Next-generation sequencing reveals a Novel NSCLC ALK F1174V mutation and confirms ALK G1202R mutation confers high-level resistance to alectinib (CH5424802/RO5424802) in ALK-rearranged NSCLC patients who progressed on crizotinib.

Acquired secondary mutations in the anaplastic lymphoma kinase (ALK) gene have been identified in ALK-rearranged (ALK+) non-small-cell lung cancer (NSCLC) patients who developed disease progression while on crizotinib treatment. Here, we identified a novel secondary acquired NSCLC ALK F1174V mutation by comprehensive next-generation sequencing in one ALK+ NSCLC patient who progressed on crizotinib after a prolonged partial response to crizotinib. In a second case, we identified a secondary acquired ALK G1202R, which also confers resistance to alectinib (CH5424802/RO5424802), a second-generation ALK inhibitor that can inhibit ALK gatekeeper L1196M mutation in vitro. ALK G1202R is located at the solvent front of the ALK kinase domain and exhibits a high level of resistance to all other ALK inhibitors currently in clinical development in vitro. Comprehensive genomic profiling of resistant tumor is increasingly important in tailoring treatment decisions after disease progression on crizotinib in ALK+ NSCLC given the promise of second-generation ALK inhibitors and other therapeutic strategies.

Heart rate decrease during crizotinib treatment and potential correlation to clinical response.

BACKGROUND: Crizotinib is used for the treatment of advanced anaplastic lymphoma kinase (ALK)-rearranged nonsmall cell lung cancer (NSCLC). Sinus bradycardia (SB) is a side effect listed in its package insert. We investigated the frequency and timing of SB, patient characteristics associated with SB during crizotinib treatment, and potential correlation between heart rate (HR) changes and clinical response to crizotinib. METHODS: A retrospective chart review was conducted of the timing and frequency of SB, patient characteristics, and clinical response of patients to crizotinib treatment. RESULTS: Forty-twp patients who had ALK-rearranged or mesenchymal epithelial transition (MET)-amplified NSCLC and received treatment with oral crizotinib 250 mg twice daily who were enrolled in 2 crizotinib trials (PROFILE 1001 and PROFILE 1005) were analyzed. There was an average decrease of 26.1 beats per minute (bpm) from the pretreatment HR among all patients during crizotinib treatment. Twenty-nine patients (69%) experienced at least 1 episode of SB (HR, <60 bpm). The average time to the lowest HR recorded was 18.6 weeks (range, 5-72 weeks). Patients who experienced SB were significantly older (aged 55.8 years vs 47.8 years; P = .0336), had a lower pretreatment HR (mean, 77.9 bpm vs 100.6 bpm; P = .002), and were on crizotinib longer (52.9 weeks vs 24.6 weeks; P = .0050) than patients who did not experience SB. The overall response rate (P = .0195) and the maximum tumor shrinkage (P = .0205) were significantly greater in patients who experienced SB. CONCLUSIONS: HR decrease is common during crizotinib treatment. It remains to be determined whether the correlation between HR decrease and clinical response to crizotinib reflects a biomarker of drug efficacy or a time/cumulative dose-dependent phenomenon.

ROS1 rearrangements define a unique molecular class of lung cancers.

PURPOSE: Chromosomal rearrangements involving the ROS1 receptor tyrosine kinase gene have recently been described in a subset of non-small-cell lung cancers (NSCLCs). Because little is known about these tumors, we examined the clinical characteristics and treatment outcomes of patients with NSCLC with ROS1 rearrangement. PATIENTS AND METHODS: Using a ROS1 fluorescent in situ hybridization (FISH) assay, we screened 1,073 patients with NSCLC and correlated ROS1 rearrangement status with clinical characteristics, overall survival, and when available, ALK rearrangement status. In vitro studies assessed the responsiveness of cells with ROS1 rearrangement to the tyrosine kinase inhibitor crizotinib. The clinical response of one patient with ROS1-rearranged NSCLC to crizotinib was investigated as part of an expanded phase I cohort. RESULTS: Of 1,073 tumors screened, 18 (1.7%) were ROS1 rearranged by FISH, and 31 (2.9%) were ALK rearranged. Compared with the ROS1-negative group, patients with ROS1 rearrangements were significantly younger and more likely to be never-smokers (each P < .001). All of the ROS1-positive tumors were adenocarcinomas, with a tendency toward higher grade. ROS1-positive and -negative groups showed no difference in overall survival. The HCC78 ROS1-rearranged NSCLC cell line and 293 cells transfected with CD74-ROS1 showed evidence of sensitivity to crizotinib. The patient treated with crizotinib showed tumor shrinkage, with a near complete response. CONCLUSION: ROS1 rearrangement defines a molecular subset of NSCLC with distinct clinical characteristics that are similar to those observed in patients with ALK-rearranged NSCLC. Crizotinib shows in vitro activity and early evidence of clinical activity in ROS1-rearranged NSCLC.

Activity of crizotinib (PF02341066), a dual mesenchymal-epithelial transition (MET) and anaplastic lymphoma kinase (ALK) inhibitor, in a non-small cell lung cancer patient with de novo MET amplification.

Crizotinib is a dual MET and ALK inhibitor. Currently, clinical development of crizotinib is focused primarily on ALK rearranged non-small cell lung cancer (NSCLC). Here we report an NSCLC patient with de novo MET amplification but no ALK rearrangement who achieved a rapid and durable response to crizotinib indicating is also a bona fide MET inhibitor.

Region specific neuron loss in the aged canine hippocampus is reduced by enrichment.

Neuron loss within the hippocampus and entorhinal cortex occurs as a function of age in humans. We first tested the hypothesis that neuron loss occurs in the aged dog. The total unilateral number of neurons in the canine entorhinal cortex and subdivisions of the hippocampus from the left hemisphere were estimated using the optical fractionator. The brains from 5 old (13.0-15.0 years old) and 5 young (3.4-4.5 years old) beagle dogs were analyzed. The hilus of the hippocampus showed a significant loss of neurons (approximately 30%) in the aged dog brain compared to young. Differences were not detected in the remaining hippocampal subfields and entorhinal cortex. We further tested the hypothesis that an antioxidant fortified food or behavioral enrichment would reduce the age-related loss of hilar neurons. Behaviorally enriched aged dogs had more neurons in the hilus (approximately 18%) compared to aged controls. These results suggest that the aged canine hippocampus in the left hemisphere shows selective neuron loss and that behavioral enrichment may reduce this loss.

Neurogenesis decreases with age in the canine hippocampus and correlates with cognitive function.

New neurons are continually produced in the adult mammalian brain from progenitor cells located in specific brain regions, including the subgranular zone (SGZ) of the dentate gyrus of the hippocampus. We hypothesized that neurogenesis occurs in the canine brain and is reduced with age. We examined neurogenesis in the hippocampus of five young and five aged animals using doublecortin (DCX) and bromodeoxyuridine (BrdU) immunostaining. The total unilateral number of new neurons in the canine SGZ and granule cell layer (GCL) was estimated using stereological techniques based upon unbiased principles of systematic uniformly random sampling. Animals received 25mg/kg of BrdU once a day for 5 days and were euthanized 9 days after the last injection. We found evidence of neurogenesis in the canine brain and that cell genesis and neurogenesis are greatly reduced in the SGZ/GCL of aged animals compared to young. We further tested the hypothesis that an antioxidant fortified food or behavioral enrichment would improve neurogenesis in the aged canine brain and neurogenesis may correlate with cognitive function. Aged animals were treated for 2.8 years and tissue was available for six that received the antioxidant food, five that received the enrichment and six receiving both treatments. There were no significant differences in the absolute number of DCX or DCX-BrdU neurons or BrdU nuclei between the treatment groups compared to control animals. The number of DCX-positive neurons and double-labeled DCX-BrdU-positive neurons, but not BrdU-positive nuclei alone, significantly correlated with performance on several cognitive tasks including spatial memory and discrimination learning. These results suggest that new neurons in the aged canine dentate gyrus may participate in modulating cognitive functions.

Neuroprotective effects of cognitive enrichment.

Cognitive enrichment early in life, as indicated by level of education, complexity of work environment or nature of leisure activities, appears to protect against the development of age-associated cognitive decline and also dementia. These effects are more robust for measures of crystallized intelligence than for measures of fluid intelligence and depend on the ability of the brain to compensate for pathological changes associated with aging. This compensatory ability is referred to as cognitive reserve. The cognitive reserve hypothesis suggests that cognitive enrichment promotes utilization of available functions. Alternatively, late life cognitive changes in cognition may be linked to a factor, such as cholinergic dysfunction, that is also present early in life and contributes to the reduced levels of early life cognitive enrichment. Beneficial effects of environmental enrichment early in life have also been observed in rodents and primates. Research with rodents indicates that these changes have structural correlates, which likely include increased synapses in specific brain regions. Dogs also show age-dependent cognitive decline, and both longitudinal and cross-sectional studies indicate that this decline can be attenuated by cognitive enrichment. Furthermore, cognitive enrichment has differential effects, improving some functions more than others. From a neurobiological perspective, behavioral enrichment in the dog may act to promote neurogenesis later in life. This can be distinguished from nutritional interventions with antioxidants, which appear to attenuate the development of neuropathology. These results suggest that a combination of behavioral and nutritional or pharmacological interventions may be optimal for reducing the rate of age-dependent cognitive decline.