Trajectory Modeling and Response Prediction in Transcranial Magnetic Stimulation for Depression.

Repetitive transcranial magnetic stimulation (rTMS) therapy could be improved by better and earlier prediction of response. Latent class mixture (LCMM) and non-linear mixed effects (NLME) modelling have been applied to model the trajectories of antidepressant response (or non-response) to TMS, but it is not known whether such models can predict clinical outcomes. We compared LCMM and NLME approaches to model the antidepressant response to TMS in a naturalistic sample of 238 patients receiving rTMS for treatment resistant depression (TRD), across multiple coils and protocols. We then compared the predictive power of those models. LCMM trajectories were influenced largely by baseline symptom severity, but baseline symptoms provided little predictive power for later antidepressant response. Rather, the optimal LCMM model was a nonlinear two-class model that accounted for baseline symptoms. This model accurately predicted patient response at 4 weeks of treatment (AUC = 0.70, 95% CI = [0.52-0.87]), but not before. NLME offered slightly improved predictive performance at 4 weeks of treatment (AUC = 0.76, 95% CI = [0.58 - 0.94], but likewise, not before. In showing the predictive validity of these approaches to model response trajectories to rTMS, we provided preliminary evidence that trajectory modeling could be used to guide future treatment decisions.

A MOZ-TIF2 leukemia mouse model displays KAT6-dependent H3K23 propionylation and overexpression of a set of active developmental genes.

Aberrant regulation of chromatin modifiers is a common occurrence across many cancer types, and a key priority is to determine how specific alterations of these proteins, often enzymes, can be targeted therapeutically. MOZ, a histone acyltransferase, is recurrently fused to coactivators CBP, p300, and TIF2 in cases of acute myeloid leukemia (AML). Using either pharmacological inhibition or targeted protein degradation in a mouse model for MOZ-TIF2-driven leukemia, we show that KAT6 (MOZ/MORF) enzymatic activity and the MOZ-TIF2 protein are necessary for indefinite proliferation in cell culture. MOZ-TIF2 directly regulates a small subset of genes encoding developmental transcription factors, augmenting their high expression. Furthermore, transcription levels in MOZ-TIF2 cells positively correlate with enrichment of histone H3 propionylation at lysine 23 (H3K23pr), a recently appreciated histone acylation associated with gene activation. Unexpectedly, we also show that MOZ-TIF2 and MLL-AF9 regulate transcription of unique gene sets, and their cellular models exhibit distinct sensitivities to multiple small-molecule inhibitors directed against AML pathways. This is despite the shared genetic pathways of wild-type MOZ and MLL. Overall, our data provide insight into how aberrant regulation of MOZ contributes to leukemogenesis. We anticipate that these experiments will inform future work identifying targeted therapies in the treatment of AML and other diseases involving MOZ-induced transcriptional dysregulation.

Mezigdomide is effective alone and in combination with menin inhibition in preclinical models of KMT2A-r and NPM1c AML.

ABSTRACT: Small molecules that target the menin-KMT2A protein-protein interaction (menin inhibitors) have recently entered clinical trials in lysine methyltransferase 2A (KMT2A or MLL1)-rearranged (KMT2A-r) and nucleophosmin-mutant (NPM1c) acute myeloid leukemia (AML) and are demonstrating encouraging results. However, rationally chosen combination therapy is needed to improve responses and prevent resistance. We have previously identified IKZF1/IKAROS as a target in KMT2A-r AML and shown in preclinical models that IKAROS protein degradation with lenalidomide or iberdomide has modest single-agent activity yet can synergize with menin inhibitors. Recently, the novel IKAROS degrader mezigdomide was developed with greatly enhanced IKAROS protein degradation. In this study, we show that mezigdomide has increased preclinical activity in vitro as a single-agent in KMT2A-r and NPM1c AML cell lines, including sensitivity in cell lines resistant to lenalidomide and iberdomide. Further, we demonstrate that mezigdomide has the greatest capacity to synergize with and induce apoptosis in combination with menin inhibitors, including in MEN1 mutant models. We show that the superior activity of mezigdomide compared with lenalidomide or iberdomide is due to its increased depth, rate, and duration of IKAROS protein degradation. Single-agent mezigdomide was efficacious in 5 patient-derived xenograft models of KMT2A-r and 1 NPM1c AML. The combination of mezigdomide with the menin inhibitor VTP-50469 increased survival and prevented and overcame MEN1 mutations that mediate resistance in patients receiving menin inhibitor monotherapy. These results support prioritization of mezigdomide for early phase clinical trials in KMT2A-r and NPM1c AML, either as a single agent or in combination with menin inhibitors.

KDM6A epigenetically regulates subtype plasticity in small cell lung cancer.

Small cell lung cancer (SCLC) exists broadly in four molecular subtypes: ASCL1, NEUROD1, POU2F3 and Inflammatory. Initially, SCLC subtypes were thought to be mutually exclusive, but recent evidence shows intra-tumoural subtype heterogeneity and plasticity between subtypes. Here, using a CRISPR-based autochthonous SCLC genetically engineered mouse model to study the consequences of KDM6A/UTX inactivation, we show that KDM6A inactivation induced plasticity from ASCL1 to NEUROD1 resulting in SCLC tumours that express both ASCL1 and NEUROD1. Mechanistically, KDM6A normally maintains an active chromatin state that favours the ASCL1 subtype with its loss decreasing H3K4me1 and increasing H3K27me3 at enhancers of neuroendocrine genes leading to a cell state that is primed for ASCL1-to-NEUROD1 subtype switching. This work identifies KDM6A as an epigenetic regulator that controls ASCL1 to NEUROD1 subtype plasticity and provides an autochthonous SCLC genetically engineered mouse model to model ASCL1 and NEUROD1 subtype heterogeneity and plasticity, which is found in 35-40% of human SCLCs.

IKAROS and MENIN coordinate therapeutically actionable leukemogenic gene expression in MLL-r acute myeloid leukemia.

Acute myeloid leukemia (AML) remains difficult to treat and requires new therapeutic approaches. Potent inhibitors of the chromatin-associated protein MENIN have recently entered human clinical trials, opening new therapeutic opportunities for some genetic subtypes of this disease. Using genome-scale functional genetic screens, we identified IKAROS (encoded by IKZF1) as an essential transcription factor in KMT2A (MLL1)-rearranged (MLL-r) AML that maintains leukemogenic gene expression while also repressing pathways for tumor suppression, immune regulation and cellular differentiation. Furthermore, IKAROS displays an unexpected functional cooperativity and extensive chromatin co-occupancy with mixed lineage leukemia (MLL)1-MENIN and the regulator MEIS1 and an extensive hematopoietic transcriptional complex involving homeobox (HOX)A10, MEIS1 and IKAROS. This dependency could be therapeutically exploited by inducing IKAROS protein degradation with immunomodulatory imide drugs (IMiDs). Finally, we demonstrate that combined IKAROS degradation and MENIN inhibition effectively disrupts leukemogenic transcriptional networks, resulting in synergistic killing of leukemia cells and providing a paradigm for improved drug targeting of transcription and an opportunity for rapid clinical translation.

MLL::AF9 degradation induces rapid changes in transcriptional elongation and subsequent loss of an active chromatin landscape.

MLL rearrangements produce fusion oncoproteins that drive leukemia development, but the direct effects of MLL-fusion inactivation remain poorly defined. We designed models with degradable MLL::AF9 where treatment with small molecules induces rapid degradation. We leveraged the kinetics of this system to identify a core subset of MLL::AF9 target genes where MLL::AF9 degradation induces changes in transcriptional elongation within 15 minutes. MLL::AF9 degradation subsequently causes loss of a transcriptionally active chromatin landscape. We used this insight to assess the effectiveness of small molecules that target members of the MLL::AF9 multiprotein complex, specifically DOT1L and MENIN. Combined DOT1L/MENIN inhibition resembles MLL::AF9 degradation, whereas single-agent treatment has more modest effects on MLL::AF9 occupancy and gene expression. Our data show that MLL::AF9 degradation leads to decreases in transcriptional elongation prior to changes in chromatin landscape at select loci and that combined inhibition of chromatin complexes releases the MLL::AF9 oncoprotein from chromatin globally.

Deep brain stimulation for psychiatric disorders: From focal brain targets to cognitive networks.

Deep brain stimulation (DBS) is a promising intervention for treatment-resistant psychiatric disorders, particularly major depressive disorder (MDD) and obsessive-compulsive disorder (OCD). Up to 90% of patients who have not recovered with therapy or medication have reported benefit from DBS in open-label studies. Response rates in randomized controlled trials (RCTs), however, have been much lower. This has been argued to arise from surgical variability between sites, and recent psychiatric DBS research has focused on refining targeting through personalized imaging. Much less attention has been given to the fact that psychiatric disorders arise from dysfunction in distributed brain networks, and that DBS likely acts by altering communication within those networks. This is in part because psychiatric DBS research relies on subjective rating scales that make it difficult to identify network biomarkers. Here, we overview recent DBS RCT results in OCD and MDD, as well as the follow-on imaging studies. We present evidence for a new approach to studying DBS' mechanisms of action, focused on measuring objective cognitive/emotional deficits that underpin these and many other mental disorders. Further, we suggest that a focus on cognition could lead to reliable network biomarkers at an electrophysiologic level, especially those related to inter-regional synchrony of the local field potential (LFP). Developing the network neuroscience of DBS has the potential to finally unlock the potential of this highly specific therapy.

Case Report of Dual-Site Neurostimulation and Chronic Recording of Cortico-Striatal Circuitry in a Patient With Treatment Refractory Obsessive Compulsive Disorder.

Psychiatric disorders are increasingly understood as dysfunctions of hyper- or hypoconnectivity in distributed brain circuits. A prototypical example is obsessive compulsive disorder (OCD), which has been repeatedly linked to hyper-connectivity of cortico-striatal-thalamo-cortical (CSTC) loops. Deep brain stimulation (DBS) and lesions of CSTC structures have shown promise for treating both OCD and related disorders involving over-expression of automatic/habitual behaviors. Physiologically, we propose that this CSTC hyper-connectivity may be reflected in high synchrony of neural firing between loop structures, which could be measured as coherent oscillations in the local field potential (LFP). Here we report the results from the pilot patient in an Early Feasibility study (https://clinicaltrials.gov/ct2/show/NCT03184454) in which we use the Medtronic Activa PC+ S device to simultaneously record and stimulate in the supplementary motor area (SMA) and ventral capsule/ventral striatum (VC/VS). We hypothesized that frequency-mismatched stimulation should disrupt coherence and reduce compulsive symptoms. The patient reported subjective improvement in OCD symptoms and showed evidence of improved cognitive control with the addition of cortical stimulation, but these changes were not reflected in primary rating scales specific to OCD and depression, or during blinded cortical stimulation. This subjective improvement was correlated with increased SMA and VC/VS coherence in the alpha, beta, and gamma bands, signals which persisted after correcting for stimulation artifacts. We discuss the implications of this research, and propose future directions for research in network modulation in OCD and more broadly across psychiatric disorders.

It's Not What You Say But How You Say It: Targeting RNA Methylation in AML.

In this month's issue of Cancer Cell, Su et al. (2020) describe two small-molecule inhibitors of the RNA demethylase FTO that demonstrate significant anti-tumor effects in various models of acute myeloid leukemia.

Using eye-tracking to parse object recognition: Priming activates primarily a parts-based but also a late-emerging features-based representation.

Biederman and Cooper (Cognitive Psychology, 23, 393-419, 1991), using parts-deleted and features-deleted stimuli, presented evidence that object priming occurs at the level of the objects' parts, but not features. A control condition confirmed that some priming was accrued by the (non-visual) object concept that the stimulus represented, but all visual-level priming appeared to be at the more global level of the parts of objects, rather than the local level of the individual features (edges, vertices). This outcome has long been viewed as an important piece of supporting evidence for the existence of structural descriptions (e.g., Biederman, Psychological Review, 94, 115, 1987). The original report used a naming response task, and concluded that stimuli presenting half of the parts of an object primed only those parts, whereas half-features-deleted stimuli primed both themselves and the "complementary" half, containing the features deleted from the first image, equally. The current study adapts an eye-tracking approach to enable examination of the time course of priming across an exposure to both the primed image and unprimed competitors. Parts-deleted images primed themselves quickly and exclusively, replicating the finding of Biederman and Cooper (1991). Features-deleted images showed a deviation across time, however; initially a features-deleted prime attracted looking to itself and to its complement equally, but later on, looking to the target deviated upward, demonstrating an ability to distinguish between the two versions. The outcome of the present tests provide support for the primacy of a structural parts description, while also demonstrating the existence of multiple types of representations, both global and local.

A Menin-MLL Inhibitor Induces Specific Chromatin Changes and Eradicates Disease in Models of MLL-Rearranged Leukemia.

Inhibition of the Menin (MEN1) and MLL (MLL1, KMT2A) interaction is a potential therapeutic strategy for MLL-rearranged (MLL-r) leukemia. Structure-based design yielded the potent, highly selective, and orally bioavailable small-molecule inhibitor VTP50469. Cell lines carrying MLL rearrangements were selectively responsive to VTP50469. VTP50469 displaced Menin from protein complexes and inhibited chromatin occupancy of MLL at select genes. Loss of MLL binding led to changes in gene expression, differentiation, and apoptosis. Patient-derived xenograft (PDX) models derived from patients with either MLL-r acute myeloid leukemia or MLL-r acute lymphoblastic leukemia (ALL) showed dramatic reductions of leukemia burden when treated with VTP50469. Multiple mice engrafted with MLL-r ALL remained disease free for more than 1 year after treatment. These data support rapid translation of this approach to clinical trials.

How self-generated labelling shapes transfer of learning during early childhood: The role of individual differences.

Multiple factors influence imitation during toddlerhood, including task complexity, social contingency, and individual differences. We conducted a secondary data analysis of individual differences in self-generated labelling using data collected from a complex puzzle imitation task with 355 2- to 3-year-olds. This analysis indicated that toddlers' ability to label the completed puzzle (fish or boat) was associated with better imitation performance. Labelling occurs during social interactions; therefore, our second analysis tested how labelling differed as a function of the level of social scaffolding in each condition. This analysis revealed that self-generated labelling was lower when the social demonstrator was removed and the task was presented on a touchscreen. This study is one of the first to examine self-generated labelling during a complex imitation task in toddlers and increases our understanding of the complexity of memory processing needed for imitation learning. Statement of contribution What is already known on this subject? Toddlers exhibit a transfer of learning deficit from 2D media, including books, TV, and tablets. Self-generated labelling enhances children's learning, through attentional and cognitive mechanisms. Children are sensitive to reduced social cues in screen media contributing to the transfer deficit. What does this study add? Self-generated labelling is associated with better goal imitation performance. Self-generated labelling occurs more frequently under social conditions.

Objective clinical pain analysis using serum cyclooxygenase-2 and inducible nitric oxide synthase in American patients.

BACKGROUND: Pain is a multidimensional condition of multiple origins. Determining both intensity and underlying cause are critical for effective management. Utilization of painkillers does not follow any guidelines relying on biomarkers, which effectively eliminates objective treatment. The aim of this study was to evaluate the use of serum cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) as pain biomarkers. This work could significantly advance the diagnosis and treatment of pain. METHODS: We assessed the potential utility of serum COX-2 and iNOS as objective measures of pain in a sample of American patients. Pain was scaled between level 0-5 in accordance with the level reported by the patients. Blood samples were collected from 102 patients in the emergency room. Sandwich ELISA was used to determine the COX-2 and iNOS levels in the blood serum while statistical analysis was performed using Pearson product-moment correlation coefficients, Regression and Receiver Operating Characteristics (ROC) analyses. The biomarker results were also compared with self-reports of pain by the patients using conventional pain ratings and patients were asked to report the cause of the pain. Pain levels were clustered into four groups as 0 [self-reported 0], 1 [self-reported as 1], 2 [self-reported as 2 and 3] and 3 [self-reported as 4 and 5]. Co-expression of COX-2 and iNOS could significantly alter pain development and its sensitization. Therefore, iNOS dependent COX-2 levels were employed as categorized level. RESULTS: Self-reported pain levels did not show a correlation with the serum level of COX-2 and iNOS. The lack of correlation is attributed to multiple reasons including patients' intake of painkillers prior to participation, painkiller intake habit, chronic diseases, and subjectivity of self-reported pain. Increased serum COX-2 levels were reported in relation to the subtypes of these health issues. Further, 83% of the patients who reported pain also showed the presence of COX-2 in serum, while only 53% of the patients showed the presence of iNOS in serum. Moderate relation was found between the clustered pain level and categorized COX-2 and iNOS- levels. CONCLUSIONS: The findings support the requirement of further studies to use COX-2 and iNOS as prognostic biomarkers for objective quantification of pain at the clinical level.

Functional and Structural Characterization of a (+)-Limonene Synthase from Citrus sinensis.

Terpenes make up the largest and most diverse class of natural compounds and have important commercial and medical applications. Limonene is a cyclic monoterpene (C10) present in nature as two enantiomers, (+) and (-), which are produced by different enzymes. The mechanism of production of the (-)-enantiomer has been studied in great detail, but to understand how enantiomeric selectivity is achieved in this class of enzymes, it is important to develop a thorough biochemical description of enzymes that generate (+)-limonene, as well. Here we report the first cloning and biochemical characterization of a (+)-limonene synthase from navel orange (Citrus sinensis). The enzyme obeys classical Michaelis-Menten kinetics and produces exclusively the (+)-enantiomer. We have determined the crystal structure of the apoprotein in an "open" conformation at 2.3 Å resolution. Comparison with the structure of (-)-limonene synthase (Mentha spicata), which is representative of a fully closed conformation (Protein Data Bank entry 2ONG ), reveals that the short H-α1 helix moves nearly 5 Å inward upon substrate binding, and a conserved Tyr flips to point its hydroxyl group into the active site.

Loss of RasGAP Tumor Suppressors Underlies the Aggressive Nature of Luminal B Breast Cancers.

Luminal breast cancers are typically estrogen receptor-positive and generally have the best prognosis. However, a subset of luminal tumors, namely luminal B cancers, frequently metastasize and recur. Unfortunately, the causal events that drive their progression are unknown, and therefore it is difficult to identify individuals who are likely to relapse and should receive escalated treatment. Here, we identify a bifunctional RasGAP tumor suppressor whose expression is lost in almost 50% of luminal B tumors. Moreover, we show that two RasGAP genes are concomitantly suppressed in the most aggressive luminal malignancies. Importantly, these genes cooperatively regulate two major oncogenic pathways, RAS and NF-κB, through distinct domains, and when inactivated drive the metastasis of luminal tumors in vivo Finally, although the cooperative effects on RAS drive invasion, NF-κB activation triggers epithelial-to-mesenchymal transition and is required for metastasis. Collectively, these studies reveal important mechanistic insight into the pathogenesis of luminal B tumors and provide functionally relevant prognostic biomarkers that may guide treatment decisions. SIGNIFICANCE: The lack of insight into mechanisms that underlie the aggressive behavior of luminal B breast cancers impairs treatment decisions and therapeutic advances. Here, we show that two RasGAP tumor suppressors are concomitantly suppressed in aggressive luminal B tumors and demonstrate that they drive metastasis by activating RAS and NF-κB. Cancer Discov; 7(2); 202-17. ©2016 AACR.See related commentary by Sears and Gray, p. 131This article is highlighted in the In This Issue feature, p. 115.

The persistence of symptom burden: symptom experience and quality of life of cancer patients across one year.

PURPOSE: The purpose of this longitudinal study was to track the symptom experience in a sample of cancer patients, determine the persistence of cancer symptoms and symptom burden, and examine the relationship between symptoms and QOL over time. METHODS: Five hundred forty-two patients provided longitudinal data, completing surveys over a 12-month period. Patients had breast, colorectal, gynecologic, lung, or prostate cancer with stage 1, 2, or 3 disease. Surveys included the Memorial Symptom Assessment Scale and the Functional Assessment of Cancer Therapy-General Scale and were administered every 3 months. Demographic and clinical information and comorbidities were collected from the tumor registry. RESULTS: The number and type of symptoms experienced by patients varied by cancer type, but about 90% of patients reported one or more symptoms--with prostate cancer patients reporting fewer symptoms and colorectal patients, more symptoms. Prostate patients also had the lowest symptom burden at every time point. Overall, symptom burden decreased over time, as did the Physical subscale for the MSAS. Quality of life was stable over time, except for physical well-being, which improved. Quality of life was negatively correlated with symptom burden at every time point. CONCLUSIONS: The differences in symptom experience by cancer type suggest that assessment and management of symptoms must be individually tailored or at least adjusted by cancer type. While symptom burden decreased over time, residual symptom burden was still noteworthy. As quality of life was persistently negatively correlated with symptom burden, the results suggest the need for comprehensive symptom assessment and management.

Patient and provider concordance on symptoms during the oncology outpatient clinic visit.

BACKGROUND: Cancer patients experience multiple symptoms, with specific symptoms varying by cancer type. Problems in communication between patients and health care providers (HCPs) can interfere with effective symptom assessment and management. OBJECTIVE: To address gaps in previous research by prospectively examining concordance between HCPs and patients on identifying patients' symptoms by using an identical tool for patients and HCPs at the time of the oncology clinic visit. METHODS: 94 patients completed measures of symptom experience and medical comorbidities before seeing their oncology medical team. HCPs were informed of a patient's participation in the study before seeing the patient in clinic. Immediately after the clinic visit, HCPs completed a symptom survey in which they noted the patient's symptoms. RESULTS: Patients reported more symptoms than the HCPs endorsed. The highest level of concordance for any symptom fell in the moderate agreement range. Kappa values reflecting concordance between patients and HCPs were not significantly different between the various patient-HCP pairs. No demographic or clinical variables for patients were found to be statistically related to the level of agreement on patients' symptoms. LIMITATIONS: The use of a small convenience sample size drawn from 3 specialty oncology outpatient clinics may limit the generalizability of the results to other types of cancer. The distribution of cancer stage was weighted toward stages III and IV, likely contributing to the number of symptoms. CONCLUSIONS: The level of agreement between HCPs and oncology patients on patient symptoms is weak. Concordance levels were similar, regardless of the type of HCP.

Inhibition of platelet activation by lachrymatory factor synthase (LFS)-silenced (tearless) onion juice.

Onion and garlic are renowned for their roles as functional foods. The health benefits of garlic are attributed to di-2-propenyl thiosulfinate (allicin), a sulfur compound found in disrupted garlic but not found in disrupted onion. Recently, onions have been grown with repressed lachrymatory factor synthase (LFS) activity, which causes these onions to produce increased amounts of di-1-propenyl thiosulfinate, an isomer of allicin. This investigation into the key health attributes of LFS-silenced (tearless) onions demonstrates that they have some attributes more similar to garlic and that this is likely due to the production of novel thiosulfinate or metabolites. The key finding was that collagen-induced in vitro platelet aggregation was significantly reduced by tearless onion extract over normal onion extract. Thiosulfinate or derived compounds were shown not to be responsible for the observed changes in the inflammatory response of AGS (stomach adenocarcinoma) cells to tumor necrosis factor alpha (TNFα) when pretreated with model onion juices. A preliminary rat feeding trial indicated that the tearless onions may also play a key role in reducing weight gain.

The RasGAP gene, RASAL2, is a tumor and metastasis suppressor.

RAS genes are commonly mutated in cancer; however, RAS mutations are rare in breast cancer, despite frequent hyperactivation of Ras and ERK. Here, we report that the RasGAP gene, RASAL2, functions as a tumor and metastasis suppressor. RASAL2 is mutated or suppressed in human breast cancer, and RASAL2 ablation promotes tumor growth, progression, and metastasis in mouse models. In human breast cancer, RASAL2 loss is associated with metastatic disease; low RASAL2 levels correlate with recurrence of luminal B tumors; and RASAL2 ablation promotes metastasis of luminal mouse tumors. Additional data reveal a broader role for RASAL2 inactivation in other tumor types. These studies highlight the expanding role of RasGAPs and reveal an alternative mechanism of activating Ras in cancer.