Abstract: Patients with chronic pain syndromes, like fibromyalgia (FM) complain of widespread pain and tenderness, as well as non‐refreshing sleep, cognitive dysfunction, and negative mood. Several lines of evidence implicate abnormalities of central pain processing as contributors for chronic pain, including dysfunctional descending pain inhibition. One form of endogenous pain inhibition, diffuse noxious inhibitory controls (DNIC), has been found to be abnormal in some chronic pain patients and evidence exists for deficient spatial summation of pain, specifically in FM. Similar findings have been reported in patients with localized musculoskeletal pain (LMP) disorders, like neck and back pain. Whereas DNIC reduces pain through activation of nociceptive afferents, vibro‐tactile pain inhibition involves innocuous A‐beta fiber. To assess whether patients with localized or widespread chronic pain disorders have dysfunctional A‐beta related pain inhibition we enrolled 28 normal pain‐free controls (NC), 29 FM patients, and 19 subjects with neck or back pain. All received 10 s sensitivity‐adjusted noxious heat stimuli to the forearms as test stimuli. To assess endogenous analgesic mechanisms of study subjects, vibro‐tactile conditioning stimuli were simultaneously applied with test stimuli either homotopically or heterotopically. Additionally, the effect of distraction on experimental pain was assessed. Homotopic vibro‐tactile stimulation resulted in 40% heat pain reductions in all subject groups. Distraction did not seem to affect experimental pain ratings. Conclusions : Vibro‐tactile stimulation effectively recruited analgesic mechanisms not only in NC but also in patients with chronic musculoskeletal pain, including FM. Distraction did not seem to contribute to this analgesic effect.

Abstract: Based on these results, we propose that myo1c is not a tension-sensitive anchor, as seen for myo1b ( 3 ). Rather, myo1c likely functions as a slow transporter capable of generating power over a range of forces via a force-sensing mechanism that is unique amongst characterized myosins. These results provide an important molecular framework by which a substantial number of biological phenomena can be understood. The finding that myo1c likely functions as a transporter will require modifications to current models of myo1c function that define myo1c as a tension-sensing anchor—most notably, the model of slow adaptation in sensory hair cells ( 1 ). This important motor also functions in insulin-stimulated exocytosis of GLUT4 in fat and muscle cells ( 4 ), trafficking of intracellular membranes ( 5 ), antigen presentation at the immunological synapse, transcription of DNA in the nucleus, and membrane ruffling. The findings presented here will likely shed light on the molecular role of myo1c in these processes. These results also clearly demonstrate that a kinetic analysis alone is insufficient for determining the molecular role of myosin, and that mechanical experiments are needed to define tension-sensing behavior. This result will likely necessitate a reevaluation of the proposed molecular roles of other myosins that have been assigned functions based solely on kinetic characterizations. Lastly, our findings clearly demonstrate mechanochemical diversity within the myosin-I family, possibly shedding light on the evolutionary imperatives that resulted in the retention of eight distinct myosin-I genes in vertebrates. We measured the effect of applied load on the myosin-attachment lifetime using an isometric optical clamp. Surprisingly, we found the force dependence of the actomyosin-detachment rate of myo1c to be unlike that of any other characterized myosin ( Fig. P1 ). The actomyosin-detachment rate shows two distinct regimes: a force-independent regime (where the rate of ADP release limits actomyosin detachment) at forces 〈  1 pN, and a highly force-dependent regime at higher loads. We found that the primary force-sensitive transition is the myosin-isomerization step that follows ATP binding, not ADP release as in other myosins. These features are in contrast to those of myo1b, which has a much higher force sensitivity over the range of all forces that have been examined and employs ADP release as the primary force-sensitive transition. Thus, despite the similarities in the ATPase kinetics of myo1b and myo1c (including similar coupling constants), their mechanical properties are very different when assayed under load. Fig. P1. ( Top ) Cartoon showing the interaction of myo1c (blue) with actin (red) during its mechanochemical cycle. At forces 〈  1 pN, ADP release limits actomyosin detachment, similar to other myosins. At forces 〉  1 pN, the primary force-dependent transition (i.e., the isomerization following ATP binding) limits detachment. ( Bottom ) The rate of actomyosin detachment as a function of force measured using an isometric optical clamp. The data clearly show two regimes, one force-independent (yellow) and one force-dependent (grey), corresponding to the highlighted mechanochemical transitions above. This force-sensing behavior is unique amongst characterized myosins. We used stopped-flow kinetic techniques to measure the kinetic parameters that define the myo1c ATPase cycle, and we confirmed that myo1c is a low duty-ratio motor with slow-cycling kinetics in the absence of force. The kinetic rate constants measured for myo1c are very similar to those of the closely related myosin-I isoform, myo1b, which has been shown to be a tension sensor that transforms into an anchoring protein at low forces ( 〉  0.5 pN) ( 3 ). We also measured the thermodynamic coupling constant (i.e., the ratio of actomyosin and ADP affinities), a parameter that has been proposed to reveal the tension sensitivity of myosin isoforms ( 2 ). Confirming the results of previous studies, we found that myo1c has a small coupling constant, which is consistent with characterized tension-sensing motors ( 2 ). Based on these results, one might have expected myo1c to behave as a tension sensor under load. Myosin IC (myo1c) is a widely expressed actin-based motor associated with several important cellular functions in vertebrates. The molecular role of myo1c in cells is not well described, but it is often modeled as a tension sensor ( 1 ). Myosins that act as tension sensors have very long actin-attachment durations and low power outputs. In contrast, myosins that act as transport motors generate power and support motility over a range of forces. We employed ensemble kinetic and single-molecule techniques to determine if myo1c is in fact a tension sensor. We found that, although myo1c is biochemically similar to known tension-sensing motors in the absence of force ( 2 ), its kinetic behavior under load is more consistent with its functioning as a slow transporter that can generate power over a range of forces.

Abstract: Abstract 3747 Introduction: Ponatinib is a potent, oral, pan-BCR-ABL inhibitor with activity against native and mutant forms of BCR-ABL, including the tyrosine kinase inhibitor (TKI)-resistant T315I mutant. The efficacy and safety of ponatinib (45 mg orally QD) were evaluated in a phase 2, international, open-label clinical trial (PACE). These multivariate analyses explored the impact of dose intensity and several prognostic and predictive factors on clinical responses, adverse events (AEs), and laboratory changes. Methods: Enrolled patients were resistant or intolerant (R/I) to dasatinib or nilotinib, or had the T315I BCR-ABL mutation at baseline. A total of 267 chronic phase (CP), 83 accelerated phase (AP), and 94 blast phase (BP) CML/Ph+ ALL patients were assigned to 1 of 6 cohorts according to disease phase (CP-, AP-, or BP-CML/Ph+ ALL), R/I to dasatinib or nilotinib, and presence of T315I. Three CP-CML and 2 AP-CML patients were treated, but not assigned to a cohort (post-imatinib, did not have T315I at baseline); these patients were excluded from efficacy analyses and included in safety analyses. For the purposes of the efficacy multivariate analyses, AP-CML, BP-CML, and Ph+ ALL patients were combined. The baseline covariates analyzed were age, time since diagnosis, number of prior TKIs, presence or absence of the T315I mutation, neutrophil and platelet counts, and weight. The primary efficacy outcome analyzed was major cytogenetic response (MCyR) in CP-CML and major hematologic response (MaHR) for all other patients. The safety outcomes analyzed were the following AEs: pancreatitis, elevated lipase, alanine aminotransferase (ALT) increase, aspartate aminotransferase (AST) increase, rash, neutropenia, thrombocytopenia, arthralgia, and hypertriglyceridemia. The impact on neutrophils, platelets, bilirubin, ALT, AST, creatinine, lipase, and triglycerides was also examined. Binary event outcomes were analyzed using logistic regression models. Data values over time were analyzed using linear mixed effects models. Laboratory values were log-transformed. Data as of 27 April 2012 were used in these analyses. Results: Median baseline characteristics of the CP-CML R/I and T315I cohorts, respectively, were: 61 vs 51 yrs of age, 8 vs 5 yrs since initial diagnosis, 3 vs 2 prior TKIs. The median dose intensity for the CP-CML R/I and T315I cohorts was 30 and 39 mg/day, respectively. In general, other baseline characteristics were balanced between these 2 cohorts. Multivariate analysis found statistically significant associations between MCyR and increasing dose intensity (mg/day) (p 〈 0.0001) and decreasing age (p=0.046) in CP-CML. Despite the finding that CP-CML patients with the T315I mutation had a higher response rate than those without the T315I mutation (MCyR 70% vs 49%), presence of T315I was not a significant prognostic factor for response after adjusting for other covariates (p 〉 0.2). This was likely because patients with T315I received a greater dose intensity, were younger, and were previously treated with fewer TKIs. The probability of achieving MaHR in patients with AP-CML, BP-CML, and Ph+ ALL increased with increasing dose intensity (p 〈 0.001) and with higher numbers of baseline platelets (p=0.0046). As in CP-CML, similar trends in baseline characteristics were observed, and the presence of the T315I mutation was not a significant prognostic factor for MaHR. In all patients, the probability of AEs (pancreatitis, lipase increase, ALT and AST increase, thrombocytopenia, neutropenia, arthralgia, and rash) increased with increasing dose intensity. Hypertriglyceridemia was trend level associated with dose intensity (p=0.054). Presence of T315I was associated with a lower risk of thrombocytopenia (p 〈 0.0001) and neutropenia (p=0.005) after adjustment for dose intensity and the other factors. In general, younger age, less time since diagnosis, and fewer prior TKIs were associated with a lower probability of AEs. Conclusion: These findings suggest that dose intensity and factors related to extent of disease and prior treatment were most predictive of effectiveness and tolerance of ponatinib. T315I was not a significant prognostic factor for efficacy or safety after adjustment for other factors, with the exception of thrombocytopenia and neutropenia; patients with T315I had lower predicted rates of these AEs after adjustment for dose intensity and other factors in the reduced models. Disclosures: Off Label Use: Ponatinib - non FDA approved (experimental) compound. Cortes:Novartis, BMS, ARIAD, Pfizer, and Chemgenex: Consultancy, Research Funding. Kim:Novartis, Bristol Myers-Squibb, Pfizer, ARIAD, and Il-Yang: Consultancy, Employment, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Pinilla-Ibarz:Novartis : Research Funding, Speakers Bureau; Bristol-Myers Squibb: Research Funding, Speakers Bureau. le Coutre:Novartis and BMS: Honoraria. Paquette:ARIAD: Consultancy. Chuah:Novartis and Bristol Myers-Squibb: Honoraria. Nicolini:Novartis, Bristol Myers-Squibb, Pfizer, ARIAD, and Teva: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Apperley:Novartis, Bristol Myers-Squibb, and ARIAD: Honoraria, Research Funding. Talpaz:Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Bristol Myers-Squibb: Membership on an entity's Board of Directors or advisory committees, Research Funding; Incyte: Research Funding; Sanofi: Membership on an entity's Board of Directors or advisory committees, Research Funding; Millenium: Research Funding; Celgene: Research Funding; ARIAD: Research Funding; Deciphera: Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees. Abruzzese:Bristol Myers-Squibb and Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees. Rea:Bristol Myers-Squibb, Novartis, and Teva: Honoraria. Baccarani:ARIAD, Novartis, Bristol Myers-Squibb, and Pfizer: Consultancy, Honoraria, Speakers Bureau. Muller:ARIAD: Consultancy. Wong:MolecularMD Corp: Employment, Equity Ownership. Dorer:ARIAD: Employment, Equity Ownership. Knickerbocker:ARIAD: Employment, Equity Ownership. Rivera:ARIAD: Employment, Equity Ownership. Clackson:ARIAD: Employment, Equity Ownership. Turner:ARIAD: Employment, Equity Ownership. Haluska:ARIAD: Employment, Equity Ownership. Guilhot:ARIAD: Honoraria. Hochhaus:ARIAD, Novartis, Bristol Myers-Squibb, Pfizer, and MSD: Membership on an entity's Board of Directors or advisory committees, Research Funding. Hughes:Novartis, Bristol Myers-Squibb, and ARIAD: Honoraria, Research Funding. Goldman:Novartis, Bristol Myers-Squibb, and Amgen: Honoraria. Shah:Novartis: Consultancy; Bristol Myers-Squibb: Consultancy, Research Funding; ARIAD: Consultancy, Research Funding. Kantarjian:Pfizer: Research Funding; ARIAD: Research Funding; Bristol-Myers Squibb: Research Funding; Novartis: Consultancy, Research Funding.