorbia species and Momordica balsamina L., respectively. Additionally, some of those macrocyclic diterpenes, and ent-abietane diterpenic lactones, also isolated from Euphorbia species, were found to be selectively toxic to drug-resistant phenotypes.     

Protein kinases and multidrug resistance

The role of protein kinases in the multidrug resistance phenotype of cancer cell lines is discussed with an emphasis on protein kinase C and protein kinase A. Evidence that P-glycoprotein is phosphorylated by these kinases is summarised and the relationship between P-glycoprotein phosphorylation and the multidrug-resistant phenotype discussed. Results showing that protein kinase C, particularly the alpha subspecies, is overexpressed in many MDR cell lines are described: this common but by no means universal finding seems to be drug- and cell line-dependent and in only in a few cases is there a direct correlation between protein kinase C activity and multidrug resistance. From co-immunoprecipitation results it is suggested that P-glycoprotein is a specific protein kinase C receptor, as well as being a substrate. Revertant experiments provide conflicting results as to a direct relationship between expression of P-glycoprotein and protein kinase C. Evidence that protein kinase A influences P-glycoprotein expression at the gene level is well documented and the mechanisms by which this occurs are becoming clarified. Results on the relationship between protein kinase C and multidrug resistance using many inhibitors and phorbol esters are difficult to interpret because such compounds bind to P-glycoprotein. In spite of huge effort, a direct involvement of protein kinase C in regulating multidrug resistance has not yet been firmly established. However, evidence that PKC regulates a Pgp-independent mechanism of drug resistance is accumulating. This revised version was published online in August 2006 with corrections to the Cover Date.     

Molecular mechanisms of antibacterial multidrug resistance

Treatment of infections is compromised worldwide by the emergence of bacteria that are resistant to multiple antibiotics. Although classically attributed to chromosomal mutations, resistance is most commonly associated with extrachromosomal elements acquired from other bacteria in the environment. These include different types of mobile DNA segments, such as plasmids, transposons, and integrons. However, intrinsic mechanisms not commonly specified by mobile elements-such as efflux pumps that expel multiple kinds of antibiotics-are now recognized as major contributors to multidrug resistance in bacteria. Once established, multidrug-resistant organisms persist and spread worldwide, causing clinical failures in the treatment of infections and public health crises.     

Development of multidrug resistance type I Cmdr1 expression in chicken embryonic gonads

A primary role of P-glycoprotein (P-gp), encoded by the multidrug resistance type I gene, is to protect against naturally occurring xenotoxics. Recently, the preferential expression of chicken multidrug resistance type I (Cmdr1) was identified in the embryonic gonads during the early periods of development. Here we investigated the expression of Cmdr1 and P-gp in the gonads during embryogenesis, and compared to that in the ovarian follicles of domestic hens (Gallus gallus). As revealed by immunohistochemistry, P-gp was highly expressed in theca cells of mature follicles, whereas the expression was low in immature follicles. Immunohistochemical analysis showed that expression of Cmdr1-type P-gp was very low in embryonic gonads. Cmdr1 mRNA was undetectable in the gonads of 5-day embryos (E5) by RT-PCR, whereas Cmdr1 mRNA was significantly detectable in the developing gonads at E9 and E21. In the testicular tissues, germ cells were distributed along developing seminiferous cords as identified by a specific marker gene, whereas Cmdr1-type P-gp positive cells were observed evenly on testicular tissues. Collectively, it is concluded that Cmdr1 expression is initiated in the chicken ovary and testis after sexual differentiation, but expression of Cmdr1-type P-gp is very low through embryogenesis.     

Radiopharmaceuticals for assessment of multidrug resistance P-glycoprotein-mediated drug transport activity

Multidrug resistance (MDR) mediated by overexpression of MDR1 P-glycoprotein (Pgp) is one of the best characterized transporter-mediated barriers to successful chemotherapy in cancer patients. Thus, noninvasive interrogation of Pgp-mediated transport activity in vivo would be beneficial in guiding therapeutic choices. Both small organic medicinals as well as metal complexes characterized as transport substrates for Pgp are amenable to incorporation of PET or SPECT radionuclides and may enable noninvasive imaging of Pgp in cancer patients. Toward this objective, clinically approved agents, exemplified by (99m)Tc-Sestamibi and (99m)Tetrofosmin, have already shown promise for the functional evaluation of Pgp-mediated transport activity in human tumors in vivo. In addition, selected agents from an upcoming class of substituted Schiff-base gallium(III) complexes containing an N(4)O(2) donor core in their organic scaffold and capable of generating both SPECT and PET radiopharmaceuticals have also been shown to be promising for noninvasive assessment of Pgp activity in vitro and in vivo.     

Design of liposomes for circumventing the reticuloendothelial cells.

Two different aspects of liposomal drug delivery to non-RES cells have been described. In one of the systems, by incorporating neutral glycolipids, with terminal beta-galactoside residue into liposomes, it is possible to target liposomes to the liver parenchymal cells, partially bypassing the RES. Asialoganglioside seems to be the most suited for this purpose. In another approach, various factors that prolong the lifespan of circulating liposomes have been discussed. It is possible to design such liposomes by imparting hydrophilicity to the liposomal surface. The effectiveness of a number of possible candidates, such as dextran, GM1 ganglioside and PEG, has been discussed in this context.     

Glyoxalase in tumourigenesis and multidrug resistance

Since the discovery by Warburg of high aerobic glycolysis in most tumours in the 1920s, it has remained unclear how to exploit this in chemotherapy. The aim of this review is to assess the evidence for the involvement of the glyoxalase system in tumour growth and multidrug resistance and the importance of the glyoxalase system as a target for anticancer drug development and a source of biomarkers for tumour diagnosis. Increased expression of glyoxalase 1 appears to support the viability of tumour cells with high glycolytic rates. Multidrug resistance conferred by overexpression of glyoxalase 1 suggests mechanisms of toxicity of most current antitumour agents involve, in some part, accumulation of methylglyoxal to cytotoxic levels. The recent finding of glyoxalase 1 gene amplification in tumours and induction of increased glyoxalase 1 expression by malignant transformation and conventional antitumour drug treatment implies a critical role of glyoxalase 1 in innate and acquired multidrug resistance in cancer treatment. Improved understanding of glyoxalase 1 in cancer chemotherapy multidrug resistance is likely vital to achieve improvement of cancer patient survival rates. Advances made to counter glyoxalase 1-linked multidrug resistance with glyoxalase 1 inhibitors and related prodrugs has been translated from in vitro to pre-clinical in vivo studies. Further research is required urgently for next stage clinical translation. Finally, overexpression of glyoxalase 1 may be linked to multidrug resistance in chemotherapy of other disease - such as microbial infections.     

Cell-derived microvesicles and antitumoral multidrug resistance

Antitumoral chemotherapeutic treatments are often impaired by innate or acquired multidrug resistance (MDR). After four decades of MDR research, having underlined its complexity, new knowledge about the mechanisms of tumor resistance to antineoplastic drugs is a prerequisite for improving chemotherapy. Following our observations with a non-pathogenic eukaryotic microorganism, Dictyostelium discoideum, I suggest that MDR in tumor cells might be the consequence of a detoxification mechanism, mediated by cell-derived microvesicles. Recently published observations with tumoral human cells support this hypothesis. First, these cell-derived vesicles might impair chemotherapeutic efficiency of many structurally-different antineoplastic agents by preventing them to reach their intracellular target, followed by their expulsion outside the tumor cells, as observed for Dictyostelium cells. Secondly, besides their newly recognized function of intercellular communication, the cell-derived vesicles might also act as intercellular transporters of multidrug resistance proteins. Experiments are suggested for checking the hypothesis of cell-derived vesicles mediating multidrug resistance.     

Phosphorylation of the multidrug resistance associated glycoprotein

Drug-resistant cell lines derived from the mouse macrophage-like cell line J774.2 express the multidrug resistance phenotype which includes the overexpression of a membrane glycoprotein (130-140 kilodaltons). Phosphorylation of this resistant-specific glycoprotein (P-glycoprotein) in intact cells and in cell-free membrane fractions has been studied. The phosphorylated glycoprotein can be immunoprecipitated by a rabbit polyclonal antibody specific for the glycoprotein. Phosphorylation studies done with partially purified membrane fractions derived from colchicine-resistant cells indicated that (a) phosphorylation of the glycoprotein in 1 mM MgCl2 was enhanced a minimum of 2-fold by 10 microM cAMP and (b) the purified catalytic subunit of the cAMP-dependent protein kinase (protein kinase A) phosphorylated partially purified glycoprotein that was not phosphorylated by [gamma-32P]ATP alone, suggesting that autophosphorylation was not involved. These results indicate that the glycoprotein is a phosphoprotein and that at least one of the kinases responsible for its phosphorylation is a membrane-associated protein kinase A. The state of phosphorylation of the glycoprotein, which is a major component of the multidrug resistance phenotype, may be related to the role of the glycoprotein in maintaining drug resistance.