[Treatment status of tyrosine kinase inhibitor for newly-diagnosed chronic myeloid leukemia: a domestic multi-centre retrospective real-world study].

Objective: To retrospectively analyze the treatment status of tyrosine kinase inhibitors (TKI) in newly diagnosed patients with chronic myeloid leukemia (CML) in China. Methods: Data of chronic phase (CP) and accelerated phase (AP) CML patients diagnosed from January 2006 to December 2022 from 77 centers, ≥18 years old, and receiving initial imatinib, nilotinib, dasatinib or flumatinib-therapy within 6 months after diagnosis in China with complete data were retrospectively interrogated. The choice of initial TKI, current TKI medications, treatment switch and reasons, treatment responses and outcomes as well as the variables associated with them were analyzed. Results: 6 893 patients in CP (n=6 453, 93.6%) or AP (n=440, 6.4%) receiving initial imatinib (n=4 906, 71.2%), nilotinib (n=1 157, 16.8%), dasatinib (n=298, 4.3%) or flumatinib (n=532, 7.2%) -therapy. With the median follow-up of 43 (IQR 22-75) months, 1 581 (22.9%) patients switched TKI due to resistance (n=1 055, 15.3%), intolerance (n=248, 3.6%), pursuit of better efficacy (n=168, 2.4%), economic or other reasons (n=110, 1.6%). The frequency of switching TKI in AP patients was significantly-higher than that in CP patients (44.1% vs 21.5%, P<0.001), and more AP patients switched TKI due to resistance than CP patients (75.3% vs 66.1%, P=0.011). Multi-variable analyses showed that male, lower HGB concentration and ELTS intermediate/high-risk cohort were associated with lower cytogenetic and molecular responses rate and poor outcomes in CP patients; higher WBC count and initial the second-generation TKI treatment, the higher response rates; Ph(+) ACA at diagnosis, poor PFS. However, Sokal intermediate/high-risk cohort was only significantly-associated with lower CCyR and MMR rates and the poor PFS. Lower HGB concentration and larger spleen size were significantly-associated with the lower cytogenetic and molecular response rates in AP patients; initial the second-generation TKI treatment, the higher treatment response rates; lower PLT count, higher blasts and Ph(+) ACA, poorer TFS; Ph(+) ACA, poorer OS. Conclusion: At present, the vast majority of newly-diagnosed CML-CP or AP patients could benefit from TKI treatment in the long term with the good treatment responses and survival outcomes.

[To compare the efficacy and incidence of severe hematological adverse events of flumatinib and imatinib in patients newly diagnosed with chronic phase chronic myeloid leukemia].

Objective: To analyze and compare therapy responses, outcomes, and incidence of severe hematologic adverse events of flumatinib and imatinib in patients newly diagnosed with chronic phase chronic myeloid leukemia (CML) . Methods: Data of patients with chronic phase CML diagnosed between January 2006 and November 2022 from 76 centers, aged ≥18 years, and received initial flumatinib or imatinib therapy within 6 months after diagnosis in China were retrospectively interrogated. Propensity score matching (PSM) analysis was performed to reduce the bias of the initial TKI selection, and the therapy responses and outcomes of patients receiving initial flumatinib or imatinib therapy were compared. Results: A total of 4 833 adult patients with CML receiving initial imatinib (n=4 380) or flumatinib (n=453) therapy were included in the study. In the imatinib cohort, the median follow-up time was 54 [interquartile range (IQR), 31-85] months, and the 7-year cumulative incidences of CCyR, MMR, MR(4), and MR(4.5) were 95.2%, 88.4%, 78.3%, and 63.0%, respectively. The 7-year FFS, PFS, and OS rates were 71.8%, 93.0%, and 96.9%, respectively. With the median follow-up of 18 (IQR, 13-25) months in the flumatinib cohort, the 2-year cumulative incidences of CCyR, MMR, MR(4), and MR(4.5) were 95.4%, 86.5%, 58.4%, and 46.6%, respectively. The 2-year FFS, PFS, and OS rates were 80.1%, 95.0%, and 99.5%, respectively. The PSM analysis indicated that patients receiving initial flumatinib therapy had significantly higher cumulative incidences of CCyR, MMR, MR(4), and MR(4.5) and higher probabilities of FFS than those receiving the initial imatinib therapy (all P<0.001), whereas the PFS (P=0.230) and OS (P=0.268) were comparable between the two cohorts. The incidence of severe hematologic adverse events (grade≥Ⅲ) was comparable in the two cohorts. Conclusion: Patients receiving initial flumatinib therapy had higher cumulative incidences of therapy responses and higher probability of FFS than those receiving initial imatinib therapy, whereas the incidence of severe hematologic adverse events was comparable between the two cohorts.

[Effect of imatinib on the height of children with chronic myeloid leukemia in the chronic phase].

Objective: To evaluate the effect of imatinib on growth impairment in children with chronic myeloid leukemia (CML-CP) in the chronic phase. Methods: From July 2018 to July 2019, questionnaires were distributed to CML children aged <18 years at the time of diagnosis who were receiving imatinib for at least 3 months or to their parents in China. The height-for-age standard deviation score (HtSDS) and the difference of standard deviation integral (△HtSDS) were used to explore the change in height with imatinib therapy. Results: The data of 238 respondents were included; 138 (58.0% ) respondents were men. The median age at the first diagnosis of CML was 11.0 years (range, 1.4-17.9 years) , and 93 (39.0% ) respondents were at the prepuberty stage. At the time of completing the questionnaires, the median age was 15.0 years (range, 2.0-34.0 years) . The median duration of imatinib therapy was 28 months (range, 3-213 months) . Among all the respondents, the mean HtSDS when completing the questionnaires (-0.063±1.361) was significantly lower than that at the time of starting imatinib treatment (0.391±1.244) (P<0.001) . Total 71.0% respondents showed growth impairment that was more common in those starting imatinib therapy at prepubertal age than in those starting at pubertal age. Multivariate analysis showed that younger at the start of imatinib therapy (P<0.001) and longer duration of imatinib therapy (P<0.001) were significantly associated with severe growth impairment on imatinib therapy. Conclusions: Imatinib induced growth impairment in children with CML-CP. Younger the age of initiation and longer the duration of imatinib therapy, more obvious the effect of imatinib on growth impairment.

Endothelial cells are required for the cAMP regulation of cardiac contractile proteins.

The contractile proteins in mammalian cardiac muscle are regulated by a cAMP-dependent reaction that alters the activity of the actomyosin ATPase. The ATPase activity of cardiac actomyosin has also been shown to depend on factors released by small arteries in the myocardial tissue. Endothelial cells have been implicated in the regulation of the contractile force developed by isolated cardiac tissue. To determine whether endothelial cells are required for the cAMP-dependent regulation of the contractile proteins, the effect of cAMP on the actomyosin ATPase activity was measured in cryostatic sections of isolated, quickly frozen rat ventricular trabeculae. In half of the trabeculae, the endocardial endothelial cells had been damaged by a 1-sec exposure to 0.5% Triton X-100. In trabeculae with intact endothelial cells, cAMP increased actomyosin ATPase activity toward an apparently maximum value. In trabeculae with damaged endothelial cells, cAMP did not change actomyosin ATPase activity. The coronary venous effluent from an isolated heart has already been shown to modify the maximum isometric force developed by an isolated trabecula. The extent to which the force of the isolated trabecula is changed by the coronary venous effluent is closely related to the degree to which cAMP has up-regulated the actomyosin ATPase activity in the isolated heart donating the coronary effluent: the greater the degree of up-regulation of ATPase activity, the greater the increase in force produced by the effluent. These results indicate that endothelial cells are required for the cAMP-dependent regulation of cardiac contractile proteins to function, and these results further suggest that the myocardium autoregulates by modulating the cAMP regulation of contractile proteins with endothelial-derived factors.

A physiological basis for variation in the contractile properties of isolated rat heart.

1. The maximum Ca(2+)-activated force, maximum velocity of unloaded shortening and both Ca(2+)- and actin-activated ATPase activities of myosin have been measured in detergent-skinned preparations of isolated bundles of rat right ventricle after exposure of the intact tissue to different conditions of superfusion, mechanical activity and temperature. 2. Maximum Ca(2+)-activated force per unit cross-sectional area decreases with increasing cross-sectional area, and, in the absence of electrical stimulation, with the duration of superfusion. Maximum velocity of unloaded shortening is not influenced by these differences. 3. Actin-activated ATPase activity of myosin decreases as cross-sectional area increases and duration of superfusion increases, but the extent of the decrease in enzymatic activity is less than that of developed force. Ca(2+)-activated ATPase activity is independent of these differences. 4. Actin-activated ATPase activity in cryostatic sections of quickly frozen tissue is not uniform across the transverse section. In thin bundles, it is highest in the centre and lowest at the edge of the section, which correspond, respectively, to the centre and the surface of the tissue bundle. Exposure of the tissue section to 1 microM-cyclic AMP increases the actin-activated ATPase activity of myosin with the largest increase in activity occurring at or near the surface of the bundle. 5. Ca(2+)-activated ATPase activity of myosin is uniform across the transverse section and is not changed by cyclic AMP. 6. Electrical stimulation, elevated Ca2+ concentration in the superfusion medium, or isoprenaline partially or completely reverse the decline in maximum Ca(2+)-activated force produced by prolonged superfusion of the bundle before its skinning. 7. These observations are similar in many ways to those made on frog skeletal muscles by Elzinga, Howarth, Rull, Wilson & Woledge (1989a). An explanation based on the existence of a physiological mechanism for regulating the properties of force generators is proposed. Regulation of the attachment of the cross-bridge to an actin filament may be the basis for the regulatory mechanism.

A calcium independent on-off switch for cardiac force generators.

Developed force and ATPase activity of actomyosin in cardiac muscle are regulated not only by the concentration of cytosolic calcium, but also by the state of the contractile proteins. In this study, it has been shown that cardiac actomyosin ATPase, even in the presence of adequate Ca, can exist in an inactive state. Micromolar cyclic AMP activates the ATPase, inducing substantial enzymatic activity. Both active and inactive forms of myosin can co-exist in the same cells. Mammalian hearts appear to contain a physiological mechanism for altering the response of actomyosin to optimal concentrations of Ca.

Ca-independent regulation of cardiac myosin.

Calcium-independent regulation of the contractile proteins of cardiac muscle has been studied using hyperpermeable cells from rat ventricles and sections of quickly frozen rat hearts. These preparations have been used to study maximum Ca-activated force, myosin ATPase activity and the maximum velocity of unloaded shortening. Beta adrenergic activity increases the amount of force and the ATPase activity in accordance with the concentration of the V1 isozyme of myosin. V3 activity is decreased at the same time. In tissues containing only V1, there is no change in maximum velocity in response to beta adrenergic stimulation. These results indicate that beta adrenergic stimulation recruits V1 force generators and probably regulates a transition between a Ca unresponsive and a Ca responsive force generator. A 21,000 dalton protein that reproduces the effect of beta adrenergic stimulation on myosin has been isolated.

Beta-adrenergic regulation of cardiac myosin.

Calcium-independent regulation of the contractile proteins of cardiac muscle has been studied using hyperpermeable cells from rat ventricles and sections of quickly frozen rat hearts. These preparations have been used to study maximum calcium-activated force, myosin ATPase activity, and the maximum velocity of unloaded shortening. Beta-adrenergic activity increases the amount of force and the ATPase activity in accordance with the concentration of the V1 isozyme of myosin. V3 activity is decreased at the same time. In tissues containing only V1, there is no change in maximum velocity in response to beta-adrenergic stimulation. These results indicate that beta-adrenergic stimulation recruits V1 force generators and probably regulates a transition between a calcium unresponsive and a calcium responsive force generator.

Ca-independent regulation of cardiac myosin.

Calcium-independent regulation of the contractile proteins of cardiac muscle has been studied using hyperpermeable cells from rat ventricles and sections of quickly-frozen rat hearts. These preparations have been used to study maximum Ca-activated force, myosin ATPase activity and the maximum velocity of unloaded shortening. Beta adrenergic activity increases the amount of force and the ATPase activity in accordance with the concentration of the V1 isozyme of myosin. V3 activity is decreased at the same time. In tissues containing only V1, there is no change in maximum velocity in response to beta adrenergic stimulation. These results indicate that beta adrenergic stimulation recruits V1 force generators and probably regulates the transition between a Ca unresponsive and a Ca responsive force generator. This type of regulation provides the cell with the ability to operate along many different force-velocity relations.

Adrenergic regulation of myosin adenosine triphosphatase activity.

The amount of inorganic phosphate liberated by the adenosine triphosphatase activity of myosin in a thin section of cardiac tissue can be measured quantitatively by precipitation with calcium in an alkaline medium under a defined set of conditions. Specificity of the procedure for myosin adenosine triphosphatase has been confirmed by the response to inhibitors and to different degrees of contractile filament overlap. Precise quantitation of adenosine triphosphatase activity has been demonstrated by (1) constant rate over time, (2) linearity with amount of enzyme, (3) correct values for the Km of adenosine triphosphate, and (4) a similar value for Vmax to those determined by more traditional procedures. Stimulation of the beta-adrenergic system by the release of catecholamines following injection of the animal with 6-hydroxydopamine causes a rise and then a fall of both calcium- and actin-activated adenosine triphosphatase in parallel with the changes in blood levels of the transmitter. Tyramine injection of rats produces a dose related increase in myosin adenosine triphosphatase. Perfusion of isolated hearts with isoproterenol increases myosin adenosine triphosphatase in dose-related manner. Addition of cyclic adenosine monophosphate and phosphodiesterase inhibitor to the solution bathing frozen, dried sections of heart increases both calcium- and actin-activated adenosine triphosphatase activity by almost 150%. The data show that the beta-adrenergic system, through cyclic adenosine monophosphatate, regulates the enzymatic activity of myosin, independent of the concentration of calcium. The possible role of this regulatory mechanism in the physiological modulation of cardiac contractility is discussed.

The properties of cardiac contractile proteins are modulated by autonomic innervation.

The force generators in myocardial cells of rats and rabbits can exist in three different states: relaxed and calcium unresponsive; relaxed and calcium responsive; and contracted. The transition between the two calcium responsive states is produced by the abrupt rise in the concentration of calcium ions during activation by depolarization of the surface membrane. The transition between the two relaxed states is controlled by the beta-adrenergic system. Stimulation converts calcium unresponsive to calcium responsive force generators by causing the release of a 21,000 dalton regulatory factor from intracellular sites on a membrane. The factor then interacts with myosin. The regulatory system can distinguish between V1 and V3 myosins, producing a calcium responsive state in the former and a calcium unresponsive state in the latter. The result of beta-adrenergic activity is, therefore, an increase in the faster and a decrease in the slower force generators. As a result individual cardiac cells can have many different force-velocity relations.

31P-nuclear magnetic resonance analysis of perfused single frog skins.

Perfusion of single frog skins has produced stability of the 31P-nuclear magnetic resonance (NMR) spectra over periods as long as 8 h at room temperature. With this approach, relatively large phosphocreatine (PCr) signals were recorded for each of the 12 frog skins studied. The ratio of the concentration of PCr to ATP was estimated to be 0.76 +/- 0.07, a value an order of magnitude larger than that previously reported. Comparison of the ratio of the intracellular concentrations of inorganic phosphate (Pi) to ATP determined in the present NMR study with that previously estimated by chemical analysis suggests that little intracellular Pi is likely to be immobilized. Inclusion of methylphosphonate in the perfusing solutions permitted simultaneous determination of extracellular and intracellular pH from the NMR spectra alone. The methylphosphonate has been found to be nontoxic. At an extracellular pH of 7.72 +/- 0.04, the intracellular pH was 7.47 +/- 0.06.

Intracellular pH of perfused single frog skin: combined 19F- and 31P-NMR analysis.

Intracellular pH (pHc) has been determined in frog skin by applying two different methods of pH measurement, 19F and 31P nuclear magnetic resonance (NMR) analysis, to the same tissues. Results from both NMR approaches confirm an observation by Lin, Shporer, and Civan [Am. J. Physiol. 248 (Cell Physiol. 17): 1985] that acidification of the extracellular medium reverses the sign of the pH gradient present under baseline conditions. The fluorinated probe, alpha-(difluoromethyl)-alanine methyl ester, was introduced into the epithelial cells by preincubating skins for 4.7-10.4 h at room temperature in Ringer solutions containing 1 mM ester. The free amino acid was subsequently released by intracellular esterase activity, thus providing a high enough probe concentration for NMR analysis to be practicable. From measurements of short-circuit current and transepithelial resistance under base-line and experimental conditions and the appearance of phosphocreatine (PCr) in the 31P spectrum of preloaded tissues, the fluorinated probe appears to be nontoxic to frog skin. Measurement of the chemical shift of methylphosphonate relative to PCr permitted calculation of extracellular pH. Estimation of the intracellular pH was performed both by measurement of the chemical shift of inorganic phosphate (Pi) relative to PCr and by measurement of the central peak spacing of the 19F spectrum. From four direct comparisons of the two techniques in two experiments, the difference in the estimated pH was only 0.03 +/- 0.07 pH units, supporting the concept that 31P-NMR analysis is a valid method of measuring pH in this tissue.

Cyclic AMP regulation of myosin isozymes in mammalian cardiac muscle.

Hyperpermeable cells from rat heart contain a cAMP-dependent system that can increase the maximum Ca-activated force (contractility) of the contractile proteins. In two different conditions where the relative concentration of the myosin isozymes changes, i.e., hypothyroidism and aging, the size of the increase in contractility from activation of the cAMP-regulated system varies closely with the relative concentration of V1, the isozyme of myosin with the greatest Ca- and actin-activated ATPase activity. The existence of another system for the regulation of the slow isozyme V3 has been demonstrated, and it may be inhibited by beta-adrenergic activity. The possibility of cAMP-dependent myosin regulation of contraction in addition to Ca regulation of troponin is considered. Phosphorylation of the contractile proteins themselves is not required for the increased contractility.

Regulation of calcium sensitivity in perforated mammalian cardiac cells.

Sarcolemmal perforations can be produced in bundles of rat right ventricular cells by either perfusion of the heart or soaking of the bundles with a solution containing 10 mM EGTA. All cells are affected and lose approximately 40% of the surface membrane. In these cells it is possible to show cAMP regulation of contractility (maximum Ca-activated force) without cAMP regulation of Ca sensitivity (pCa for 50% of maximum Ca-activated force). Therefore, the target molecule for cAMP is different for the two regulatory systems. Both regulatory systems can be slowly washed out of the cell by 10 mM EGTA solution but not by relaxing or contraction solutions. A model for regulation of Ca sensitivity is proposed.

Regulation of cardiac contractile proteins by phosphorylation.

Several of the contractile proteins of the heart can be phosphorylated, but in studies with isolated proteins only phosphorylation of the inhibitory subunit of troponin (TnI) produces a major change in the properties of the contractile system. As TnI is phosphorylated, the concentration of calcium required for activation of contraction is increased. Phosphorylation of the tropomyosin-binding subunit of troponin (TnT) or of the light chain of myosin fails to change ATPase activity of the isolated protein system. Phosphorylation of TnI is stimulated by the beta-adrenergic system and inhibited by the cholinergic system. Maximum calcium-activated force produced by the contractile system can be increased in hyperpermeable cardiac cells by cyclic AmP (cAMP) or agents that stimulate cAMP synthesis. This change in the contractile system, which appears to be part of the physiological response to beta-adrenergic stimulation, is mediated by phosphorylation of an intermediate that then modifies the contractile system. Phosphorylation of the contractile proteins is not involved.

31P nuclear magnetic resonance analysis of frog skin.

The intracellular phosphate composition of whole and split frog skins has been studied by 31P nuclear magnetic resonance (NMR) analysis. The spectra were similar to those previously recorded from isolated epithelial cells of toad bladder. However, qualitative differences were noted in comparison with spectra from whole toad bladder. The 31P spectra from whole frog skin reflect the intracellular compositions of the epithelial cells, whereas subepithelial elements contribute significantly to the total observed 31P signals from toad bladder. Analyzed at 4 degrees C, the average phosphocreatine (PCr) and ATP concentrations of frog skin are of similar magnitude. The ratio of [PCr] to [ATP + ADP] depends on time, tissue oxygen tension, temperature, and extracellular inorganic phosphate concentration. Both this ratio and the short-circuit current (measured in parallel experiments) fell during the course of aerating frog skins in Ringer solution at room temperature. The intracellular inorganic phosphate (Pi) signal was identified. After reduction of extracellular pH, the signal did not shift immediately but subsequently did undergo an acid shift.