ABSTRACT
Peripheral neuropathy is one of the impeding neurotoxic complications of cancer chemotherapeutic agents,1 particularly tubulin inhibitors such as vincristine and taxanes, as well as platinum-based drugs2 and, more recently, proteasome inhibitors.3 As shown in the case of the chemotherapeutic agent oxaliplatin, a consequence of chemotherapy is the development of a painful peripheral neuropathy and decreases in both nerve conduction velocity and their action potential amplitudes.4 Further, binding of cisplatin to DNA in the dorsal root ganglion neurons leads to death and atrophy of the ganglia.5 Although management of neuropathic pain using antidepressants or anticonvulsants is available for agents such as Taxol, peripheral neurotoxicity represents a dose-limiting complication.6 Indeed, this toxicity has been dose limiting for many new antitubulin drugs, such as cryptophycin-52.7 Thus, there is a need for the development of new chemotherapeutic agents or analogues of present ones that have minimal or no neurotoxicity; this seems of particular importance if antitubulin, platinum-based drugs and proteasome inhibitors are to have their full potential in chemotherapy. Of the various research models available for assessing the neurotoxicity
CONTENTS
19.1 Introduction ........................................................................................................................... 541 19.2 Materials and Methods .......................................................................................................... 542
19.2.1 Culture Chemicals .................................................................................................... 542 19.2.2 Cytotoxic Agents ...................................................................................................... 542 19.2.3 PC-12 Cell Preparation ............................................................................................. 542 19.2.4 NGF-Induced Neurite Outgrowth Assay .................................................................. 543 19.2.5 Neurite Recovery after Drug Treatment ................................................................... 543 19.2.6 PC-12 Clonogenic Cell Survival Assay .................................................................... 543
19.3 Results ................................................................................................................................... 543 19.4 Discussion .............................................................................................................................544 References ...................................................................................................................................... 547
of cytotoxic drugs, PC-12 cells have been widely used,8,9 in which assessment of nerve growth factor-dependent neurite outgrowth has been the basis for dening chemotherapy-induced neurotoxicity for a variety of compounds.10-13
We have modied this method to correlate the observed inhibition of neurite outgrowth with the cytotoxicity of the agents studied on the same PC-12 cells. We have dened the method for obtaining viable single-cell suspensions of PC-12 cells that can be used for both the neurite outgrowth assay and the clonogenic cell kill assay. The importance of demonstrating any difference between neurite outgrowth inhibition and cytotoxicity seems essential in understanding both the drug’s mechanism of action and its potential reversal. A correlation between neurite outgrowth inhibition and the effect on clonogenic cell survival by four standard anticancer agents and two novel investigational anticancer agents is presented here.
