Transcriptional regulators with broad expression in the zebrafish spinal cord.

BACKGROUND: The spinal cord is a crucial part of the vertebrate CNS, controlling movements and receiving and processing sensory information from the trunk and limbs. However, there is much we do not know about how this essential organ develops. Here, we describe expression of 21 transcription factors and one transcriptional regulator in zebrafish spinal cord. RESULTS: We analyzed the expression of aurkb, foxb1a, foxb1b, her8a, homeza, ivns1abpb, mybl2b, myt1a, nr2f1b, onecut1, sall1a, sall3a, sall3b, sall4, sox2, sox19b, sp8b, tsc22d1, wdhd1, zfhx3b, znf804a, and znf1032 in wild-type and MIB E3 ubiquitin protein ligase 1 zebrafish embryos. While all of these genes are broadly expressed in spinal cord, they have distinct expression patterns from one another. Some are predominantly expressed in progenitor domains, and others in subsets of post-mitotic cells. Given the conservation of spinal cord development, and the transcription factors and transcriptional regulators that orchestrate it, we expect that these genes will have similar spinal cord expression patterns in other vertebrates, including mammals and humans. CONCLUSIONS: Our data identify 22 different transcriptional regulators that are strong candidates for playing different roles in spinal cord development. For several of these genes, this is the first published description of their spinal cord expression.

Timing of high-dose methotrexate CNS prophylaxis in DLBCL: a multicenter international analysis of 1384 patients.

Prophylactic high-dose methotrexate (HD-MTX) is often used for diffuse large B-cell lymphoma (DLBCL) patients at high risk of central nervous system (CNS) relapse, despite limited evidence demonstrating efficacy or the optimal delivery method. We conducted a retrospective, international analysis of 1384 patients receiving HD-MTX CNS prophylaxis either intercalated (i-HD-MTX) (n = 749) or at the end (n = 635) of R-CHOP/R-CHOP-like therapy (EOT). There were 78 CNS relapses (3-year rate 5.7%), with no difference between i-HD-MTX and EOT: 5.7% vs 5.8%, P = .98; 3-year difference: 0.04% (-2.0% to 3.1%). Conclusions were unchanged on adjusting for baseline prognostic factors or on 6-month landmark analysis (n = 1253). In patients with a high CNS international prognostic index (n = 600), the 3-year CNS relapse rate was 9.1%, with no difference between i-HD-MTX and EOT. On multivariable analysis, increasing age and renal/adrenal involvement were the only independent risk factors for CNS relapse. Concurrent intrathecal prophylaxis was not associated with a reduction in CNS relapse. R-CHOP delays of ≥7 days were significantly increased with i-HD-MTX vs EOT, with 308 of 1573 (19.6%) i-HD-MTX treatments resulting in a delay to subsequent R-CHOP (median 8 days). Increased risk of delay occurred in older patients when delivery was later than day 10 in the R-CHOP cycle. In summary, we found no evidence that EOT delivery increases CNS relapse risk vs i-HD-MTX. Findings in high-risk subgroups were unchanged. Rates of CNS relapse in this HD-MTX-treated cohort were similar to comparable cohorts receiving infrequent CNS prophylaxis. If HD-MTX is still considered for certain high-risk patients, delivery could be deferred until R-CHOP completion.

Quantification of the effect of in utero events on lifetime resilience in dairy cows.

Currently, the dairy industry is facing many challenges that could affect its sustainability, including climate change and public perception of the industry. As a result, interest is increasing in the concept of identifying resilient animals, those with a long productive lifespan, as well as good reproductive performance and milk yield. There is much evidence that events in utero, that is, the developmental origins of health and disease hypothesis, alter the life-course health of offspring and we hypothesized that these could alter resilience in calves, where resilience is identified using lifetime data. The aim of this study was to quantify lifetime resilience scores (LRS) using an existing scoring system, based on longevity with secondary corrections for age at first calving and calving interval, and to quantify the effects of in utero events on the LRS using 2 datasets. The first was a large dataset of cattle on 83 farms in Great Britain born from 2006 to 2015 and the second was a smaller, more granular dataset of cattle born between 2003 and 2015 in the Langhill research herd at Scotland's Rural College. Events during dam's pregnancy included health events (lameness, mastitis, use of an antibiotic or anti-inflammatory medication), the effect of heat stress as measured by temperature-humidity index, and perturbations in milk yield and quality (somatic cell count, percentage fat, percentage protein and fat:protein ratio). Daughters born to dams that experienced higher temperature-humidity indexes while they were in utero during the first and third trimesters of pregnancy had lower LRS. Daughter LRS were also lower where milk yields or median fat percentages in the first trimester were low, and when milk yields were high in the third trimester. Dam LRS was positively associated with LRS of their offspring; however, as parity of the dam increased, LRS of their calves decreased. Similarly, in the Langhill herd, dams of a higher parity produced calves with lower LRS. Additionally, dams that recorded a high maximum locomotion score in the third trimester of pregnancy were negatively associated with lower calf LRS in the Langhill herd. Our results suggest that events that occur during pregnancy have lifelong consequences for the calf's lifetime performance. However, experience of higher temperature-humidity indexes, higher dam LRS, and mothers in higher parities explained a relatively small proportion of variation in offspring LRS, which suggests that other factors play a substantial role in determining calf LRS. Although "big data" can contain a considerable amount of noise, similar findings between the 2 datasets indicate it is likely these findings are real.

Central nervous system lymphomas-Assessment and treatment and prevention of central nervous system relapse.

In this review focused on lymphoma and the central nervous system (CNS), we summarize recent developments in the management of primary (PCNSL) and secondary CNS lymphoma (SCNSL), treatment of CNS lymphoma in the older population, the neuroradiological assessment of CNS lymphoma and finally highlight the ongoing debate on optimal CNS prophylaxis. The section on PCNSL focuses on the different approaches available for frontline treatment in Europe and the United States and discusses consolidation strategies. We then highlight available strategies to treat PCNSL in the elderly population, an area of unmet need. New therapies aiming at minimizing toxicity and prioritizing quality of life are emerging for these patients. Secondary CNS lymphoma, especially in the relapsed/refractory setting is another area of unmet need, and the efficacy of CAR-T cell therapy is being explored. We provide an overview of the imaging challenges in the neuroradiological assessment of CNS lymphoma. Finally, the section on CNS prophylaxis summarizes recent findings from large retrospective studies challenging the efficacy of present approaches to prophylaxis in higher-risk patients with lymphoma.

CNS prophylaxis for diffuse large B-cell lymphoma.

CNS relapse in the brain parenchyma, eyes, or leptomeninges is an uncommon but devastating complication of diffuse large B-cell lymphoma. CNS prophylaxis strategies, typically involving intrathecal or high-dose antimetabolites, have been developed in the front-line treatment setting with the aim to reduce this subsequent risk. Clinical and biological features associated with elevated risk are increasingly well defined and are discussed in this Review. This Review summarises both the historical and current developments in this challenging field, provides a nuanced discussion regarding current reasons for and against standard prophylactic measures, outlines evidence for the timing of prophylactic measures when delivered, and reflects on possible future developments.

Pirtobrutinib in relapsed or refractory B-cell malignancies (BRUIN): a phase 1/2 study.

BACKGROUND: Covalent Bruton's tyrosine kinase (BTK) inhibitors are efficacious in multiple B-cell malignancies, but patients discontinue these agents due to resistance and intolerance. We evaluated the safety and efficacy of pirtobrutinib (working name; formerly known as LOXO-305), a highly selective, reversible BTK inhibitor, in these patients. METHODS: Patients with previously treated B-cell malignancies were enrolled in a first-in-human, multicentre, open-label, phase 1/2 trial of the BTK inhibitor pirtobrutinib. The primary endpoint was the maximum tolerated dose (phase 1) and overall response rate (ORR; phase 2). This trial is registered with ClinicalTrials.gov, NCT03740529. FINDINGS: 323 patients were treated with pirtobrutinib across seven dose levels (25 mg, 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, and 300 mg once per day) with linear dose-proportional exposures. No dose-limiting toxicities were observed and the maximum tolerated dose was not reached. The recommended phase 2 dose was 200 mg daily. Adverse events in at least 10% of 323 patients were fatigue (65 [20%]), diarrhoea (55 [17%]), and contusion (42 [13%]). The most common adverse event of grade 3 or higher was neutropenia (32 [10%]). There was no correlation between pirtobrutinib exposure and the frequency of grade 3 treatment-related adverse events. Grade 3 atrial fibrillation or flutter was not observed, and grade 3 haemorrhage was observed in one patient in the setting of mechanical trauma. Five (1%) patients discontinued treatment due to a treatment-related adverse event. In 121 efficacy evaluable patients with chronic lymphocytic leukaemia (CLL) or small lymphocytic lymphoma (SLL) treated with a previous covalent BTK inhibitor (median previous lines of treatment 4), the ORR with pirtobrutinib was 62% (95% CI 53-71). The ORR was similar in CLL patients with previous covalent BTK inhibitor resistance (53 [67%] of 79), covalent BTK inhibitor intolerance (22 [52%] of 42), BTK C481-mutant (17 [71%] of 24) and BTK wild-type (43 [66%] of 65) disease. In 52 efficacy evaluable patients with mantle cell lymphoma (MCL) previously treated with covalent BTK inhibitors, the ORR was 52% (95% CI 38-66). Of 117 patients with CLL, SLL, or MCL who responded, all but eight remain progression-free to date. INTERPRETATION: Pirtobrutinib was safe and active in multiple B-cell malignancies, including patients previously treated with covalent BTK inhibitors. Pirtobrutinib might address a growing unmet need for alternative therapies for these patients. FUNDING: Loxo Oncology.

Anteroposterior patterning of the zebrafish ear through Fgf- and Hh-dependent regulation of hmx3a expression.

In the zebrafish, Fgf and Hh signalling assign anterior and posterior identity, respectively, to the poles of the developing ear. Mis-expression of fgf3 or inhibition of Hh signalling results in double-anterior ears, including ectopic expression of hmx3a. To understand how this double-anterior pattern is established, we characterised transcriptional responses in Fgf gain-of-signalling or Hh loss-of-signalling backgrounds. Mis-expression of fgf3 resulted in rapid expansion of anterior otic markers, refining over time to give the duplicated pattern. Response to Hh inhibition was very different: initial anteroposterior asymmetry was retained, with de novo duplicate expression domains appearing later. We show that Hmx3a is required for normal anterior otic patterning, and that otic patterning defects in hmx3a-/- mutants are a close phenocopy to those seen in fgf3-/- mutants. However, neither loss nor gain of hmx3a function was sufficient to generate full ear duplications. Using our data to infer a transcriptional regulatory network required for acquisition of otic anterior identity, we can recapitulate both the wild-type and the double-anterior pattern in a mathematical model.

Evolution of lbx spinal cord expression and function.

Ladybird homeobox (Lbx) transcription factors have crucial functions in muscle and nervous system development in many animals. Amniotes have two Lbx genes, but only Lbx1 is expressed in spinal cord. In contrast, teleosts have three lbx genes and we show here that zebrafish lbx1a, lbx1b, and lbx2 are expressed by distinct spinal cell types, and that lbx1a is expressed in dI4, dI5, and dI6 interneurons, as in amniotes. Our data examining lbx expression in Scyliorhinus canicula and Xenopus tropicalis suggest that the spinal interneuron expression of zebrafish lbx1a is ancestral, whereas lbx1b has acquired a new expression pattern in spinal cord progenitor cells. lbx2 spinal expression was probably acquired in the ray-finned lineage, as this gene is not expressed in the spinal cords of either amniotes or S. canicula. We also show that the spinal function of zebrafish lbx1a is conserved with mouse Lbx1. In zebrafish lbx1a mutants, there is a reduction in the number of inhibitory spinal interneurons and an increase in the number of excitatory spinal interneurons, similar to mouse Lbx1 mutants. Interestingly, the number of inhibitory spinal interneurons is also reduced in lbx1b mutants, although in this case the number of excitatory interneurons is not increased. lbx1a;lbx1b double mutants have a similar spinal interneuron phenotype to lbx1a single mutants. Taken together these data suggest that lbx1b and lbx1a may be required in succession for correct specification of dI4 and dI6 spinal interneurons, although only lbx1a is required for suppression of excitatory fates in these cells.

Ibrutinib for central nervous system lymphoma: the Australasian Lymphoma Alliance/MD Anderson Cancer Center experience.

Primary and secondary central nervous system lymphomas (PCNSL/SCNSL) are aggressive rare malignancies with dismal outcomes. Encouraging data have emerged from Phase I/II clinical trials treating relapsed/refractory PCNSL/SCNSL with ibrutinib. We analysed 33 patients who received ibrutinib, alone or with other therapies, for PCNSL (n = 9) or SCNSL (n = 24). The objective response rate was 58% (complete response 55%). The median progression-free survival and overall survival for patients with PCNSL were both 3·1 months; for SCNSL, 10·2 and 11·5 months respectively. Only one invasive fungal infection was observed, despite concurrent or recent use of dexamethasone 8-16 mg daily in 14 patients (42%). Ibrutinib has encouraging activity in these aggressive malignancies.

Prdm12 specifies V1 interneurons through cross-repressive interactions with Dbx1 and Nkx6 genes in Xenopus.

V1 interneurons are inhibitory neurons that play an essential role in vertebrate locomotion. The molecular mechanisms underlying their genesis remain, however, largely undefined. Here, we show that the transcription factor Prdm12 is selectively expressed in p1 progenitors of the hindbrain and spinal cord in the frog embryo, and that a similar restricted expression profile is observed in the nerve cord of other vertebrates as well as of the cephalochordate amphioxus. Using frog, chick and mice, we analyzed the regulation of Prdm12 and found that its expression in the caudal neural tube is dependent on retinoic acid and Pax6, and that it is restricted to p1 progenitors, due to the repressive action of Dbx1 and Nkx6-1/2 expressed in the adjacent p0 and p2 domains. Functional studies in the frog, including genome-wide identification of its targets by RNA-seq and ChIP-Seq, reveal that vertebrate Prdm12 proteins act as a general determinant of V1 cell fate, at least in part, by directly repressing Dbx1 and Nkx6 genes. This probably occurs by recruiting the methyltransferase G9a, an activity that is not displayed by the amphioxus Prdm12 protein. Together, these findings indicate that Prdm12 promotes V1 interneurons through cross-repressive interactions with Dbx1 and Nkx6 genes, and suggest that this function might have only been acquired after the split of the vertebrate and cephalochordate lineages.

Acute Renal Vein Thrombosis: A Case Report of Successful Treatment with Mechanical Thromboaspiration.

Treatment of renal vein thromboses has previously been limited to conventional anticoagulation. We describe a case of successful treatment of acute native renal vein thrombosis (RVT) in a 61-year-old fit-and-well female using mechanical thromboaspiration without the use of thrombolysis. The report demonstrates the computed tomographic appearances of an acute RVT, the venographic images taken before and after thromboaspiration, and the magnetic resonance imaging of the kidney performed 3 months later, showing complete resolution of the thrombus and normal appearances of the kidney. We believe this case to be the first published case of a safe and successful treatment of acute native RVT using the Penumbra Indigo® mechanical aspiration system, highlighting the importance of access to skilled interventional radiologists for treatment of this rare condition.

Minor change, major difference: divergent functions of highly conserved cis-regulatory elements subsequent to whole genome duplication events.

Within the vertebrate lineage, a high proportion of duplicate genes have been retained after whole genome duplication (WGD) events. It has been proposed that many of these duplicate genes became indispensable because the ancestral gene function was divided between them. In addition, novel functions may have evolved, owing to changes in cis-regulatory elements. Functional analysis of the PAX2/5/8 gene subfamily appears to support at least the first part of this hypothesis. The collective role of these genes has been widely retained, but sub-functions have been differentially partitioned between the genes in different vertebrates. Conserved non-coding elements (CNEs) represent an interesting and readily identifiable class of putative cis-regulatory elements that have been conserved from fish to mammals, an evolutionary distance of 450 million years. Within the PAX2/5/8 gene subfamily, PAX2 is associated with the highest number of CNEs. An additional WGD experienced in the teleost lineage led to two copies of pax2, each of which retained a large proportion of these CNEs. Using a reporter gene assay in zebrafish embryos, we have exploited this rich collection of regulatory elements in order to determine whether duplicate CNEs have evolved different functions. Remarkably, we find that even highly conserved sequences exhibit more functional differences than similarities. We also discover that short flanking sequences can have a profound impact on CNE function. Therefore, if CNEs are to be used as candidate enhancers for transgenic studies or for multi-species comparative analyses, it is paramount that the CNEs are accurately delineated.

Roles of Hedgehog pathway components and retinoic acid signalling in specifying zebrafish ventral spinal cord neurons.

In mouse, Hedgehog (Hh) signalling is required for most ventral spinal neurons to form. Here, we analyse the spinal cord phenotype of zebrafish maternal-zygotic smoothened (MZsmo) mutants that completely lack Hh signalling. We find that most V3 domain cells and motoneurons are lost, whereas medial floorplate still develops normally and V2, V1 and V0v cells form in normal numbers. This phenotype resembles that of mice that lack both Hh signalling and Gli repressor activity. Ventral spinal cord progenitor domain transcription factors are not expressed at 24 hpf in zebrafish MZsmo mutants. However, pMN, p2 and p1 domain markers are expressed at early somitogenesis stages in these mutants. This suggests that Gli repressor activity does not extend into zebrafish ventral spinal cord at these stages, even in the absence of Hh signalling. Consistent with this, ectopic expression of Gli3R represses ventral progenitor domain expression at these early stages and knocking down Gli repressor activity rescues later expression. We investigated whether retinoic acid (RA) signalling specifies ventral spinal neurons in the absence of Hh signalling. The results suggest that RA is required for the correct number of many different spinal neurons to form. This is probably mediated, in part, by an effect on cell proliferation. However, V0v, V1 and V2 cells are still present, even in the absence of both Hh and RA signalling. We demonstrate that Gli1 has a Hh-independent role in specifying most of the remaining motoneurons and V3 domain cells in embryos that lack Hh signalling, but removal of Gli1 activity does not affect more dorsal neurons.

The value of bispecific antibodies in relapsed and refractory DLBCL.

Diffuse large B-cell lymphoma (DLBCL) may be cured with anti-CD20 based chemoimmunotherapy in the majority of cases, however, relapsed/refractory disease occurs in 30-40% patients, and despite significant recent therapeutic advances, continues to represent an unmet clinical need. Bispecific antibodies represent a novel class of therapy currently in development for relapsed/refractory B-cell lymphoma. This review discusses the background clinical need, mechanism of action, and clinical data including efficacy and toxicity for bispecific antibodies in DLBCL, focusing on the most advanced class in development; CD20 targeting T-cell engaging antibodies. Emerging possibilities for future use of bispecific antibodies is also discussed, including novel and cytotoxic combination regimens in relapsed and first-line settings.

Footbathing and Foot Trimming, and No Quarantine: Risks for High Prevalence of Lameness in a Random Sample of 269 Sheep Flocks in England, 2022.

Since 2004, the prevalence of lameness in sheep flocks in England has reduced as farmers have adopted evidence-based management practices to control lameness. In 2011, the Farm Animal Welfare Council proposed a target prevalence of <2% lameness in sheep by 2021. This study investigated whether that target had been achieved and determined which practices were associated with prevalence of lameness. A postal questionnaire was sent to 1000 randomly selected farmers to investigate the prevalence of lameness and management practices in 2022. The geometric mean prevalence of lameness was <2% in ewes and lambs, but the median was 3%; approximately 26% flocks had <2% lameness. Data were analysed using robust variable selection with multivariable linear models. Farmers that quarantined ewes for ≥3 weeks and did not use foot bathing or foot trimming to prevent lameness had 40-50% lower prevalence of lameness than those not using these practices. Fewer farmers (19.0%) were always using parenteral antimicrobials to treat footrot, an effective practice, than in previous research (49.7%). We conclude that the target of <2% lameness in England has been achieved by 26% of farmers, and further work is required for more farmers to follow the evidence-based management practices to minimise lameness.

Transcriptional Regulators with Broad Expression in the Zebrafish Spinal Cord.

Background: The spinal cord is a crucial part of the vertebrate CNS, controlling movements and receiving and processing sensory information from the trunk and limbs. However, there is much we do not know about how this essential organ develops. Here, we describe expression of 21 transcription factors and one transcriptional regulator in zebrafish spinal cord. Results: We analyzed the expression of aurkb, foxb1a, foxb1b, her8a, homeza, ivns1abpb, mybl2b, myt1a, nr2f1b, onecut1, sall1a, sall3a, sall3b, sall4, sox2, sox19b, sp8b, tsc22d1, wdhd1, zfhx3b, znf804a, and znf1032 in wild-type and MIB E3 ubiquitin protein ligase 1 zebrafish embryos. While all of these genes are broadly expressed in spinal cord, they have distinct expression patterns from one another. Some are predominantly expressed in progenitor domains, and others in subsets of post-mitotic cells. Given the conservation of spinal cord development, and the transcription factors and transcriptional regulators that orchestrate it, we expect that these genes will have similar spinal cord expression patterns in other vertebrates, including mammals and humans. Conclusions: Our data identify 22 different transcriptional regulators that are strong candidates for playing different roles in spinal cord development. For several of these genes, this is the first published description of their spinal cord expression.

Endovascular treatment of a critical common iliac artery stenosis using intravascular ultrasound guidance in a patient with a known severe iodine contrast allergy.

Introduction: The mainstay of endovascular treatment in peripheral arterial disease is digital subtraction angiography with an iodinated contrast. In patients with a known contrast allergy, the current most commonly used alternative is to perform carbon dioxide angiography. Operator experience and availability of carbon dioxide (CO2) angiography delivery systems are, however, limited. Intravascular ultrasound is now commonly used in combination with fluoroscopy in peripheral venous interventions and in coronary arterial disease and we propose that this also provides a safe and effective alternative option to guide endovascular intervention in patients with a known contrast allergy. Case report: We present a case describing the use of intravascular ultrasound and fluoroscopy to guide endovascular treatment of a critical left common iliac artery stenosis in a patient with known anaphylaxis to iodinated contrast. Discussion: Intravascular ultrasound is fast becoming the gold standard treatment option in peripheral venous interventions; however, the use in peripheral arterial disease is limited. We believe this is the first case that reports the use of predominantly intravascular ultrasound in endovascular peripheral arterial intervention with a contrast adjunct. Conclusion: We have demonstrated that intravascular ultrasound is a safe and effective alternative or adjunct to iodinated contrast for arterial angiography and stenting in the setting of patient contra-indications.

Integration of PET in DLBCL.

F-fluorodeoxyglucose positron emission tomography-computerized tomography (18FDG-PET/CT) is the gold-standard imaging modality for staging and response assessment for most lymphomas. This review focuses on the utility of 18FDG-PET/CT, and its role in staging, prognostication and response assessment in diffuse large B-cell lymphoma (DLBCL), including emerging possibilities for future use.