Chemotherapy-induced peripheral neuropathy in immunodeficient mice: new useful ready-to-use animal models

Cisplatin, paclitaxel, and bortezomib represent the most employed chemotherapy regimens for the treatment of genitourinary cancers, breast and lung cancers and multiple myeloma. Nevertheless, their clinical use is often associated to the development of peripheral neuropathies characterized mostly by sensory alterations and pain (Argyriou et al., 2012). Several rat models of chemotherapy-induced peripheral neuropathy (CIPN) had been established in the past to describe the mechanisms of its development and pathogenesis. However, only few cancer cell lines induce the development of cancer in the rat, while immunodeficient mice best allowed human cancers xenografts to study at the same time, the antineoplastic and neurotoxic effects of chemotherapy. Here we characterized neuropathic pain, neurophysiological and neuropathological alterations induced by chronic chemotherapy in immunodeficient nude mice. Mice were treated with effective doses of cisplatin (4 mg/Kg, i.p), paclitaxel (80 mg/Kg, i.v) and bortezomib (0.8 mg/Kg, i.v) for a 4-6 weeks period. At the end of the 6th week all chemotherapy regimens determined a significant impairment of neurophysiologic parameters, mechanical allodynia and thermal hypo-or hyperalgesia. Light microscopy analysis of dorsal root ganglia (DRG) showed that bortezomib induced morphological alterations in the sensory neurons and satellite cells as dark inclusions and clear vacuolation throughout the cytoplasm. Moreover, sporadic episodes of neuronal degeneration were evident. DRG of cisplatin-treated animals showed severe neuronal atrophy. Moreover bortezomib induced moderate to severe axonal degeneration of the myelinated fibers in the sciatic nerves. More severe changes were induced by paclitaxel where also areas of fibers loss were frequently observed and rare pathological abnormalities were present in unmyelinated fibers. Similar changes were evident in paclitaxel-treated mice (degeneration at different stage of severity in myelinated fibers, enlargement of Schwann cells, fibers loss and dark inclusions in the unmyelinated fibers). These schedules demonstrated to be effective in mimicking clinical features of painful neuropathies and allows to combine the study of peripheral neurotoxicity of chemotherapy drugs to their anti-tumour activity against cancers of human origin

A new animal model of chemotherapy induced peripheral neurotoxicity: the immune-deficient mouse

Cisplatin, paclitaxel and bortezomib are anticancer drugs widely employed in the treatment of different solid tumours even though peripheral neurotoxicity represents a major limitation in their clinical use. During the last decades many rat and mouse models of chronic chemotherapyinduced peripheral neurotoxicity (CIPN) have been characterized from the clinical, pathological, neurophysiological and behavioural point of view. These models were based on immune-competent animals, however in preclinical oncology immune-deficient mice are mainly used. In this respect, the development of immune-deficient mice models could represent a basis for the concurrent investigation of both anticancer drug efficacy and neurotoxicity in animals implanted with human-derived cancer. Moreover, in the same model, neuroprotective effects and non-interference with anticancer activity could be better studied. In this study we established the feasibility of new immune-deficient murine models of peripheral neurotoxicity induced by three anticancer drugs. Forty-eight athymic nude mice were randomized in 4 groups of 12 animals, three were treated respectively with cisplatin, paclitaxel and bortezomib, and one was left untreated. All animals were followed up for 6 weeks. They were examined at baseline, week 4 and 6 for neurophysiological functions and behavioural tests, whilst morphological and morphometric analysis were performed on dorsal root ganglia (DRG) and peripheral nerves collected after 4 and 6 weeks of treatment. The results of the study demonstrate that athymic nude mice show CIPN features similar to those observed in conventional models even if some differences must be remarked as the prolonged time of treatment required to develop a chronic neuropathy. The characterization of this new mice model of CIPN will allow studies of antineoplastic and neurotoxic effects in the same animal

Positive effect of Mesenchymal Stem Cells therapeutic administration on chronic Experimental Autoimmune Encephalomyelitis

Multiple Sclerosis (MS) is a crippling chronic disease of the Central Nervous System caused by the presence of self-antibodies which progressively damage axonal myelin sheath, leading to axonal transmission impairment and to the development of neurological symptoms. MS is characterized by a Relapsing-Remitting course, and current therapies rely only on the use of immunosuppressive drugs, which are however unable to reverse disease progression. Encouraging results have been obtained in preclinical studies with the administration of Mesenchymal Stem Cells (MSCs) before disease onset (Zappia et al., 2005). Here, we investigate the therapeutic potential of MSC administration after disease onset into an animal model of MS, represented by Dark Agouti rats affected by chronic Relapsing-Remitting Experimental Autoimmune Encephalomyelitis (EAE) (Cavaletti et al., 2004). 106 MSC were intravenously injected in EAE rats after disease onset. Clinical score was assessed daily, and after 45 days rats were sacrificed and histological analysis of spinal cords performed to evaluate the demyelinating lesions. After the first peak of disease, no further relapses were observed in EAE rats treated with MSCs, differently from what observed in EAE group. Histological analysis demonstrated the presence of demyelinated plaques in spinal cords of EAE rats, (Luxol fast Blue staining and anti-MBP immunohystochemistry). On the contrary the therapeutic schedule with MSCs significantly reduces the number and the extension of demyelinated areas in the spinal cords, confirming clinical score evaluations. These results demonstrated that MSCs ameliorate the clinical course of EAE and hamper the disease relapsing by reducing the areas of demyelinated lesions. Granted by MIUR – FIRB Futuro in Ricerca 2008 Prot. N° RBFR08VSVI_001

Oxaliplatin-induced peripheral neurotoxicity: morphological characterization in different mouse strains

Oxaliplatin is one of the most effective anticancer drug, particularly employed in the treatment of colorectal cancer, but one of the major limitation in its use is peripheral neurotoxicity. Oxaliplatin induced peripheral neurotoxicity (OIPN) has a high incidence and is frequently long lasting or permanent. Neuropathy is characterized by distal sensory impairment initially in the legs, then extending to the arms. A prominent manifestation of sensitive damage is ataxia. Besides chronic neurotoxicity, many patients experience an acute, rapidly developing cold-induced sensory neuropathy, usually resolving within one week. OIPN clinical manifestations reflect the involvement of dorsal root ganglia (DRG) as primary target of the drug toxicity. Although this assumption is largely accepted and some pathogenetic hypothesis have been proposed, mechanisms at the basis of OIPN need to be clearly defined. OIPN may vary in frequency and severity among different cancer patients despite equal treatment schedules. A genetic susceptibility for more severe oxaliplatin-induced peripheral neurotoxicity (OIPN) has been suggested but never confirmed. Therefore we designed a study to assess the phenotypic differences induced by oxaliplatin treatment in six different mice strains (Balb c, AJ, C57Bl6, FVB, DBA, CD1) aiming at identifying the more and less severely affected. Animals were treated with OHP 3.5 mg/Kg/iv twice weekly x 4 weeks and evaluated before and after treatment. In all strains we performed a multimodal characterization of its neurotoxicity through morphological and morphometrical assessment in caudal nerve and DRG at light and electron microscopy, intra-epidermal nerve fibers density quantification, evaluation of mechanical and cold allodynia/hypoaesteshesia, caudal and digital nerve conduction velocity, activity of wide dynamic range (WDR) neurons of the spinal dorsal horn. Our preliminary data suggest that all the strains show signs of OIPN but not the same modifications in the parameters examined. We will show these results with particular attention to morphological data. This study suggests that genetic variability might have a role in the type and severity of OHP-induced peripheral damage

Anti-tumor Efficacy Assessment of the Sigma Receptor Pan Modulator RC-106. A Promising Therapeutic Tool for Pancreatic Cancer

Introduction: Pancreatic cancer (PC) is one of the most lethal tumor worldwide, with no prognosis improvement over the past 20-years. The silent progressive nature of this neoplasia hampers the early diagnosis, and the surgical resection of the tumor, thus chemotherapy remains the only available therapeutic option. Sigma receptors (SRs) are a class of receptors proposed as new cancer therapeutic targets due to their over-expression in tumor cells and their involvement in cancer biology. The main localization of these receptors strongly suggests their potential role in ER unfolded protein response (ER-UPR), a condition frequently occurring in several pathological settings, including cancer. Our group has recently identified RC-106, a novel pan-SR modulator with good in vitro antiproliferative activities toward a panel of different cancer cell lines. In the present study, we investigated the in vitro properties and pharmacological profile of RC-106 in PC cell lines with the aim to identify a potential lead candidate for the treatment of this tumor.Methods: Pancreatic cancer cell lines Panc-1, Capan-1, and Capan-2 have been used in all experiments. S1R and TMEM97/S2R expression in PC cell lines was quantified by Real-Time qRT-PCR and Western Blot experiments. MTS assay was used to assess the antiproliferative effect of RC-106. The apoptotic properties of RC-106 was evaluated by TUNEL and caspase activation assays. GRP78/BiP, ATF4, and CHOP was quantified to evaluate ER-UPR. Proteasome activity was investigated by a specific fluorescent-based assay. Scratch wound healing assay was used to asses RC-106 effect on cell migration. In addition, we delineated the in vivo pharmacokinetic profile and pancreas distribution of RC-106 in male CD-1 mice.Results: Panc-1, Capan-1, and Capan-2 express both SRs. RC-106 exerts an antiproliferative and pro-apoptotic effect in all examined cell lines. Cells exposure to RC-106 induces the increase of the expression of ER-UPR related proteins, and the inhibition of proteasome activity. Moreover, RC-106 is able to decrease PC cell lines motility. The in vivo results show that RC-106 is more concentrated in pancreas than plasma.Conclusion: Overall, our data evidenced that the pan-SR modulator RC-106 is an optimal candidate for in vivo studies in animal models of PC

OATP1B2 deficiency protects against paclitaxel-induced neurotoxicity

Paclitaxel is among the most widely used anticancer drugs and is known to cause a dose-limiting peripheral neurotoxicity, the initiating mechanisms of which remain unknown. Here, we identified the murine solute carrier organic anion–transporting polypeptide B2 (OATP1B2) as a mediator of paclitaxel-induced neurotoxicity. Additionally, using established tests to assess acute and chronic paclitaxel-induced neurotoxicity, we found that genetic or pharmacologic knockout of OATP1B2 protected mice from mechanically induced allodynia, thermal hyperalgesia, and changes in digital maximal action potential amplitudes. The function of this transport system was inhibited by the tyrosine kinase inhibitor nilotinib through a noncompetitive mechanism, without compromising the anticancer properties of paclitaxel. Collectively, our findings reveal a pathway that explains the fundamental basis of paclitaxel-induced neurotoxicity, with potential implications for its therapeutic management

Current View in Platinum Drug Mechanisms of Peripheral Neurotoxicity

Peripheral neurotoxicity is the dose-limiting factor for clinical use of platinum derivatives, a class of anticancer drugs which includes cisplatin, carboplatin, and oxaliplatin. In particular cisplatin and oxaliplatin induce a severe peripheral neurotoxicity while carboplatin is less neurotoxic. The mechanisms proposed to explain these drugs’ neurotoxicity are dorsal root ganglia alteration, oxidative stress involvement, and mitochondrial dysfunction. Oxaliplatin also causes an acute and reversible neuropathy, supposed to be due by transient dysfunction of the voltage-gated sodium channels of sensory neurons. Recent studies suggest that individual genetic variation may play a role in the pathogenesis of platinum drug neurotoxicity. Even though all these mechanisms have been investigated, the pathogenesis is far from clearly defined. In this review we will summarize the current knowledge and the most up-to-date hypotheses on the mechanisms of platinum drug-induced peripheral neurotoxicity

Current View in Platinum Drug Mechanisms of Peripheral Neurotoxicity

Peripheral neurotoxicity is the dose-limiting factor for clinical use of platinum derivatives, a class of anticancer drugs which includes cisplatin, carboplatin, and oxaliplatin. In particular cisplatin and oxaliplatin induce a severe peripheral neurotoxicity while carboplatin is less neurotoxic. The mechanisms proposed to explain these drugs’ neurotoxicity are dorsal root ganglia alteration, oxidative stress involvement, and mitochondrial dysfunction. Oxaliplatin also causes an acute and reversible neuropathy, supposed to be due by transient dysfunction of the voltage-gated sodium channels of sensory neurons. Recent studies suggest that individual genetic variation may play a role in the pathogenesis of platinum drug neurotoxicity. Even though all these mechanisms have been investigated, the pathogenesis is far from clearly defined. In this review we will summarize the current knowledge and the most up-to-date hypotheses on the mechanisms of platinum drug-induced peripheral neurotoxicity

Addressing the Need of a Translational Approach in Peripheral Neuropathy Research: Morphology Meets Function

Peripheral neuropathies (PNs) are a type of common disease that hampers the quality of life of affected people. Treatment, in most cases, is just symptomatic and often ineffective. To improve drug discovery in this field, preclinical evidence is warranted. In vivo rodent models allow a multiparametric approach to test new therapeutic strategies, since they can allow pathogenetic and morphological studies different from the clinical setting. However, human readouts are warranted to promptly translate data from the bench to the bedside. A feasible solution would be neurophysiology, performed similarly at both sides. We describe a simple protocol that reproduces the standard clinical protocol of a neurophysiology hospital department. We devised the optimal montage for sensory and motor recordings (neurography) in mice, and we also implemented F wave testing and a short electromyography (EMG) protocol at rest. We challenged this algorithm by comparing control animals (BALB/c mice) with a model of mild neuropathy to grasp even subtle changes. The neurophysiological results were confirmed with neuropathology. The treatment group showed all expected alterations. Moreover, the neurophysiology matched the neuropathological analyses. Therefore, our protocol can be suggested to promptly translate data from the bench to the bedside and vice versa