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Environment International 39 (2012) 73–86

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Environment International
journal homepage: www.elsevier.com/locate/envint

Anticancer drugs in surface waters


What can we say about the occurrence and environmental significance of cytotoxic,
cytostatic and endocrine therapy drugs?
Jean-Philippe Besse a,⁎, Jean-François Latour b, 1, Jeanne Garric a, 2
a
Cemagref, UR Milieux Aquatiques Ecologie et Pollution (MAEP), Laboratoire d'écotoxicologie/Laboratoire d'analyses physico-chimiques des milieux aquatiques, 3 bis quai Chauveau,
CP 220, F-69226 Lyon, France
b
Pharmacie, Centre Léon Bérard, 28 rue Laennec 69008 Lyon, France

a r t i c l e i n f o a b s t r a c t

Article history: This study considers the implications and research needs arising from anticancer (also referred to as antineo-
Received 20 June 2011 plastic) drugs being released into the aquatic environment, for the entire therapeutic classes used: cytotoxic,
Accepted 6 October 2011 cytostatic and endocrine therapy drugs.
Available online 8 November 2011
A categorization approach, based on French consumption amounts, allowed to highlight parent molecules
and several metabolites on which further occurrence and ecotoxicological studies should be conducted.
Keywords:
Anticancer drugs
Investigations of consumption trends at a national and a local scale show an increase in the use of anticancer
Antineoplastic drugs between 2004 and 2008, thus leading to increased levels released in the environment. It therefore
Pharmaceuticals appears necessary to continue surveying their presence in surface waters and in wastewater treatment
Surface water plant (WWTP) effluents.
Ecotoxicity Furthermore, due to the rise of anticancer home treatments, most of the prescribed molecules are now available
Occurrence in town pharmacies. Consequently, hospital effluents are no longer the main expected entry route of anticancer
drugs into the aquatic environment.
Concerning ecotoxicological risks, current knowledge remains insufficient to support a definitive conclusion.
Risk posed by cytotoxic molecules is still not well documented and it is not possible to conclude on their
long-term effects on non-target organisms. To date, ecotoxicological effects have been assessed using standardized
or in vitro assays. Such tests however may not be suitable for anticancer drugs, and further work should focus on
full-life cycle or even multigenerational tests.
Environmental significance (i.e. occurrence and effects) of cytostatics (protein kinases inhibitors and monoclonal
antibodies), if any, is not documented. Protein kinases inhibitors, in particular, deserve further investigation due
to their universal mode of action.
Finally, concerning endocrine therapy drugs, molecules such as antiestrogen Tamoxifen and its active metabolites,
could be of concern.
Overall, to accurately assess the ecotoxicological risk of anticancer drugs, we discuss the need to break away from
tests on isolated molecules and to test effects of mixtures at the low ng.l− 1 range.
© 2011 Elsevier Ltd. All rights reserved.

1. Introduction negligible amounts and because they are likely to exert biological and/
or adverse effects on aquatic organisms, they can be considered as envi-
Since their first detection in the aquatic environment in the 1970s ronmental contaminants. Subsequently, there is a need to target phar-
(Hignite and Aznaroff, 1977), growing attention has been paid to phar- maceuticals with environmental significance, quantify them, and
maceuticals. As these compounds reach the environment in non- assess their ecotoxicological risk (Fent et al., 2006; Halling-Sørensen
et al., 1998; Kolpin et al., 2002; Ternes, 1998). Among pharmaceuticals,
drugs used for cancer treatment (referred to as anticancer or antineo-
plastic drugs) are suspected to represent a specific risk for aquatic
⁎ Corresponding author. Tel.: + 33 4 72 20 87 87, + 33 4 72 20 89 05; fax: + 33 4 78 non-target species (Kümmerer, 2001).
47 78 75. Currently, two main groups of drugs are used: cytotoxic drugs and
E-mail addresses: jean-philippe.besse@cemagref.fr (J.-P. Besse),
latour@lyon.fnclcc.fr (J.-F. Latour), jeanne.garric@cemagref.fr (J. Garric).
endocrine therapy drugs. Due to their intended function, the first ones
1
Tel.: + 33 4 78 78 28 28. could exert cytotoxic, genotoxic, mutagenic, carcinogenic or teratogenic
2
Tel.: + 33 4 72 20 87 87; fax: + 33 4 78 47 78 75. effects on aquatic species. Moreover, as they often act with structure

0160-4120/$ – see front matter © 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.envint.2011.10.002
74 J.-P. Besse et al. / Environment International 39 (2012) 73–86

Table 1
A classification of anticancer drugs (cytotoxic, cytostatic and hormonally active drugs).

ATC class Class Mechanism of action Detailed mechanism

Antineoplastic Cytotoxics which interact Alkylating agents Attach one or more nucleophile groups to the DNA
agents (ATC L01) directly with DNA and inhibit or alter its transcription
Platinum complexes Inhibit DNA replication by binding to it
Intercalating agents Act by breaking single-stranded DNA
Cytotoxics which interact Antimetabolites Structural analogs of purine, pyrimidine or folic acid,
indirectly with DNA act by blocking enzyme activity and disrupting DNA synthesis
Cytotoxic antibiotics Intercalate between DNA base pairs and thus disturb the synthesis
and/or function of nucleic acids
Mitotic spindle inhibitors Halt chromosome segregation by inhibiting mitotic spindle formation
Topoisomerase inhibitors Induce or stabilize DNA damage by blocking religation of double stranded DNA breaks
Cytostatics Protein kinase inhibitors Interact with protein kinases, involved in the regulation of many
biological processes (cell growth, migration,survival..),
most of the compounds used are Tyrosine kinase inhibitors
Monoclonal antibodies Block tumoral cells extracellular receptors
Endocrine therapy Hormones and related GnRH and LhRH analogs Interfere with gonadotropin release
(ATC L02) Anti-hormones and related Antiestrogens Direct estrogen receptor antagonism
Antiaromatase Inhibit the estrogen biosynthesis
Antiandrogens Inhibit the binding of testosterone to its receptor

and function of DNA, some authors consider that all eukaryote organisms structure and mechanism of action: cytotoxics, strictly speaking, and
might be susceptible to their toxicity (Johnson et al., 2008). cytostatics.
Drugs used in endocrine therapy, due to their specific hormonal or Cytotoxic drugs are compounds that interact with DNA. They
anti-hormonal properties can be considered as endocrine disruptors cause metabolic and morphological alterations of the cell, leading to
and therefore require consideration. its death, through different mechanisms of action (Table 1); they
Overall, considering their distinctive mechanisms of action, anti- are reported to exert genotoxic, mutagenic and carcinogenic effects.
cancer drugs should be screened and assessed for environmental Cytostatic agents display mechanisms of action somewhat different,
risk, according to the EMEA guidelines (EMEA, 2006). However, as leading to a decrease in tumoral cell proliferation.
the EMEA guidelines only require environmental assessment for Cytostatics do not interact with DNA and can achieve their effects
newly authorized pharmaceuticals, most of the anticancer drugs are through various mechanisms, such as blocking cell growth factors, or
not submitted to such a procedure. indirectly by recruiting cytotoxic cells, such as macrophages and
Some authors have questioned the environmental occurrence and monocytes, which will contribute to the tumoral cell death. Cytostatics
risk of cytotoxic (Henschel et al., 1997; Johnson et al., 2008; Kümmerer include monoclonal antibodies (ATC class L01XC) and anti-protein
and Al-Ahmad, 1997; Straub, 2010; Zounkova et al., 2010), and endo- kinases (ATC class L01XE); most of the protein kinase inhibitors
crine therapy drugs (Runnalls et al., 2010). Others have modeled their used are tyrosine kinase inhibitors (Table 1).
release in hospital effluents (Mullot, 2009; Mullot et al., 2010). Never- Aside from these two classes, other molecules are used for treatment
theless, it appears that only a few drugs have been assessed and that of hormone-dependant cancers. They are referred to as endocrine
no focused effort has been made to ascertain which molecules are likely therapy drugs, and are classed into two groups: hormone analogs and
to pose a risk to the aquatic environment. related agents (L02A), and hormone antagonists and related agents
This paper therefore aims at achieving three goals: (L02B).
Hormone analogs used in cancer treatment are molecules that
• providing an overview of the occurrence of anticancer drugs in
interfere with gonadotropin release. These molecules are either
the aquatic environment by calculating predicted environmental
GnRH analogs or LhRH analogs. 3 Hormone antagonists include several
concentrations (PECs), based on French consumption data;
non-steroidal antiestrogens and antiandrogens: antiestrogens act either
• conducting a preliminary risk characterization for these molecules
by inhibiting the estrogen biosynthesis (antiaromatase compounds) or
and give hints for further ecotoxicological testing;
by direct estrogen receptor antagonism (Table 1). Antiandrogens are
• investigating consumption trends.
compounds with a non-steroidal structure that inhibit the binding of
In this study, only the risk for surface waters is discussed. Risk testosterone to its receptor (Table 1).
related to drinking waters has been discussed by other authors For convenience, the term anticancer drug will be used throughout
(Johnson et al., 2008; Rowney et al., 2009); risk associated with this document when accounting for the all types of molecules
sludge and sediment is not well documented, however some existing described above (cytotoxics, cytostatics and endocrine therapy).
data suggest that hydrophobic anticancers and platinum derivatives
may partition on particles (Lenz et al., 2005; Thomas et al., 2007);
finally, risk related to groundwater, if any, has not yet been documented. 3. Materials and methods

2. Classification of anticancer drugs, cytotoxic, cytostatic and To gather and investigate pharmacological, metabolism and
endocrine therapy drugs ecotoxicological data for anticancer compounds, scientific literature
was reviewed as along with the following databases and books: the
In studies conducted on occurrence and effects of anticancer drugs Banque Claude Bernard (BCB, 2011), the Micromedex Drugdex®
in the environment, the terms cytotoxic or cytostatic are both used databank (Micromedex Drugdex©, 2011), the IARC database
without discrimination to account for anticancer drugs in general. (www.iarc.fr), and the BC cancer agency database (www.bccancer.
However, cytotoxic and cytostatic agents can differ greatly in their
mechanism of action. 3
Some progestagens, namely progesterone and medroxyprogesterone, can be used
Cytotoxic drugs belong to the L01 class of the Anatomical Therapeutic in the treatment of some cancers. For these molecules, expected concentrations and ef-
Chemical (ATC) classification (www.whocc.no/atcddd). Cytotoxic agents fects on the aquatic environment have been discussed elsewhere (Besse and Garric,
can be classified in two main groups, on the basis of their chemical 2009).
J.-P. Besse et al. / Environment International 39 (2012) 73–86 75

bc.ca), and the Drug bank database (www.drugbank.ca). Gathered 100%; 12 molecules have excretion rate values equal to or lower than 10%, while 14
molecules have values greater than or equal to 50%. PECs refined by metabolism are
data on metabolism are provided in Appendix A.
given in Table 2.
Preliminary exposure assessment was implemented by calculating A recent study (Rowney et al., 2009) calculated PEC values for 10 cytotoxic molecules
PECs, as described by Besse et al. (2008), using the following Eq. (1): in WWTP influent and effluent in England. If converted to values for WWTP effluent (i.e.
without the dilution factor of 10), our refined PEC are close to those calculated in this
amount  Fexcreta  Fstp study, except for Fluorouracil, for which our PEC is one order of magnitude higher. Such
PEC ¼ ð1Þ a discrepancy is notably linked to the fact that this molecule is highly removed in
WWinhab  hab  Dilution  365
WWTPs (Mahnik et al., 2004; Rowney et al., 2009; Straub, 2010), which was not taken
into account in our calculation.
PECs are expressed in mg.l − 1 using the following parameters: Only a few cytotoxics have been searched and/or detected in the aquatic environment,
consumption is the quantity (mg.year − 1) of an active molecule and concentrations are generally very low, in the ng.l− 1 range or below detection limits
consumed by the population over 1 year in a defined zone (generally (Table 3). As WWTP removal rates were not taken into account, calculated PECs for
a country); hab is the number of inhabitants and 100 the correction cytotoxics appear consistent with measured concentrations in wastewaters, but are
generally higher than measured levels in surface waters.
factor for the percentages; 365 is the number of days per year
For cytostatics, to our knowledge, no occurrence data are available for these
(day.year − 1); WWinhab is the volume of wastewater per person molecules, as environmental research has not yet focused on this class of molecules.
per day (default value = 200 l.inhabitants − 1.day − 1); dilution is the Among the investigated endocrine therapy drugs, only a few have been searched in
dilution factor from wastewater treatment plant (WWTP) effluents the environment (Table 3). Tamoxifen is reported with maximum concentrations in
WWTP effluent of 42 ng.l− l (Ashton et al., 2004), and of 102 ng.l− l and of 25 ng.l− l in
to surface waters (default value set at 10); Fexcreta is the excretion
WWTP effluent and in surface water respectively (Coetsier et al., 2009); nevertheless,
fraction of the active molecule; Fstp is the fraction of emission of in the bulk of measurements, concentrations are below the detection limit. For
the drug from WWTPs directed to surface water, which can be other compounds, only data from Liu et al. (2010) are available, with Letrozole and
defined as (1-WWTP removal fraction). In most cases, WWTP removal Anastrozole measured with a high frequency, and with maximum respective concentrations
rates were not available and therefore we assumed an Fstp value of 1, of 2.38 and 3.70 ng.l− l in hospital effluent, and of 0.3 and 0.4 ng.l− l in WWTP
effluent (Table 3). Only conservative PECs could be determined for these molecules,
which corresponds to a worst-case scenario (i.e. no removal in
calculated values for Letrozole and Anastrozole are one order of magnitude higher
WWTPs). than the reported ones whereas calculated value for Tamoxifen is in the range of
Calculated PECs and investigated consumption trends are based field measurements (Table 2).
on national and local data sets. At the national scale, French actual
amounts, kindly provided by the French Health Products Safety Agency 4.2. Consumption trends for cytotoxic and cytostatic compounds
(Agence Française de Sécurité Sanitaire des Produits de Santé, AFSSAPS,
Paris) were used: two data sets for cytotoxic (ATC L01) compounds for Data from the French Health Products Safety Agency for cytotoxic and cytostatic
drugs (AFSSAPS, 2006 and AFSSAPS, 2009a) show that national consumption amounts
years 2004 and 2008 (respectively AFSSAPS, 2006; and AFSSAPS,
have increased between 2004 and 2008 (Fig. 1a and b); total amounts reached 13 t in
2009a); and one data set for endocrine therapy drugs (ATC L02) for 2004 and have increased to 17.5 t in 2008 (Appendix B).
year 2008 (AFSSAPS, 2009b). Results also show that antimetabolites, mainly represented by Capecitabine and
At the local scale, consumption data for years 2004 and 2008, Fluorouracil (90% of the total amounts of antimetabolites in 2008), and Hydroxycarbamide
originating from a regional specialized cancer care centre, the Centre (Hydroxyurea) are the main classes used in France (Fig. 1a and b). Hydroxycarbamide
(ATC class L01XX05) is by far the most consumed anticancer drug in France. Its consump-
Léon Bérard (CLB) located in Lyon, France, were used. tion represents 44% and 39% of the total amounts for 2004 and 2008 respectively, and it is
Gathered data are given in Appendices B and C, respectively for almost completely available in town pharmacies. Due to this specific consumption pattern,
AFSSAPS and CLB data. we chose to consider this molecule separately from others. Antimetabolites and alkylating
agents are the main classes used in hospitals (Fig. 1a and b; Appendix B).
4. Results If consumed amounts have increased for almost all anticancer chemical classes,
except for plant alkaloids (Fig. 1b; Appendix B), this is not the case for all individual
4.1. Exposure assessment drugs: for example, Mitotane consumption has doubled over the past 4 years, whereas a
nearby 40% decline in consumption amounts can be observed for Methotrexate.
Based on Eq. (1), PEC values were determined for a wide range of anticancer drugs. Consumption trends also indicate that some molecules have completely disappeared,
The calculated values are presented in Table 2 for years 2004 and 2008. PECs proposed replaced by new ones, presumably more efficient or with less adverse effects. To this
here provide a rough insight of the national contamination, not accounting for local extent, it can be observed that the use of cytostatic molecules (protein kinase inhibitors
specificities. and monoclonal antibodies) has increased from 2004 to 2008 and that consumption
Due to scarcity of data, WWTP removal rates were not taken into account in the amounts have risen for all the compounds belonging to this family (from 4% to 8% of
following calculations. Two PEC values were determined for each molecule: a conservative the total amounts). Among cytostatics, Protein kinase inhibitors are almost completely
PEC, assuming no human metabolism, and a second one, refined by excretion rates, when delivered in town pharmacies (Fig. 1b).
available. The most interesting point however, is the distinct shift that can be observed in
Conservative PECs are to be considered as a representation of what currently enters consumption patterns between 2004 and 2008 (Fig. 1a and b; Appendix B). In 2004,
the aquatic environment rather than what is in reality occurring in surface water. On 82% of anticancer drugs were delivered in hospitals (regardless of Hydroxycarbamide,
the basis of such a worst-case scenario (i.e. 100% excretion and 0% removal in see Appendix B for details), while in 2008 this proportion was only about 35%. This
WWTPs), calculated values appear to be very low, with only 6 molecules displaying a shift is observed for almost all classes of anticancer drugs, and notably for antimetab-
conservative PEC higher than 10 ng.l− 1 (the EMEA threshold value for the environmental olites (Fig. 1a and b). This is of importance for the assessment of entry routes of anti-
risk assessment of pharmaceuticals; EMEA, 2006) and 22 molecules with a PEC higher cancers in the aquatic environment (see Section 5.2).
than 1 ng.l− 1. Overall, nearly half of the anticancers used display very low conservative Local data from the Centre Léon Bérard (Lyon, France) also show that consump-
PECs, lower than 0.1 ng.l− 1 (Table 2). Compared to most of the other pharmaceutical tion amounts have increased between 2004 and 2008 (Fig. 2a and b). CLB total
classes, conservative PECs for anticancer drugs are much lower. They are up to 3 orders of amounts consumed were of 22 kg in 2004 and have increased to 26 kg in 2008 (Ap-
magnitude lower than anti-inflammatory drugs, antihypertensives and certain antibiotics. pendix C).
They are however in the lower range of serotoninergic antidepressants (Besse et al., Not all the molecules used at the national scale are used at the CLB (Appendix C).
2008), and in the same range than synthetic estrogens and progestagens (Besse and Garric, Nevertheless the CLB local consumption trends are similar to the national ones, and the
2009). main chemical classes used are antimetabolites and alkylating agents, which confirms
For cytotoxics, only three molecules have a conservative PEC greater than 10 ng.l− 1: observations made at the national scale for hospitals (Fig. 2a; 1a and 1b). As Hydroxy-
Hydroxycarbamide, Capecitabine and Fluorouracil. For cytostatic compounds, Imatinib is carbamide is almost completely delivered in town pharmacies, it is logically used in
the only molecule displaying a conservative value higher than 10 ng.l− 1, while seven very low amounts at the CLB (Appendix C). Contrary to national trends, the use of
molecules have a PEC higher than 1 ng.l− 1 (Table 2). Finally, for endocrine therapy plant alkaloids has increased between 2004 and 2008 at the CLB. Overall, and despite
drugs, calculated values are also under the ng.l− 1 range, except for a few molecules: differences observed for a limited number of molecules (probably linked to local patterns
the antiandrogens Bicalutamide, Flutamide (PEC higher than 10 ng.l− 1); the antiandrogen in chemotherapy protocols), a good correlation is observed between national and the CLB
Nilutamide, the antiestrogen Tamoxifen and the antiaromatase Exemestane (PECs higher local data for cytotoxic molecules (Appendix C).
than 1 ng.l− 1). Concerning cytostatic molecules, the CLB data show that the use of monoclonal
Metabolism data were retrieved for approximately one-third of the anticancer drugs antibodies has considerably risen between 2004 and 2008 as consumed amounts
investigated here; interestingly, such data are mainly available for the most consumed have increased by a factor of 3 (Fig. 2a; Appendix C). Protein kinase inhibitors are
(and therefore the most studied) drugs. Excretion rates vary from less than 5% up to not used at all at the CLB, which confirms observations made at the national scale.
76 J.-P. Besse et al. / Environment International 39 (2012) 73–86

Table 2
Predicted environmental concentrations (PECs) for anticancer drugs, based on their consumption in France for years 2004 and 2008 (data from AFSSAPS, 2006, 2009a, 2009b).
Concentrations expressed in ng.l− 1. Molecules are sorted by decreasing consumption amounts (for year 2008). WWTP removal rates were not taken into account in the calculation of
the following values.

Molecule Type of Chemical class Total amounts Total amounts Conservative PEC Conservative PEC F excreta Refined PEC Refined PEC
molecule 2004 (kg) 2008 (kg) 2004 (ng l− 1) 2008 (ng l− 1) 2004 (ng.l− 1) 2008 (ng.l− 1)

Hydroxycarbamide Cytotoxic Other 5756.67 6838.63 131.43 156.13 0.50 65.72 78.07
Capecitabine Cytotoxic Antimetabolite 2620.99 5134.94 59.84 117.24 0.03 1.80 3.52
Fluorouracil Cytotoxic Antimetabolite 1690.24 1733.20 38.59 39.57 0.20 7.72 7.91
Imatinib Cytostatic Tk inhibitor 583.68 873.90 13.33 19.95 0.25 3.33 4.99
Bicalutamide Endocrine Antiandrogen n.a. 863.00 n.a. 19.70 0.55 n.a. 10.84
Flutamide Endocrine Antiandrogen n.a. 521.00 n.a. 11.90 b 0.10 n.a. b 1.19
Gemcitabin Cytotoxic Antimetabolite 339.21 379.28 7.74 8.66 0.10 0.77 0.87
Tamoxifen Endocrine Antiestrogen n.a. 377.00 n.a. 8.61 n.a. n.a. n.a.
Cyclophosphamide Cytotoxic Alkylating agent 281.84 305.73 6.43 6.98 > 0.25 > 1.61 >1.74
Estramustine Cytotoxic Other 388.38 287.62 8.87 6.57 0.05 0.44 0.33
Mitotane Cytotoxic Other 95.90 233.75 2.19 5.34 0.60 1.31 3.20
Exemestane Endocrine Antiaromatase n.a. 182.00 n.a. 4.16 n.a. n.a. n.a.
Nilutamide Endocrine Antiandrogen n.a. 169.00 n.a. 3.86 b 0.03 n.a. b0.12
Erlotinib Cytostatic Tk inhibitor 0.00 148.85 0.00 3.40 b 0.02 0.00 b0.07
Cytarabine Cytotoxic Antimetabolite 117.41 133.59 2.68 3.05 b 0.10 b 0.27 b0.31
Lapatinib Cytostatic Tk inhibitor 0.00 116.20 0.00 2.65 0.70 0.00 1.86
Ifosfamide Cytotoxic Alkylating agent 121.38 103.04 2.77 2.35 0.50 1.39 1.18
Mercaptopurine Cytotoxic Antimetabolite 102.04 94.84 2.33 2.17 0.07 0.16 0.15
Bevacizumab Cytostatic Monoclonal ab 0.00 87.12 0.00 1.99 n.a. n.a. n.a.
Carboplatin Cytotoxic Platinum derivative 64.38 83.59 1.47 1.91 1.00 1.47 1.91
Methotrexate Cytotoxic Antimetabolite 117.63 74.73 2.69 1.71 0.90 2.42 1.54
Rituximab Cytostatic Monoclonal ab 32.52 72.08 0.74 1.65 n.a. n.a. n.a.
Pipobroman Cytotoxic Alkylating agent 63.06 66.93 1.44 1.53 n.a. n.a. n.a.
Nilotinib Cytostatic Tk inhibitor 0.00 58.71 0.00 1.34 0.60 0.00 0.80
Trastuzumab Cytostatic Monoclonal ab 15.05 56.01 0.34 1.28 n.a. n.a. n.a.
Cetuximab Cytostatic Monoclonal ab 7.38 55.03 0.17 1.26 n.a. n.a. n.a.
Temozolomide Cytotoxic Alkylating agent 29.23 53.54 0.67 1.22 0.10 0.07 0.12
Irinotecan Cytotoxic Other 33.89 46.58 0.77 1.06 > 0.50 > 0.38 >0.53
Etoposide Cytotoxic Plant alcaloid 332.84 41.11 7.60 0.94 0.93 7.07 0.87
Tegafur Cytotoxic Antimetabolite 86.51 37.31 1.98 0.85 n.a. n.a. n.a.
Paclitaxel Cytotoxic Plant alcaloid 27.29 38.75 0.62 0.88 0.18 0.11 0.16
Pemetrexed Cytotoxic Antimetabolite 0.92 37.31 0.02 0.85 0.90 0.02 0.77
Procarbazine Cytotoxic Methylhydrazine 16.48 34.55 0.38 0.79 b 0.20 b 0.08 b0.16
Letrozole Endocrine Antiaromatase n.a. 34.30 n.a. 0.78 n.a. n.a. n.a.
Oxaliplatin Cytotoxic Platinum derivative 20.34 33.47 0.46 0.76 n.a. n.a. n.a.
Anastrozole Endocrine Antiaromatase n.a. 31.70 n.a. 0.72 n.a. n.a. n.a.
Dacarbazine Cytotoxic Alkylating agent 18.65 29.45 0.43 0.67 0.50 0.22 0.34
Docetaxel Cytotoxic Plant alcaloid 16.41 27.47 0.37 0.63 b 0.08 b 0.03 b0.05
Bexarotene Cytotoxic Other 8.24 23.62 0.19 0.54 n.a. n.a. n.a.
Cisplatin Cytotoxic Platinum derivative 17.31 22.74 0.40 0.52 n.a. n.a. n.a.
Sunitinib Cytostatic Pk inhibitor 0.00 20.10 0.00 0.46 n.a. n.a. n.a.
Epirubicin Cytotoxic Cytotoxic ATB 18.80 17.59 0.43 0.40 n.a. n.a. n.a.
Doxorubicin Cytotoxic Cytotoxic ATB 7.94 16.82 0.18 0.38 0.50 0.09 0.19
Vinorelbine Cytotoxic Plant alcaloid 6.29 12.97 0.14 0.30 0.11 0.02 0.03
Streptozocin Cytotoxic Cytotoxic ATB 7.43 8.38 0.17 0.19 0.10 0.02 0.02
Chlorambucil Cytotoxic Alkylating agent 6.65 8.18 0.15 0.19 n.a. n.a. n.a.
Fulvestrant Endocrine Antiestrogen n.a. 6.70 n.a. 0.15 n.a. n.a. n.a.
Fludarabine Cytotoxic Antimetabolite 3.13 5.51 0.07 0.13 n.a. n.a. n.a.
Melphalan Cytotoxic Alkylating agent 3.80 4.86 0.09 0.11 n.a. n.a. n.a.
Tretinoin Cytotoxic other 1.84 3.28 0.04 0.07 n.a. n.a. n.a.
Leuprorelin Endocrine Lh-RH analog n.a. 3.12 n.a. 0.07 n.a. n.a. n.a.
Lomustine Cytotoxic Cytotoxic ATB 0.37 3.09 0.01 0.07 n.a. n.a. n.a.
Altretamin Cytotoxic Other 3.21 3.06 0.07 0.07 n.a. n.a. n.a.
Mitomycin C Cytotoxic Cytotoxic ATB 2.04 3.01 0.05 0.07 > 0.10 > 5.00 E-3 >7.00 E-3
Aminolevulinate Cytotoxic Photodynamic th. 0.00 2.39 0.00 0.05 n.a. n.a. n.a.
Thioguanine Cytotoxic Antimetabolite 1.93 2.23 0.04 0.05 n.a. n.a. n.a.
Triptorelin Endocrine Lh-RH analog n.a. 2.00 n.a. 0.05 n.a. n.a. n.a.
Carmustine Cytotoxic Cytotoxic ATB 1.41 1.81 0.03 0.04 n.a. n.a. n.a.
Mitoguazone Cytotoxic Other 1.03 1.68 0.02 0.04 n.a. n.a. n.a.
Fotemustine Cytotoxic Cytotoxic ATB 1.46 1.31 0.03 0.03 n.a. n.a. n.a.
Panitumumab Cytostatic Monoclonal ab 0.00 1.28 0.00 0.03 n.a. n.a. n.a.
Anagrelide Cytotoxic Other 0.00 1.23 0.00 0.03 n.a. n.a. n.a.
Temsirolimus Cytostatic Pk inhibitor 0.00 1.19 0.00 0.03 n.a. n.a. n.a.
Goserelin Endocrine GnRH analog n.a. 1.14 n.a. 0.03 n.a. n.a. n.a.
Toremifen Endocrine Antiestrogen n.a. 1.00 n.a. 0.02 n.a. n.a. n.a.
Bleomycine Cytotoxic Cytotoxic ATB 0.72 0.88 0.02 0.02 n.a. n.a. n.a.
Daunorubicin Cytotoxic Cytotoxic ATB 0.77 0.76 0.02 0.02 n.a. n.a. n.a.
Alemtuzumab Cytostatic Monoclonal ab 0.00 0.72 0.00 0.02 n.a. n.a. n.a.
Vinblastine Cytotoxic Plant alcaloid 0.32 0.71 0.01 0.02 n.a. n.a. n.a.
Amsacrine Cytotoxic Other 0.16 0.39 4.00 E-3 9.00 E-3 0.20 0.80 E-3 1.80 E-3
Thiotepa Cytotoxic Alkylating agent 0.21 0.35 5.00 E-3 8.00 E-3 n.a. n.a. n.a.
Mitoxantrone Cytotoxic Cytotoxic ATB 0.29 0.27 7.00 E-3 6.00 E-3 n.a. n.a. n.a.
J.-P. Besse et al. / Environment International 39 (2012) 73–86 77

Table 2 (continued)
Molecule Type of Chemical class Total amounts Total amounts Conservative PEC Conservative PEC F excreta Refined PEC Refined PEC
molecule 2004 (kg) 2008 (kg) 2004 (ng l− 1) 2008 (ng l− 1) 2004 (ng.l− 1) 2008 (ng.l− 1)

Miltefosine Cytotoxic Other 0.00 0.22 0.00 5.00 E-3 n.a. n.a. n.a.
Bortezomib Cytotoxic Other 0.00 0.22 0.00 5.00 E-3 n.a. n.a. n.a.
Topotecan Cytotoxic Other 0.10 0.21 2.00 E-3 5.00 E-3 n.a. n.a. n.a.
Idarubicine Cytotoxic Cytotoxic ATB 0.13 0.17 3.00 E-3 4.00 E-3 n.a. n.a. n.a.
Vincristine Cytotoxic Plant alcaloid 0.14 0.15 3.00 E-3 3.00 E-3 n.a. n.a. n.a.
Clofarabine Cytotoxic Antimetabolite 0.00 0.14 0.00 3.00 E-3 n.a. n.a. n.a.
Pirarubicine Cytotoxic Cytotoxic ATB 0.03 0.13 1.00 E-3 3.00 E-3 n.a. n.a. n.a.
Vindesine Cytotoxic Plant alcaloid 0.05 0.06 1.00 E-3 1.00 E-3 n.a. n.a. n.a.
Arsenic trioxyde Cytotoxic Other 0.02 0.05 1.00 E-3 1.00 E-3 n.a. n.a. n.a.
Buserelin Endocrine GnRH analog n.a. 0.05 n.a. 1.00 E-3 n.a. n.a. n.a.
Porfimer sodium Cytotoxic Photodynamic th. 0.01 0.04 0.00 1.00 E-3 n.a. n.a. n.a.
Cladribine Cytotoxic Antimetabolite 0.02 0.03 0.00 1.00 E-3 n.a. n.a. n.a.
Busulfan Cytotoxic Alkylating agent 0.15 0.03 1.00 E-3 1.00 E-3 n.a. n.a. n.a.
Raltitrexed Cytotoxic Antimetabolite 0.03 0.03 1.00 E-3 1.00 E-3 0.50 0.50 E-3 0.50 E-3
Pentostatine Cytotoxic Other 0.00 0.02 0.00 1.00 E-3 n.a. n.a. n.a.
Alitretinoin Cytotoxic Other 0.00 18.00 E-3 0.00 4.00 E-4 n.a. n.a. n.a.
Trabectedine Cytotoxic Plant alcaloid 0.00 13.00 E-3 0.00 3.00 E-4 n.a. n.a. n.a.
Chlormetine Cytotoxic Alkylating agent 0.56 8.00 E-3 1.00 E-3 2.00 E-4 n.a. n.a. n.a.
Ibritumomab tiuxetan Cytotoxic Photodynamic th. 78.00 3.00 E-3 1.78 1.10-4 n.a. n.a. n.a.

Tk inhibitor: Tyrosine kinase inhibitor.


Pk inhibitor: Protein kinase inhibitor.
Monoclonal ab: Monoclonal antibody.
Cytotoxic ATB: Cytotoxic antibiotic.
Photodynamic th.: Photodynamic therapy.
n.a.: not available or incomplete data.
Fexcreta: fraction excreted as unchanged drug.

Table 3
Concentrations of anticancer drugs in different environmental samples (expressed in ng.l− 1). WWTP : wastewater treatment plant; nd: not detected; m: mean; md: median; max:
maximum.

Molecule Sample Concentration (ng.l− 1) References

Cyclophosphamide Surface water nd–64.8 Moldovan (2006)


WWTP effluent 0.6 (md) Zuccato et al. (2005)
Hospital effluent 20–4500 Steger-Hartmann et al. (1996)
WWTP influent nd–143 Steger-Hartmann et al. (1996)
WWTP effluent nd–17 Steger-Hartmann et al. (1996)
Surface water 0.05–0.17 Buerge et al. (2006)
Hospital effluent 30–900 Catastini et al. (2008)
WWTP effluent 300 Catastini et al. (2008)
Ifosfamide Hospital effluent nd–1914 (md = 109) Kümmerer et al. (1997)
WWTP influent nd–40 Kümmerer et al. (1997)
WWTP influent nd (md)–2900 (max) Ternes (1998)
Surface water nd (md)–20 (max) Ternes (1998)
Surface water b0.05–0.14 Buerge et al. (2006)
Hospital effluent 30–900 Catastini et al. (2008)
WWTP effluent 100 Catastini et al. (2008)
Hospital effluent b2–10,647 (max) Yin et al. (2010)
Daunorubicin Hospital effluent 260–1350 Mahnik et al. (2007)
Etoposide Hospital effluent 110–600 Catastini et al. (2008)
Hospital effluent b5–380 (max) Yin et al. (2010)
Methotrexate Hospital effluent nd–200 Catastini et al. (2008)
WWTP effluent 30 Catastini et al. (2008)
Hospital effluent b2–4689 (max) Yin et al. (2010)
Bleomycin WWTP influent 15.8 (m) Aherne et al. (1990)
Surface water 8.5 (m) Aherne et al. (1990)
Hospital effluent b8600–124,000 Mahnik et al. (2004)
Hospital effluent nd–30 Catastini et al. (2008)
Tamoxifen Surface water 27–212 Roberts and Thomas (2006)
WWTP effluent nd–42 (max) Ashton et al. (2004)
WWTP effluent 102 (max) Coetsier et al. (2009)
Surface water 25 (max) Coetsier et al. (2009)
Hospital effluent 0.2–8.2 Liu et al. (2010)
WWTP influent 0.28 Liu et al. (2010)
Letrozole Hospital effluent 0.20–2.38 Liu et al. (2010)
WWTP influent 0.28–0.8 Liu et al. (2010)
WWP effluent 0.27–0.6 Liu et al. (2010)
Anastrozole Hospital effluent 0.3–3.7 Liu et al. (2010)
WWTP influent 0.12–0.32 Liu et al. (2010)
WWP effluent 0.3 Liu et al. (2010)
78 J.-P. Besse et al. / Environment International 39 (2012) 73–86

a a
8000 9000

7000 8000
7000
6000
6000
5000
5000
kg 4000 g
4000
3000 3000
2000 2000
1000 1000
0 0
Antimetabolites Hydroxycarbamide Alkylating agents Antimetabolites
Pharmacies 2004 Pharmacies 2008 Hospital 2004 Hospital 2008
Complete hosp 2004 Complete hosp 2008 Outpatients 2004 Outpatients 2008

b b
1200
1400
1000
1200
800
1000
kg 600
800
400 g
600
200
400
0
Alkylating Alcaloids and Cytotoxic Platinum Monoclonal Protein Other 200
agents related antibiotics derivatives antibodies kinase
inhibitors 0
Pharmacies 2004 Pharmacies 2008 Hospital 2004 Hospital 2008 Alcaloids and Cytotoxic Platinum Monoclonal Other
related antibiotics derivatives antibodies
Fig. 1. a. Consumption trends in French town pharmacies and hospitals for years 2004 Complete hosp. 2004 Complete hosp 2008 Outpatients 2004 Outpatients 2008
and 2008, for the two most consumed anticancer chemical classes (data from AFSSAPS,
2006, 2009a). Amounts are given in kg. Due to its high consumption amounts, Fig. 2. a. Consumption trends for anticancer drugs at the Centre Léon Bérard (Lyon,
Hydroxycarbamide was treated separately from other molecules (see Section 4.2). France); sorted by “Complete hospitalization” and “Outpatients” for years 2004 and
b. Consumption trends in French town pharmacies and hospitals for years 2004 and 2008 for the two most consumed anticancer chemical classes (CLB data; Appendix C).
2008, for the other anticancer chemical classes (data from AFSSAPS, 2006, 2009a). Amounts are given in grams. Complete hosp: Complete hospitalization. b. Consumption
Amounts are given in kg. Due to its high consumption amounts, Hydroxycarbamide trends for anticancer drugs at the Centre Léon Bérard (Lyon, France); sorted by
was treated separately from other molecules (see Section 4.2). “Complete hospitalization” and “Outpatients” for years 2004 and 2008 for the other
anticancer chemical classes (CLB data; Appendix C). Amounts are given in grams.
Complete hosp: Complete hospitalization.

Results are displayed in Table 4. Preferential molecules are:


5. Discussion Hydroxycarbamide, Capecitabine, Cyclophosphamide, Ifosfamide,
Mitotane, Imatinib, Lapatinib, Tamoxifen and Flutamide. For each
5.1. Targeting anticancer drugs to be searched in surface waters molecule, its inclusion to the list is discussed along with subsequent
recommendations.
5.1.1. Parent molecules Even though irrelevant to the standpoint of the risk, such a list is
For anticancer drugs, scientific knowledge is limited, both on useful to i) help to define environmental tracers of anticancers
ecotoxicological and occurrence data. Consequently, and pragmatically, contamination, ii) highlight molecules on which further occurrence
in order to gain knowledge on these molecules, it seems sound to first and ecotoxicological studies should be conducted and iii) determine, for
focus on compounds which are the most likely to be present in effluents the implementation of ecotoxicological assays, exposure concentration
or surface waters: indeed, searching for anticancer contaminants at the ranges consistent with the environmental reality.
ng.l− 1 range means facing analytical challenges. Therefore, as a first
step, we consider that it is better to focus primarily on molecules for 5.1.2. Metabolites
which there is a higher probability of detection in the environment. As in previous works (Besse and Garric, 2008; Besse et al., 2008),
From this perspective, we built a “preferential” (rather than priority) special attention was paid to metabolites. Metabolism of anticancer
list, based on exposure only, which was implemented as follows: compounds is complex and leads to the formation of multiple
Considering the range of measured concentrations in the environ- pharmacologically active and inactive metabolites. However, if antican-
ment, the number of molecules with a conservative PEC lower than cer drug metabolism is well documented (Appendix A), quantitative
1 ng.l− 1, and the analytical feasibility, we chose arbitrarily to set a available data (i.e. excretion rates) are limited and only allow calculating
threshold value at 1 ng.l− 1 in order to rank anticancer drugs. Molecules PECs for a very few metabolites.
with a conservative PEC (i.e. without taking into account metabolism A preferential list was established for metabolites following the
and WWTP removal rates) lower than this value were a priori excluded same rules than for parent compounds. Metabolites considered here
from the list of preferential molecules. were those reported to be pharmacologically active and/or those
Among compounds with a PEC higher than 1 ng.l− 1, metabolism, with an excretion rate higher than 10% (Besse et al., 2008; EMEA,
WWTP removal rates and biodegradation data were reviewed exhaus- 2006). Results and discussions are presented in Table 5. For metabolites,
tively, and refined PECs were calculated when possible (using Eq. (1)). degradation data and environmental behavior are more than limited, so
Molecules with a refined PEC higher than 1 ng.l− 1 were considered as it remains uncertain to which extent they can occur in the aquatic
preferential compounds. For compounds with a conservative PEC environment.
higher than 1 ng.l− 1 and for which neither reliable excretion data nor Here we briefly discuss a few of the selected metabolites, as they
reliable biodegradation data were available, we considered that it was appear to display specific properties. 7-Hydroxymethotrexate, which
not possible to conclude. pharmacological activity is controversial (Fabre and Goldman, 1985;
J.-P. Besse et al. / Environment International 39 (2012) 73–86 79

Table 4
Preferential list of anticancer drugs for the aquatic environment, based on calculated PECs, available pharmacological, metabolism and biodegradation data.

Molecule Mechanism PEC Excretion WWTP Refined Conclusion Discussion


of action (ng.l− 1) fraction removal rate/ PEC (ng.l− 1)
Biodegradation

Hydroxycarbamide Other 156.13 0.5 nd 78.1 Preferential 1) Hydroxycarbamide (Hydroxyurea) is by far


the most consumed cytotoxic molecule (44% and 39%
of the total amounts consumed in 2004 and 2008 respectively)
2) Need to confirm its occurrence in wastewaters
and surface waters and if necessary to conduct
ecotoxicological assays
Capecitabine Antimetabolite 117.24 0.03 0.15a 2.9 Preferential 1) Precursor of Fluorouracil
2) Despite a very low excretion fraction, the refined
PEC is still higher than 1 ng.l− 1
3) Even if the highest biodegradation reported value is used,
the refined PEC is higher than 1 ng.l− 1
4) Need to assess occurrence of this molecule
in the environment
Fluorouracil Antimetabolite 39.57 0.2b >0.9c b0.8 Non 1) No need to assess for its occurrence considering
preferential current scientific knowledge
Imatinib Tk inhibitor 19.95 0.25 nd 5 Preferential 1) No data available on environmental behavior
2) Need to confirm its presence in the aquatic environment
d
Bicalutamide Antiandrogen 19.7 0.55 not readily 15.7 Preferential 1) Possibly only weakly active if (S)-enantiomer is the main
biodegradablee excreted compound (see reference d)
2) Need to confirm its presence in the aquatic environment
Flutamide Antiandrogen 11.9 b0.1 nd b1.2 Non 1) No need to assess for occurrence considering
preferential current scientific knowledge
Gemcitabine Antimetabolite 8.66 0.1 0.4f 0.52 Non 1) No need to assess for occurrence considering
preferential current scientific knowledge
Tamoxifen Antiestrogen 8.61 nd Not readily nd Preferential 1) Only limited data on excretion
biodegradableg 2) Could be excreted in significant amounts in feces
as unchanged drug or glucuronide
(BCB, 2011; Kisanga et al., 2005)
3) Already detected in surface waters and wastewaters
Cyclophosphamide Alkylating 6.98 >0.25 0h >1.75 Preferential 1) Expected to occur in the environment at the
agent ng.l− 1 range due to its very low biodegradability
(Kümmerer et al., 2000)
2) Refined PECs are consistent with field measurements
Estramustine Alkylating 6.57 nd nd nd nd 1) Only approximately 75% of the drug is orally absorbed, and
agent unchanged Estramustine can be excreted through biliary route
2) This molecule might be present in the aquatic environment
at concentrations higher than the ng.l− 1, but metabolism
and degradation data are too scarce to allow drawing any
conclusion.
Mitotane Other 5.34 0.6 nd 3.2 Preferential 1) No data available on environmental behavior
2) Need to confirm its presence in the aquatic environment
Exemestane Antiaromatase 4.16 nd >0.9i b0.4 Non 1) No need to assess for occurrence considering current
preferential scientific knowledge
Nilutamide Antiandrogen 3.86 b0.02 nd 0.07 Non 1) No need to assess for occurrence considering current
preferential scientific knowledge
Erlotinib Tk inhibitor 3.4 b0.02 Not readily b0.07 Non 1) No need to assess for occurrence considering current
biodegradablej preferential scientific knowledge
Cytarabine Antimetabolite 3.05 b0.1 0.5k b0.15 Non 1) No need to assess for occurrence considering current
preferential scientific knowledge
Lapatinib Tk inhibitor 2.65 0.7l nd 1.82 Preferential 1) The excretion rate used here is the maximum reported
value and might not be representative of the general case.
2) If median excretion value is used (Appendix A),
Lapatinib is no longer a preferential compound
Ifosfamide Nitrogen 2.35 0.5 0m 1.18 Preferential 1) Expected to occur in the environment at the
mustard ng.l− 1 range due to its very low biodegradability
(Kümmerer et al., 1997, 2000)
2) Refined PECs are consistent with field measurements
Mercaptopurine Antimetabolite 2.17 0.07 nd 0.15 Non 1) No need to assess for its occurrence considering
preferential current scientific knowledge
Bevacizumab mab 1.99 nd nd nd nd 1) Data too scarce to conclude
Carboplatin Platinum 1.91 1 0.65n 0.67 Non 1) Reported to be removed in wastewater and
derivative preferential during water treatment processes,
2) Adsorption to sludge and solid particles
(Lenz et al., 2005, 2007)
Methotrexate Antimetabolite 1.71 0.9 0.95o 0.07 Non 1) No need to assess for its occurrence considering
preferential current scientific knowledge
2) Refined PEC could be higher as Methotrexate
is also used for the treatment of Rheumatoid
polyarthritis under another ATC class (L04)
Rituximab mab 1.65 nd nd nd nd 1) Data too scarce to conclude
Pipobroman Alkylating 1.53 nd nd nd nd 1) Data too scarce to conclude
agent

(continued on next page)


80 J.-P. Besse et al. / Environment International 39 (2012) 73–86

Table 4 (continued)
Molecule Mechanism PEC Excretion WWTP Refined Conclusion Discussion
of action (ng.l− 1) fraction removal rate/ PEC (ng.l− 1)
Biodegradation

Nilotinib Tk inhibitor 1.34 0.6 nd 0.8 Non 1) No need to assess for occurrence considering
preferential current scientific knowledge
Trastuzumab mab 1.28 nd nd nd nd 1) Data too scarce to conclude
Cetuximab mab 1.26 nd nd nd nd 1) Data too scarce to conclude
Temozolomide Alkylating 1.22 0.1 nd 0.12 Non 1) No need to assess for occurrence considering current
agent preferential scientific knowledge
Irinotecan Other 1.06 >0.5 nd >0.5 nd 1) Data too scarce to conclude

nd: not determined.


Tk inhibitor: tyrosine kinase inhibitor
mab: monoclonal antibody.
a
OECD 302B, 15% removal at 7 days, up to 58% removal at 28 days (reported values from Straub, 2010).
b
Taking into account the fraction of Capecitabine excreted as Fluorouracil.
c
Batch experiment, incubated with activated sludge (Mahnik et al., 2004); complete biodegradation with test OECD 303A (Kiffmeyer et al., 1998); reported not biodegradable in
the closed bottle test and Zahn–Wellens test (Kümmerer and Al-Ahmad, 1997).
d
Data from FASS, 2011. Bicalutamide is a racemate, which activity resides almost exclusively in the (R)-enantiomer, and a part of the excreted Bicalutamide is the inactive
(S)-enantiomer (Cockshott, 2004).
e
Not readily biodegraded: OECD 301C, OECD 302A (FASS, 2011).
f
Closed bottle test and Zahn–Wellens test, from 42% to 50%, depending on tests conditions (Kümmerer and Al-Ahmad, 1997).
g
Not readily biodegraded: OECD 301C (FASS, 2011).
h
Suggested to be persistent in WWTP (Buerge et al., 2006); reported not biodegradable in closed bottle test and Zahn–Wellens test (Kümmerer et al., 2000).
i
Not considered to be persistent, OECD 308 (Ericson, 2007).
j
Not readily biodegraded, OECD 301C (FASS, 2011).
k
Closed bottle test and Zahn–Wellens test, from 50% to more than 95%, depending on test conditions (Kümmerer and Al-Ahmad, 1997).
l
Considering maximal excretion rate (median excretion rate of 27%, see Appendix A for details).
m
Reported not biodegradable in closed bottle test and Zahn–Wellens test (Kümmerer et al., 2000).
n
From 65% to 82% removal rate with membrane bioreactor alone or with advanced sewage treatment processes (Lenz et al., 2007).
o
OECD confirmatory test (Kiffmeyer et al., 1998); reported as readily biodegradable with OECD 301F test (Henschel et al., 1997).

Lingg et al., 2005; Seither et al., 1989), has been reported to appear influence levels of anticancers entering the aquatic environment,
during biodegradation experiments of Methotrexate, and further- it therefore appears necessary to continue surveying their presence
more, not biodegradable (Kiffmeyer et al., 1998). It remains unclear in WWTP effluents and surface waters.
to which extent the biodegradation of Methotrexate can lead to the Consumption data also show that hospital effluents may not be the
formation of 7-hydroxymethotrexate. Nevertheless, as this metabolite primary input source of anticancer drugs in the environment. Indeed,
is potentially active, not readily biodegradable, and as it could be a distinct shift in consumption trends has been observed between
found in the aquatic environment at levels higher than the parent 2004 and 2008, which appears to be linked with the generalization
compound, occurrence studies should be conducted. of home treatment in order to improve patients' comfort. The fact
The antiestrogenic Tamoxifen is metabolized into two active that an increasing number of molecules originally restricted to hospitals
metabolites, Hydroxy-Desmethyltamoxifen (Endoxifen) and 4- are now available in town pharmacies, combined with the development
Hydroxytamoxifen. The two metabolites have been reported to be of such home treatments, leads to a more diffuse rejection of anticancers
100 times more potent and active than the parent compound and throughout a given town. This contradicts the generally admitted idea
are considered responsible for the pharmacological activity (BCB, that hospital effluents are the primary input source of anticancer drugs
2011; Micromedex Drugdex©, 2011; Wu et al., 2009; Zheng et al., in the environment.
2007). Available metabolism data do not allow determining a reliable Local consumption data from the Centre Léon Berard (CLB), in
PEC for these metabolites, but they have been detected in bile and Lyon, which allows monitoring the distribution patterns of anticancer
urine (Kisanga et al., 2005). Considering their high antiestrogenic activity drugs depending on the type of hospitalization (complete hospitalization
and their potential presence in the aquatic environment, these metabo- and outpatients), provide a meaningful illustration for this. The hospital-
lites should be investigated for occurrence, and if detected in the aquatic ization type strongly influences the entrance pathway of anticancer
environment, tested for ecotoxicity, as they might be of greater concern drugs in the sewage system. Molecules consumed by patients during
than the parent Tamoxifen. complete hospitalization at the CLB are 100% evacuated with the center's
Searching for metabolites in surface waters can be costly and hard to effluents. On the contrary, only up to 20% of the drugs delivered to
achieve due to limited available standards, and because they are expected outpatients are considered consumed on site. Therefore, only 20% of
to be present at low concentrations. Nevertheless, pharmaceutical metab- outpatient delivered drugs are eliminated with hospital effluents, while
olites, which have already been detected in surface waters and sediment the remaining 80% are consumed by patients at home and thus are
(Langford and Thomas, 2010), and subsequently anticancer metabolites, discharged into the sewer system diffusely throughout the town and
require consideration as they can exert biological effects on aquatic organ- its suburbs. In 2004 and 2008, amounts delivered to outpatients
isms. Moreover, they should be taken into account when assessing the accounted for approximately half of the total amounts consumed at
risk of pharmaceuticals (Besse et al., 2008; Celiz et al., 2009; EMEA, the CLB. Based on available AFSSAPS and CLB data, we established a the-
2006). To this extent, the work provided here is incomplete as only oretical scheme of anticancer drugs inputs in the aquatic environment
human metabolites were considered, but no environmental degradation (Fig. 3). These results are of importance for anticancer drug risk
compounds such as photolysis by-products. Nevertheless, current knowl- management.
edge is probably insufficient to support any conclusion on the latter. Data available here seem to indicate that this shift in consumption
trends is observed both at a national and a local scale. However, such an
5.2. Implications of consumption trends assumption should be confirmed with other recent national and local
data sources, originating from other hospitals and cancer care centers.
National and local consumption data both show an increase in the Finally, if hospital effluents are no longer the main expected entry
consumption of anticancer drugs. As consumption trends directly route of anticancer drugs in the environment, they nevertheless
J.-P. Besse et al. / Environment International 39 (2012) 73–86 81

Table 5
Preferential list of anticancer metabolites for the aquatic environment, based on calculated PECs, available pharmacological, metabolism, and biodegradation data.

Molecule Active Parent compound Excretion PEC Conclusion Discussion


(PEC, ng.l− 1) fraction (ng.l− 1)

N-desmethylmetabolite Yes Imatinib (19.95) 0.13 2.6 Preferential 1) No degradation data available
(CGP74588) 2) Need to confirm occurrence in waters
Hydroxy-bicalutamide No Bicalutamide 0.32 6.3 Preferential 1) No degradation data available
(19.7) 2) Need to confirm occurrence in waters
Hydroxy-flutamide Yes Flutamide (11.5) nd nd nd 1) Data too scarce to conclude
2′,2′-Difluorodeoxyuridine Yes Gemcitabine 0.9 7.8 Preferential 1) Reported to be pharmacologically active
(dFdU) (8.66) (Benyumov et al., 2011; Veltkamp et al., 2008)
2) Suspected to contribute to the toxicity of Gemcitabine
(Micromedex Drugdex©, 2011)
3) Occurrence and, if necessary, ecotoxicity studies should be conducted
4-Hydroxy-N- Yes Tamoxifen (8.61) nd nd Preferential NB: otherwise indicated, the following considerations concern the two
desmethyltamoxifen metabolites of Tamoxifen:
(Endoxifen) 1) 100 times more affinity to estrogen receptor than Tamoxifen
(BCB, 2011; Wu et al., 2009; Zheng et al., 2007)
2) 100 times more potent than Tamoxifen
(Wu et al., 2009; Zheng et al., 2007)
3) Could be excreted as glucuronide conjugates
4) Detected in urine, but no excretion rates are available
(Kisanga et al., 2005)
5) Hydroxy-tamoxifen could be the main metabolite of Tamoxifen
6) No degradation data are available
7) Occurrence is to be confirmed and if, necessary, ecotoxicity studies should
be conducted for these two metabolites
4-Hydroxy-tamoxifen Yes Tamoxifen (8.61) nd nd Preferential
Phosphoramide mustard Yes Cyclophosphamide 0.4 b1 Non 1) Considered responsible for pharmacological effects
(6.98) preferential 2) Reported to be rapidly hydrolyzed: half-life at pH 7.4
and ambient temperature of 2 h (Joqueviel et al., 1998)
3) No need to assess for its occurrence considering current scientific
knowledge
Estromustine Yes Estramustine nd nd nd 1) Reported to be cytotoxic (BCB, 2011)
(6.57) 2) Neither excretion nor biodegradation data are available
3) Not possible to conclude on its occurrence in the environment
17-hydro-exemestane Yes Exemestane (4.16) nd nd nd 1) Data are too scarce to conclude
N-Desmethylmetabolite Yes Erlotinib (3.4) 0.03 0.1 Non 1) No need to assess for occurrence, considering
(OSI-420) preferential current scientific knowledge
Carboxylic acid metabolite nd Erlotinib (3.4) 0.3 1 Preferential 1) No degradation data available
2) Need to confirm occurrence in water
Uracil-arabinoside No Cytarabine (3.05) 0.9 2.74 Preferential 1) No degradation data available
2) Need to confirm occurrence in water
7-Hydroxy-methotrexate Yes Methotrexate 0.15 nd Preferential 1) Pharmacological activity unclear (see Section 5.1.2)
(1.71) 2) Reported to appear during biodegradation
experiment of Methotrexate, (Kiffmeyer et al., 1998)
3) Reported as not biodegradable (Kiffmeyer et al., 1998)

nd: not determined.

remain a specific input source for specific pharmaceuticals (i.e. re- and new chronic data, built using standardized chronic assays, it
stricted to hospitals) and other macro and micropollutants (Verlicchi was concluded that there was no exposure level of concern in the
et al., 2010), or even pathogenic microorganisms (Emmanuel et al., aquatic environment for these molecules.
2009; Prado et al., 2011). Furthermore, some chronic data, based on standardized chronic
Overall, variability and trends in consumption amounts illustrate tests on Daphnia magna, suggest that Cytarabine and Gemcitabine are
the need to make consumption data public and easily accessible for also of limited environmental concern at their current environmental
anticancer drugs but also for all other pharmaceutical products. Public exposure level (Zounkova et al., 2010).
access of consumption amounts allows determining what is currently As quoted by several authors (Johnson et al., 2008; Kümmerer and
entering the aquatic environment and what are the input pathways. Al-Ahmad, 1997), genotoxic effects of cytotoxic drugs are expected to
Subsequent exposure assessments, such as the one implemented be a major issue for the aquatic environment. Studies conducted until
here, allow presenting an exhaustive overview of molecules likely to now, and based on in vitro assays, suggest that the environmental
be of environmental concern and are helpful for any further occurrence concentrations of tested cytotoxics are under concentrations required
or ecotoxicological investigation. Furthermore, they are the basis of any to cause genotoxic effects (Ferk et al., 2009; Zounkova et al., 2010, 2007).
comprehensive risk assessment. Overall, it appears that most of the molecules have been tested only
by acute standardized tests or genotoxic in vitro assays (Table 6a), mak-
5.3. Risk considerations ing it difficult to conclude on their environmental risk. Therefore, stud-
ies need to be implemented in order to acquire relevant
5.3.1. Cytotoxic compounds ecotoxicological data. To this extent, the suitability of standardized
Currently, only very few ecotoxicological data are available for tests and endpoints for the assessment of cytotoxic drugs is arguable:
cytotoxic drugs (Table 6a). Recently, an extensive risk assessment if acute tests measuring survival endpoints are clearly inadequate, as
based on the EMEA guideline (EMEA, 2006) was conducted for shown on other pharmaceutical classes (Kümmerer et al., 2004), the
Capecitabine and Fluorouracil (Straub, 2010). On the basis of PECs, use of algal growth, Daphnia magna reproduction, and even fish sub-
available measured concentrations, available ecotoxicological data, chronic tests can be questioned, as they do not seem suitable for
82 J.-P. Besse et al. / Environment International 39 (2012) 73–86

Total delivered amounts or surface waters. However, since they only measure subcellular
effects, it is difficult to relate observed effects on a single compound
78% of the 22% of the (or even a mixture) with potential environmental effects at the indi-
total amountsa total amountsa vidual or population level. We therefore consider that no conclusion
Town Hospitals
on environmental genotoxic effects should be drawn on the sole
basis of in vitro assays. The in vivo comet assay, which is increasingly
10.3%b 11.7%b used to assess genotoxicity in organisms exposed to contaminants
8.2%c in situ or in laboratory experiments (Frenzilli et al., 2009; Lacaze
Consumption at Complete
Outpatients et al., 2010), could be a more suitable tool to investigate the geno-
home hospitalization
toxic effects of cytotoxic compounds on aquatic species. A possible per-
2.1%d 11.7% spective could be the assessment of genotoxic effects on spermatozoa
and oocytes of exposed aquatic organisms (Lacaze et al., 2011). Germi-
Diffuse rejection
in sewer system
Hospital effluents nal DNA cell damage, related to poor fertilization rate and defective
embryonic development (Aitken and De Iuliis, 2007), can lead to re-
86.2% 13.8% production impairment, which is a relevant ecotoxicological endpoint
for anticancer drugs at the individual, population, and community scale.
urban WWTP

Fig. 3. Theoretical input pathways for anticancer drugs in the aquatic environment
with relative contribution of each entry pathway. Based on gathered French data 5.3.2. Cytostatic compounds
from the AFSSAPS (national consumption amounts) and from the Centre Léon Bérard Cytostatics are a recent class of anticancer drugs considered as
(CLB; specialized cancer care center); see Section 4.2 and Appendix B for details.
a
promising for the improvement of treatment efficiency. Two sub-
: Using AFSSAPS data (AFSSAPS, 2009a). b: Using CLB data, approximately 53% of the
total amounts were delivered for complete hospitalization and 47% for outpatients, at groups can be found among this class: Protein kinase inhibitors (in
the CLB in 2008 (Appendix C). c: Approximately 80% of the anticancer drugs delivered particular tyrosine kinase inhibitors) and monoclonal antibodies.
to outpatients are considered consumed and rejected at home (i.e. outside the CLB). Tyrosine kinases belong to the protein kinase enzyme family. Protein
d
: Approximately 20% of the anticancer drugs delivered to outpatients are considered kinases are key regulators of cell function that constitute one of the largest
consumed and rejected onsite.
and most functionally diverse gene families. By adding phosphate groups
to substrate proteins, they direct the activity, localization and overall
determining long-term effects of exposure to low concentrations of function of many proteins. Protein kinases therefore mediate most of
DNA interacting molecules. To our knowledge, no experiments have the signal transduction in eukaryotic cells, regulating cellular metabolism,
been conducted on long-term toxicity of cytotoxic drugs in fish, mol- transcription, cell cycle progression, cytoskeletal rearrangement and cell
luscs, or invertebrates other than Daphnia magna. Even if hard to imple- movement, apoptosis, and differentiation. The diversity of essential
ment, full-life cycle or multigenerational assays on fish sound more functions mediated by kinases is shown by the conservation of some
relevant for assessing environmental chronic effects of cytotoxics. 50 distinct kinase families between yeast, invertebrate and mammalian
Moreover, as cytotoxics may contribute to the genotoxicity kinomes (Manning et al., 2002). Currently, nothing is known on their
detected in certain effluent or surface water samples (Jolibois and occurrence and biological effects (if any) on aquatic non-target organ-
Guerbet, 2005; Reifferscheid and Grummt, 2000), their genotoxic isms, and only limited data are available for Erlotinib (Table 6a). Never-
potential should be tested. To do so, we consider that in vivo assays theless, considering their universal mode of action and increasing
should be more suitable than in vitro tests. Indeed, in vitro assays consumption amounts, they should be taken into consideration and
are useful tools for screening genotoxic effects, notably in effluents assessed for occurrence, and if need be, for ecotoxicity.

Table 6a
Review of existing ecotoxicological data for anticancer drugs (cytotoxic and cytostatic molecules). Concentrations are expressed in mg.l− 1.

Molecule Organism Endpoint Value Concentration (mg.l− 1) References

Methotrexate V.fischerii Luminescence EC50 1220 Henschel et al. (1997)


S.subspicatus Growth EC50 72 h 260 Henschel et al. (1997)
T. pyriformis Growth EC50 48 h 45 Henschel et al. (1997)
D. magna Immobilization EC50 48 h >1000 Henschel et al. (1997)
B. rerio Survival EC50 96 h 85 Henschel et al. (1997)
X. laevis Growth EC50 96 h 0.015 Bantle et al. (1994)
Fluorouracil V. fischerii Luminescence EC50 0.12 Backhaus et al. 2000
P. promelas Growth LOEC 120 h 20 De young et al. (1996)
D. magna Reproduction LOEC 21 days 0.05 Zounkova et al. (2010)
D. magna Reproduction NOEC 21 days 0.0028 Straub (2010)
A. flos-aquae Growth NOEC 72 h 0.002 Straub (2010)
Capecitabine D. magna Reproduction EC50 48 h >850 Straub (2010)
P. subcapitata Growth NOEC 72 h 0.14 Straub (2010)
Cladribin D. magna Immobilization EC50 48 h 233 FDA-CDER (1996)
Cytarabine D. magna Reproduction LOEC 21 days 3.7 Zounkova et al. (2010)
Gemcitabin D. magna Reproduction LOEC 21 days >1.0 Zounkova et al. (2010)
P. subcapitata Growth EC50 72 h 0.57 FASS (2011)
D.magna Immobilization EC50 48 h >0.99 FASS (2011)
P. promelas Survival LC50 96 h >1000 FASS (2011)
O. mykiss Survival LC50 96 h >1000 FASS (2011)
Paclitaxel D. magna Immobilization EC50 48 h >0.74 FDA-CDER (1996)
Thiotepa D. magna Immobilization EC50 48 h 546 FDA-CDER (1996)
Erlotinib S. capricornutum Growth NOEC 72 h 0.14 FASS (2011)
D. magna Reproduction NOEC 48 h 0.7 FASS (2011)
O. mykiss Survival NOEC 14 days 0.02 FASS (2011)
J.-P. Besse et al. / Environment International 39 (2012) 73–86 83

Monoclonal antibodies are molecules that target growth factor 5.3.3.3. Antiestrogens. Among the molecules investigated, Tamoxifen is
receptors in order to exert a direct effect on the growth and survival the only one for which some occurrence and ecotoxicological data are
of cancer cells by antagonizing ligand-receptor signaling. Monoclonal available (Table 6b). A full-life cycle study has been conducted on
antibodies can target to cell surface antigens and directly elicit apoptotic Pimephales promelas (Williams et al., 2007). Several endpoints were
signaling (Ludwig et al., 2003). Conservative PEC calculated here are measured and an overall NOEC for F0 and F1 generations was
very low for this class of compound. Considering available knowledge, reported to be equal to 5.12 μg.l − 1, which is far above expected con-
it is currently impossible to conclude on the environmental significance centrations in water. Authors therefore considered that Tamoxifen is
of this class of molecules; again, there is a need for further scientific not expected to pose a risk at the current environmental levels of
investigation. exposure. However, since Tamoxifen is metabolized into two more
potent metabolites, potentially present in the aquatic environment,
5.3.3. Endocrine therapy drugs the latter should be taken into account when assessing Tamoxifen
Due to their mechanism of action, exposure to such molecules ecotoxicological effects and environmental risk.
could be hazardous for aquatic species, notably fishes, batracians or No data are available for Toremifen, however considering its very low
molluscs, displaying steroid receptors similar to mammals. conservative PEC (about 0.02 ng.l− l), risk for the aquatic environment is
expected to be limited.

5.3.3.1. Hormone analogs. Chronic administration of these molecules


in human results in a significant reduction of plasma testosterone. 5.3.3.4. Antiandrogens. To our knowledge, no occurrence data for the
Considering this, occurrence of these molecules in the aquatic envi- aquatic environment are available, and only a few ecotoxicological
ronment may be harmful for sensitive non target-organisms. However, studies have been conducted on these molecules (Table 6b). Calculated
given their very low consumption amounts and very low PEC values PECs for these molecules are low, with the higher PEC observed for Bica-
(lower than 0.1 ng.l− 1, see Table 2), the associated risk is expected to lutamide, which is notably reported as not biodegradable (Table 4). Con-
be limited relative to other endocrine disruptors, even if data remain sidering PEC values, risk posed by Nilutamide and Fulvestrant is expected
too scarce to draw any definitive conclusion. to be limited relative to other compounds from the same class. Ecotoxico-
logical data on fish indicate toxic concentrations far above PEC for Fluta-
mide, Nilutamide and even Bicalutamide (Table 6b). Overall,
5.3.3.2. Anti-aromatase molecules. These molecules act by inhibiting
considering PECs calculated here and limited available ecotoxicological
the aromatase activity, which converts androgens into estrogens. All
data, risk is expected to be limited relative to other endocrine dis-
compounds have very low PECs, either conservative or refined
rupters, but further works are needed prior to conclude definitely.
(Tables 2 and 4). Very few ecotoxicological data are available for these
molecules (Table 6b). Available data for Letrozole (Sun et al., 2007)
and some data available for Fadrozole (Gust et al., 2010; Panter et al., 5.3.3.5. 5-α-reductase inhibitors. Finasteride and Dutasteride are 5-α-
2004), an anti-aromatase molecule no more used in human medicine, reductase inhibitors that inhibit the conversion of testosterone to
report effects at the μg.l− 1 level. Studies comparing the in vitro inhibitory dihydrotestosterone. These molecules are not strictly speaking anticancer
aromatase activity of different molecules (Trösken et al., 2004, 2006) drugs but considering their mechanism of action and indications (i.e.
show that Letrozole is the most potent aromatase inhibitor among treatment of prostatic hyperplasia), we discuss them in this paper. No
those tested, with an inhibitory activity stronger than Procholoraz, occurrence studies have been conducted for these two molecules in the
Flusilazole and Imazalil (molecules used as fungicides in agriculture). aquatic environment. As gathered consumption amounts were incom-
Considering their mechanism of action and antiaromatase potency, plete (Finasteride is also used in the treatment of androgenic alopecia),
these molecules could lead to a disruption of reproductive functions in no reliable PEC could be calculated. Nevertheless, available consumption
non-target aquatic organisms, especially molluscs and fishes. However, data indicate that such a PEC is expected to be in the ng.l− 1 range.
given the very low PEC values for compounds investigated here, and Moreover, Finasteride consumption has increased since 2004.
given the lack of ecotoxicological data, it is not possible to conclude Only limited ecotoxicological data are available for Finasteride; which
for these molecules. report impairment of gonad development and sexual differentiation in

Table 6b
Review of existing ecotoxicological data for anticancer drugs (endocrine therapy drugs). Concentrations are expressed in μg.l− 1.

Molecule Organism Endpoint Value Concentration (μg.l− 1) References

Letrozole O. latipes Fecundity LOEC 21 days 5 Sun et al. (2007)


O. latipes Fertility LOEC 21 days 5 Sun et al. (2007)
O. latipes Increase in genotypic F1 males LOEC 21 days 5 Sun et al. (2007)
Tamoxifen P. promelas Overall a NOEC 284 days 5 Williams et al. (2007)
P. promelas F1 larvae growth b 28 days 0.08 Williams et al. (2007)
P. promelas F1 growth c 112 days 0.01 Williams et al. (2007)
P. promelas Increase in Vtg, F1 males d 112 days 0.01 Williams et al. (2007)
A. tonsa Larval development EC50 5 days 49,000 Andersen et al. (2001)
Nilutamide Green algae Growth NOEC 1000 FDA-CDER (1996)
Flutamide B. calicyflorus Fertilization of sexual females LOEC 96 h 1 Preston et al. (2000)
G. aculeatus Spiggin inhibition LOEC 21 days 500 Sebire et al. (2008)
G. aculeatus Male behavior LOEC 21 days 100 Sebire et al. (2008)
P. promelas Testis alterations LOEC 21 days 62 Jensen et al. (2004)
P. promelas Increase of estradiol plasma levels LOEC 21 days 651 Jensen et al. (2004)
Bicalutamide P. promelas Overall e NOEC f 10 FASS (2011)
a
NOEC value considered significant for all endpoints in F0 and F1 generations.
b
Statistically significant decrease in F1 larvae length and weight at 28 days post hatch.
c
Statistically significant decrease in F1 individuals length and weight at 112 days post hatch, observed at this concentration only.
d
Statistically significant increase in vitellogenin in F1 males at 112 days post hatch, observed at this concentration only.
e
Overall endpoints : reproduction, weight, length, secondary sexual characteristics, time to hatch, hatchability, survival and histology.
84 J.-P. Besse et al. / Environment International 39 (2012) 73–86

male tadpoles at the mg.l− 1 range (Urbatzka et al., 2009). As for other It is our point of view that many steps remain to be taken concerning
antiandrogens, data remain too scarce to make any conclusion on the environmental impact of anticancer drugs, and that assessing the
these two molecules. environmental risk they pose is a challenging though interesting and
necessary issue for the coming years.
5.4. Assessing the environmental effects of anticancer drugs To summarize our opinion on anticancer drugs, there is a need to:

As quoted by other authors (Johnson et al., 2008), we consider • assess the occurrence of parent anticancer drugs and metabolites
that it is still not possible to draw any definitive conclusions on highlighted here;
risks posed by these molecules. Regarding EMEA guidelines, risks • break away from “classical” risk assessment based on standardized
associated to anticancer drugs are generally discussed relative to tests;
isolated molecules, and so are subjected to ecotoxicological assays. • assess the effect of environmental mixtures of anticancer drugs at
Nevertheless, the true question is: what are the effects related to a environmentally realistic concentrations;
chronic exposure to a mixture of several molecules with similar mech- • focus on in vivo long term, even full life cycle tests, for the assessment
anisms of action, each at very low concentrations in the ng.l − 1 range of ecotoxicity, genotoxicity and/or endocrine disrupting effects of
(or even less)? Biotests on isolated molecules, relying on standardized anticancer drugs.
endpoints, provide meaningless information in the case or anticancer
drugs. Although currently necessary for regulatory considerations, we Acknowledgments
question the suitability of these tests for assessing environmental ef-
fects of such compounds. The authors wish to thank the AFSSAPS and particularly Paul
Due to their mechanisms of action and very low expected environ- Houeto, for kindly sharing the pharmaceutical consumption data.
mental concentrations, anticancer drugs require taking a step forward They also appreciate Pr François Locher, from the Faculty of Pharmacy of
from “classic” standardized approaches. It is our opinion that there is a Lyon, and Monique Boucquin, from the “Pharmaceutical Documentation
need to better account for the environmental reality and complexity by and Information Center (CDIP)” of Lyon hospices, for providing pharma-
testing mixtures of anticancers at environmentally realistic concentra- ceutical information. Finally the authors are very grateful to Ashley
tions (i.e. at the ng.l− 1 range). Testing for mixtures is still an emerging Tilghman-Sibille for proofreading and correcting the English, and also to
and complicated field but existing work shows how important this the two anonymous reviewers, for their helpful comments that allowed
issue is (Brian et al., 2007; Sárria et al., 2011; Sun et al., 2009). us improving this article.
Mixtures tests for anticancer drugs could be implemented according
to i) an “environmental” approach, taking into account molecules likely Supplementary data (Appendices A, B and C)
to be present in the environment; and ii) a “pharmacological” approach
based on the combination of molecules used in cancer treatment. Supplementary data to this article can be found online at doi:10.
Chemotherapy protocols are often based on combinations of several 1016/j.envint.2011.10.002.
molecules with different targets, kinetics, and mechanisms of action, in
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