Association of Pembrolizumab With Tumor Response and Survival Among Patients With Advanced Melanoma.

IMPORTANCE: The programmed death 1 (PD-1) pathway limits immune responses to melanoma and can be blocked with the humanized anti-PD-1 monoclonal antibody pembrolizumab. OBJECTIVE: To characterize the association of pembrolizumab with tumor response and overall survival among patients with advanced melanoma. DESIGN, SETTINGS, AND PARTICIPANTS: Open-label, multicohort, phase 1b clinical trials (enrollment, December 2011-September 2013). Median duration of follow-up was 21 months. The study was performed in academic medical centers in Australia, Canada, France, and the United States. Eligible patients were aged 18 years and older and had advanced or metastatic melanoma. Data were pooled from 655 enrolled patients (135 from a nonrandomized cohort [n = 87 ipilimumab naive; n = 48 ipilimumab treated] and 520 from randomized cohorts [n = 226 ipilimumab naive; n = 294 ipilimumab treated]). Cutoff dates were April 18, 2014, for safety analyses and October 18, 2014, for efficacy analyses. EXPOSURES: Pembrolizumab 10 mg/kg every 2 weeks, 10 mg/kg every 3 weeks, or 2 mg/kg every 3 weeks continued until disease progression, intolerable toxicity, or investigator decision. MAIN OUTCOMES AND MEASURES: The primary end point was confirmed objective response rate (best overall response of complete response or partial response) in patients with measurable disease at baseline per independent central review. Secondary end points included toxicity, duration of response, progression-free survival, and overall survival. RESULTS: Among the 655 patients (median [range] age, 61 [18-94] years; 405 [62%] men), 581 had measurable disease at baseline. An objective response was reported in 194 of 581 patients (33% [95% CI, 30%-37%]) and in 60 of 133 treatment-naive patients (45% [95% CI, 36% to 54%]). Overall, 74% (152/205) of responses were ongoing at the time of data cutoff; 44% (90/205) of patients had response duration for at least 1 year and 79% (162/205) had response duration for at least 6 months. Twelve-month progression-free survival rates were 35% (95% CI, 31%-39%) in the total population and 52% (95% CI, 43%-60%) among treatment-naive patients. Median overall survival in the total population was 23 months (95% CI, 20-29) with a 12-month survival rate of 66% (95% CI, 62%-69%) and a 24-month survival rate of 49% (95% CI, 44%-53%). In treatment-naive patients, median overall survival was 31 months (95% CI, 24 to not reached) with a 12-month survival rate of 73% (95% CI, 65%-79%) and a 24-month survival rate of 60% (95% CI, 51%-68%). Ninety-two of 655 patients (14%) experienced at least 1 treatment-related grade 3 or 4 adverse event (AE) and 27 of 655 (4%) patients discontinued treatment because of a treatment-related AE. Treatment-related serious AEs were reported in 59 patients (9%). There were no drug-related deaths. CONCLUSIONS AND RELEVANCE: Among patients with advanced melanoma, pembrolizumab administration was associated with an overall objective response rate of 33%, 12-month progression-free survival rate of 35%, and median overall survival of 23 months; grade 3 or 4 treatment-related AEs occurred in 14%. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01295827.

Phase I Study of Pembrolizumab (MK-3475; Anti-PD-1 Monoclonal Antibody) in Patients with Advanced Solid Tumors.

PURPOSE: This phase I study evaluated the safety, maximum tolerated dose, antitumor activity, and pharmacokinetics and pharmacodynamics of pembrolizumab in patients with advanced solid tumors. EXPERIMENTAL DESIGN: In a 3 + 3 dose escalation study, 10 patients received pembrolizumab 1, 3, or 10 mg/kg intravenously every 2 weeks until progression or intolerable toxicity. Seven additional patients received 10 mg/kg every 2 weeks. Thirteen patients participated in a 3-week intrapatient dose escalation (dose range, 0.005-10 mg/kg) followed by 2 or 10 mg/kg every 3 weeks. Tumor response was assessed by Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1. RESULTS: No dose-limiting toxicities were observed. Maximum administered dose was 10 mg/kg every 2 weeks. One patient with melanoma and one with Merkel cell carcinoma experienced complete responses of 57 and 56+ weeks' duration, respectively. Three patients with melanoma experienced partial responses. Fifteen patients with various malignancies experienced stable disease. One patient died of cryptococcal infection 92 days after pembrolizumab discontinuation, following prolonged corticosteroid use for grade 2 gastritis considered drug related. Pembrolizumab exhibited pharmacokinetic characteristics typical of humanized monoclonal antibodies. Maximum serum target engagement was reached with trough levels of doses greater than or equal to 1 mg/kg every 3 weeks. Mechanism-based translational models with a focus on intratumor exposure prediction suggested robust clinical activity would be observed at doses ≥2 mg/kg every 3 weeks. CONCLUSIONS: Pembrolizumab was well tolerated and associated with durable antitumor activity in multiple solid tumors. The lowest dose with full potential for antitumor activity was 2 mg/kg every 3 weeks.

Relationship between sodium channel NaV1.3 expression and neuropathic pain behavior in rats.

A multitude of voltage-gated sodium channel subtypes (NaV1) are expressed in primary sensory neurons where they influence excitability via their role in the generation and propagation of action potentials. Peripheral nerve injury alters the expression of several NaV1subtypes, but among these only NaV1.3 is up-regulated in dorsal root ganglia (DRG) neurons. The increased expression of NaV1.3 implicates this subtype in the development and maintenance of neuropathic pain, but its contribution to neuropathic pain behavior has not been examined. Using the spared nerve injury (SNI) model, we found that peripheral nerve lesion increased NaV1.3-like immunoreactivity (-LI) in DRG neurons and that mechanical allodynia was partially alleviated following oral administration of two NaV1 blockers, mexiletine (30 and 100 mg/kg, p.o.) and lamotrigine (30 and 100 mg/kg, p.o.). Intrathecal administration of antisense oligonucleotides (4 days) selective for NaV1.3 decreased NaV1.3 immunostaining in the DRG by 50% in the SNI model, but did not attenuate mechanical or cold allodynia. Moreover, we found that only 18% of NaV1.3 positive neurons also expressed activated transcription factor-3 (ATF3), a marker of injured neurons. We then selectively axotomized a cutaneous nerve (sural) and a muscle nerve (gastrocnemius) in order to identify if NaV1.3 up-regulation is dependent on cutaneous and/or muscle afferent activation and found that the numbers of neurons expressing NaV1.3 was proportional to the magnitude of the injury, but independent of the nature of innervation. These results suggest that NaV1.3 increases in primary sensory neurons that are not directly damaged in response to injury. Thus, although NaV1.3 is up-regulated in a subpopulation of DRG neurons after injury, reduction in the expression of NaV1.3 subtype alone is not sufficient to influence the NaV1-dependent behavioral hypersensitivity associated with nerve injury.

Induction of CX3CL1 expression in astrocytes and CX3CR1 in microglia in the spinal cord of a rat model of neuropathic pain.

UNLABELLED: Previous studies have shown that chemokines might play a role in the pathology of chronic pain. The purpose of this study was to provide an immunohistochemical description of the distribution of CX3CL1 (fractalkine) and its receptor CX3CR1 in the rat spinal cord in a model of inflammatory pain induced by unilateral intraplantar complete Freund's adjuvant (CFA) and in a model of neuropathic pain induced by L5 spinal nerve ligation (modified Chung model or mSNL). In naïve rats, CX3CL1 is found in the cytoplasm of neurons as shown by colocalization of CX3XL1 and NeuN. Similar distribution of CX3CL1 was observed after CFA, whereas after mSNL, CX3CL1 was not only observed in neurons but also found in astrocytes, as shown by colocalization of CX3CL1 and GFAP. Weak immunoreactivity for the CX3CL1 receptor, CX3CR1, was found in microglia in the spinal cord of either naïve rats or rats with inflammation. However, after spinal nerve injury, CX3CR1-LI was upregulated in microglia throughout the dorsal horn. PERSPECTIVE: This study shows that spinal nerve injury, but not peripheral inflammation, induces the expression of a chemokine, CX3CL1 (fractalkine), in astrocytes and upregulates CX3CR1 in microglia in the spinal cord. This selective regulation of CX3CL1 and its receptor, CX3CR1, suggests that these chemokines may represent new targets for the treatment of neuropathic pain.

Contribution of the tetrodotoxin-resistant voltage-gated sodium channel NaV1.9 to sensory transmission and nociceptive behavior.

The transmission of pain signals after injury or inflammation depends in part on increased excitability of primary sensory neurons. Nociceptive neurons express multiple subtypes of voltage-gated sodium channels (NaV1s), each of which possesses unique features that may influence primary afferent excitability. Here, we examined the contribution of NaV1.9 to nociceptive signaling by studying the electrophysiological and behavioral phenotypes of mice with a disruption of the SCN11A gene, which encodes NaV1.9. Our results confirm that NaV1.9 underlies the persistent tetrodotoxin-resistant current in small-diameter dorsal root ganglion neurons but suggest that this current contributes little to mechanical thermal responsiveness in the absence of injury or to mechanical hypersensitivity after nerve injury or inflammation. However, the expression of NaV1.9 contributes to the persistent thermal hypersensitivity and spontaneous pain behavior after peripheral inflammation. These results suggest that inflammatory mediators modify the function of NaV1.9 to maintain inflammation-induced hyperalgesia.

Impaired neuropathic pain responses in mice lacking the chemokine receptor CCR2.

Mice lacking the chemokine receptor chemotactic cytokine receptor 2 (CCR2) have a marked attenuation of monocyte recruitment in response to various inflammatory stimuli and a reduction of inflammatory lesions in models of demyelinating disease. In the present study, we compared nociceptive responses in inflammatory and neuropathic models of pain in CCR2 knockout and wild-type mice. In acute pain tests, responses were equivalent in CCR2 knockout and wild-type mice. In models of inflammatory pain, CCR2 knockout mice showed a 70% reduction in phase 2 of the intraplantar formalin-evoked pain response but only a modest (20-30%) and nonsignificant reduction of mechanical allodynia after intraplantar Freund's adjuvant (CFA). In a model of neuropathic pain, the development of mechanical allodynia was totally abrogated in CCR2 knockout mice. CFA administration induced marked up-regulation of CCR2 mRNA in the skin and a moderate increase in the sciatic nerve and dorsal root ganglia (DRG). In response to nerve ligation, persistent and marked up-regulation of CCR2 mRNA was evident in the nerve and DRG. Disruption of Schwann cells in response to nerve lesion resulted in infiltration of CCR2-positive monocytes/macrophages not only to the neuroma but also to the DRG. Chronic pain also resulted in the appearance of activated CCR2-positive microglia in the spinal cord. Collectively, these data suggest that the recruitment and activation of macrophages and microglia peripherally and in neural tissue may contribute to both inflammatory and neuropathic pain states. Accordingly, blockade of the CCR2 receptor may provide a novel therapeutic modality for the treatment of chronic pain.

Distribution of the voltage gated sodium channel Na(v)1.3-like immunoreactivity in the adult rat central nervous system.

Voltage-gated sodium channels are transmembrane proteins responsible for the initiation and propagation of action potentials. One subtype, Na(v)1.3 (brain type III) is tetrodotoxin sensitive and fast inactivated. Na(v)1.3 has been shown to be expressed at low levels in the adult rat, but to be upregulated after sciatic nerve axotomy in the dorsal root ganglia. In the present study, we used immunohistochemistry to look at the distribution of Na(v)1.3 in the adult rat central nervous system. We used a polyclonal antibody, raised against residues 511-524. This epitope corresponds to the sequence located in the intracellular loop between domains I and II of Na(v)1.3 and is specific for this sodium channel subtype. We found Na(v)1.3-like immunoreactivity (-LI) neurons in the cerebral cortex, hippocampal formation, colliculi, and mesencephalic reticular formation. Na(v)1.3-LI was observed in fiber tracts such as the corpus callosum, anterior commissure, corticofugal fibers, lateral lemniscus, and cerebellar peduncles. Na(v)1.3-LI was particularly intense in sensory nerve tracts such as the mesencephalic trigeminal tract, vestibulospinal tract, or spinal trigeminal tract. In the spinal cord, Na(v)1.3-LI was intense throughout the white matter and the dorsal roots. In the spinal cord grey matter, Na(v)1.3-LI fibers terminate in the deep laminae of the dorsal horn and in the ventral horn. Na(v)1.3-LI was also found in motoneurons as well as in ventral roots. This study shows that Na(v)1.3 is present at the protein level in the adult rat.