Beyond MABEL: An Integrative Approach to First in Human Dose Selection of Immunomodulators by the Health and Environmental Sciences Institute (HESI) Immuno-Safety Technical Committee (ITC).

Administration of a new drug candidate in a first-in-human (FIH) clinical trial is a particularly challenging phase in drug development and is especially true for immunomodulators, which are a diverse and complex class of drugs with a broad range of mechanisms of action and associated safety risks. Risk is generally greater for immunostimulators, in which safety concerns are associated with acute toxicity, compared to immunosuppressors, where the risks are related to chronic effects. Current methodologies for FIH dose selection for immunostimulators are focused primarily on identifying the minimum anticipated biological effect level (MABEL), which has often resulted in sub-therapeutic doses, leading to long and costly escalation phases. The Health and Environmental Sciences Institute (HESI) - Immuno-Safety Technical Committee (ITC) organized a project to address this issue through two complementary approaches: (i) an industry survey on FIH dose selection strategies and (ii) detailed case studies for immunomodulators in oncology and non-oncology indications. Key messages from the industry survey responses highlighted a preference toward more dynamic PK/PD approaches as in vitro assays are seemingly not representative of true physiological conditions for immunomodulators. These principles are highlighted in case studies. To address the above themes, we have proposed a revised decision tree, which expands on the guidance by the IQ MABEL Working Group (Leach et al. 2021). This approach facilitates a more refined recommendation of FIH dose selection for immunomodulators, allowing for a nuanced consideration of their mechanisms of action (MOAs) and the associated risk-to-benefit ratio, among other factors.

Recovery Animals in Toxicology Studies: An Innovation and Quality Consortium Perspective on Best Practices With Case Study Examples.

The inclusion of recovery animals in nonclinical safety studies that support clinical trials is undertaken with a wide diversity of approaches even while operating under harmonized regulatory guidance. While empirical evaluation of reversibility may enhance the overall nonclinical risk assessment, there are often overlooked opportunities to reduce recovery animal use by leveraging robust scientific and regulatory information. In the past, there were several attempts to benchmark recovery practices; however, recommendations have not been consistently applied across the pharmaceutical industry. A working group (WG) sponsored by the 3Rs Translational and Predictive Sciences Leadership Group of the IQ Consortium conducted a survey of current industry practice related to the evaluation of reversibility/recovery in repeat dose toxicity studies. Discussion among the WG representatives included member company strategies and case studies that highlight challenges and opportunities for continuous refinements in the use of recovery animals. The case studies presented in this paper demonstrate increasing alignment with the Society of Toxicologic Pathology recommendations (2013) towards (1) excluding recovery phase cohorts by default (include only when scientifically justified), (2) minimizing the number of recovery groups (e.g., control and one dose level), and (3) excluding controls in the recovery cohort by leveraging external and/or dosing phase data. Recovery group exclusion and decisions regarding the timing of reversibility evaluation may be driven by indication, modality, and/or other scientific or strategic factors using a weight of evidence approach. The results and recommendations discussed present opportunities to further decrease animal use without impacting the quality of human risk assessment.

Non-Human Primate Husbandry and Impact on Non-Human Primates Health: Outcomes From an IQ DruSafe/3RS Industrial Benchmark Survey.

The presence of health issues (diarrhea, poor body condition) in non-human primates can impact animal welfare, confound toxicity study data, and lead to animal exclusion from studies. A working group cosponsored by DruSafe and 3Rs Translational and Predictive Sciences Leadership Groups of the IQ Consortium conducted a survey to benchmark quarantine, pre-study screening, husbandry, and veterinary care practices and their impact on NHP health. Nineteen companies participated in the survey providing separate responses for studies conducted in-house and at Contract Research Organizations from 3 regions (North America (NA), Europe and Asia) for an aggregate of 33 responses. A majority of responding companies conducted studies at North America CROs (39%) or in-house (36%) using primarily Chinese (33%) or Cambodian (27%) and to a lesser extent Vietnam (18%) or Mauritian (15%) origin NHPs. Forty-Five percent of responses had pre-study health issues (fecal abnormalities, etc.) on ≥ 1 studies with the highest incidence observed in Vietnam origin NHPs (80%). The survey suggested variable pre-screening and quarantine practices across facilities. Husbandry practices including behavioral assessments, environmental enrichment and consistent diets were associated with a lower incidence of health issues. The survey also benchmarked approaches used to diagnose and manage abnormal feces in NHPs and has provided strategies to minimize impact on NHP health. The survey highlighted opportunities for harmonizing screening criteria across industry and for improving tracking and sharing of health screening results, leading to further refinement of NHP veterinary care practices, higher quality studies, and reduced NHP use.

Toxicologic Pathology Forum: Opinion on Not Euthanizing Control Animals in the Recovery Phase of Non-Rodent Toxicology Studies.

Nonclinical toxicology studies that are required to support human clinical trials of new drug candidates are generally conducted in a rodent and a non-rodent species. These studies typically contain a vehicle control group and low, intermediate, and high dose test article groups. In addition, a dosing-free recovery phase is sometimes included to determine reversibility of potential toxicities observed during the dosing phase and may include additional animals in the vehicle control and one or more dose groups. Typically, reversibility is determined by comparing the test article-related changes in the dosing phase animals to concurrent recovery phase animals at the same dose level. Therefore, for interpretation of reversibility, it is not always essential to euthanize the recovery vehicle control animals. In the absence of recovery vehicle control tissues, the pathologist's experience, historical control database, digital or glass slide repositories, or literature can be used to interpret the findings in the context of background pathology of the species/strain/age. Therefore, in most studies, the default approach could be not to euthanize recovery vehicle control animals. This article provides opinions on scenarios that may or may not necessitate euthanasia of recovery phase vehicle control animals in nonclinical toxicology studies involving dogs and nonhuman primates.

Correction to: Behavioral Artistry: Examining the Relationship Between the Interpersonal Skills and Effective Practice Repertoires of Applied Behavior Analysis Practitioners.

The article Behavioral Artistry: Examining the Relationship Between the Interpersonal Skills and Effective Practice Repertoires of Applied Behavior Analysis Practitioners, written by Kevin Callahan, was originally published electronically on the publisher's internet portal (currently SpringerLink) on August 29, 2019 without open access. With the author(s)' decision to opt for Open Choice the copyright of the article changed on August, 2019 to © The Author(s) 2019 and the article is forthwith distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits use, duplication, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license and indicate if changes were made.

Behavioral Artistry: Examining the Relationship Between the Interpersonal Skills and Effective Practice Repertoires of Applied Behavior Analysis Practitioners.

This study investigated interpersonal skills associated with the concept of behavioral artistry (BA), a repertoire of practitioner behaviors including care, attentiveness, and creativity, among others, associated with the effective delivery of applied behavior analysis (ABA) treatment. Survey results indicated parents of children with autism preferred BA descriptors for ABA therapists over non-BA descriptors. A separate survey of 212 university students on a standardized personality assessment revealed students majoring and/or working in the field of ABA had lower levels of BA than those in other human services professions. Practitioners with higher BA scores were observed and rated more positively in their delivery of ABA for children with autism. Implications for training/supervising effective ABA practitioners within a BA model are discussed.

Arterial Thickness and Stiffness Are Independently Associated with Left Ventricular Strain.

BACKGROUND: The aim of this study was to examine the association between myocardial strain and arterial thickness and stiffness in young adults. Increased common carotid artery intima media thickness and peripheral arterial stiffness are known to precede coronary artery disease and cardiovascular (CV) events such as myocardial infarction and congestive heart failure. However, subclinical cardiac dysfunction can be detected in high-risk adults by myocardial strain echocardiography. The authors hypothesized that increased carotid artery intima media thickness would be associated with abnormal myocardial strain in young subjects who had obesity and type 2 diabetes mellitus. METHODS: CV risk factors were collected in 338 young adults participating in a prospective, cross-sectional study. The CV parameters collected included intima-media thickness, peripheral arterial stiffness by brachial distensibility, and myocardial strain and strain rate. General linear models were constructed to determine if vascular structure and function measures were independently associated with myocardial strain and strain rate. RESULTS: A linear relationship was found between global longitudinal strain obtained from the four-chamber view and global strain rate in systole and carotid intima-media thickness (four-chamber global longitudinal strain: ß = 3.0, CV risk factor-adjusted R2 = 0.34; global strain rate in systole: ß = 0.0053, R2 = 0.21; P ≤ .0001) and between four-chamber global longitudinal strain and lower brachial distensibility (ß = -0.42, R2 = 0.22; P < .001). CONCLUSIONS: Adverse changes in vascular structure and function are simultaneously present with reduced myocardial systolic function.

Structure-Based Screen Identification of a Mammalian Ste20-like Kinase 4 (MST4) Inhibitor with Therapeutic Potential for Pituitary Tumors.

Pituitary tumors of the gonadotrope lineage are often large and invasive, resulting in hypopituitarism. No medical treatments are currently available. Using a combined genetic and genomic screen of individual human gonadotrope pituitary tumor samples, we recently identified the mammalian sterile-20 like kinase 4 (MST4) as a protumorigenic effector, driving increased pituitary cell proliferation and survival in response to a hypoxic microenvironment. To identify novel inhibitors of the MST4 kinase for potential future clinical use, computational-based virtual library screening was used to dock the SelleckChem kinase inhibitor library into the ATP-binding site of the MST4 crystal structure. Several inhibitor candidates were identified with the potential to bind with high affinity. Using a TR-FRET in vitro recombinant kinase assay, hesperadin, initially described as an Aurora kinase inhibitor, exhibited potent inhibition of the MST4 kinase at nanomolar concentrations. The LßT2 gonadotrope pituitary cell hypoxic model was used to test the ability of this inhibitor to antagonize MST4 actions. Under short-term severe hypoxia (1% O2), MST4 protection from hypoxia-induced apoptosis was abrogated in the presence of hesperadin. Similarly, under chronic hypoxia (5%), hesperadin blocked the proliferative and colony-forming actions of MST4 as well as the ability to activate specific downstream signaling and hypoxia-inducible factor-1 effectors. Together, these data identify hesperadin as the first potent, selective inhibitor of the MST4 kinase with the capacity to block pituitary tumor cell growth in a hypoxic microenvironment.

Novel Interaction of Class IIb Histone Deacetylase 6 (HDAC6) with Class IIa HDAC9 Controls Gonadotropin Releasing Hormone (GnRH) Neuronal Cell Survival and Movement.

The impact of histone deacetylases (HDACs) in the control of gonadotropin releasing hormone (GnRH) neuronal development is unknown. We identified an increase in many HDACs in GT1-7 (differentiated) compared with NLT (undifferentiated) GnRH neuronal cell lines. Increased HDAC9 mRNA and protein and specific deacetylase activity in GT1-7 cells suggested a functional role. Introduction of HDAC9 in NLT cells protected from serum withdrawal induced apoptosis and impaired basal neuronal cell movement. Conversely, silencing of endogenous HDAC9 in GT1-7 cells increased apoptosis and cell movement. Comparison of WT and mutant HDAC9 constructs demonstrated that the HDAC9 pro-survival effects required combined cytoplasmic and nuclear localization, whereas the effects on cell movement required a cytoplasmic site of action. Co-immunoprecipitation demonstrated a novel interaction of HDAC9 selectively with the Class IIb HDAC6. HDAC6 was also up-regulated at the mRNA and protein levels, and HDAC6 catalytic activity was significantly increased in GT1-7 compared with NLT cells. HDAC9 interacted with HDAC6 through its second catalytic domain. Silencing of HDAC6, HDAC9, or both, in GT1-7 cells augmented apoptosis compared with controls. HDAC6 and -9 had additive effects to promote cell survival via modulating the BAX/BCL2 pathway. Silencing of HDAC6 resulted in an activation of movement of GT1-7 cells with induction in acetylation of α-tubulin. Inhibition of HDAC6 and HDAC9 together resulted in an additive effect to increase cell movement but did not alter the acetylation of αtubulin. Together, these studies identify a novel interaction of Class IIa HDAC9 with Class IIb HDAC6 to modulate cell movement and survival in GnRH neurons.

Loss of Growth arrest specific gene 6 (Gas6) results in altered GnRH neuron migration, delayed vaginal opening and sexual maturation in mice.

Prior work has shown the importance of TAM (Tyro3, Axl, Mer) receptor tyrosine kinases in GnRH neuronal development and reproductive function. It is unclear if TAM receptor actions are dependent on ligand activation for their functional effects; thus, we characterized reproductive phenotype of ligand Growth arrest specific gene (Gas6) null mice. Gas6 null mice showed delayed vaginal opening and delayed first estrus. Animals eventually attained normal estrous cycles as adults. The GnRH neuronal population was significantly decreased in Gas6 null adults and embryos, but the final positioning of cell bodies in the hypothalamus was normal. Vaginal tissue showed up-regulation of TAM receptor mRNAs in the absence of the ligand. These data confirm that Gas6 plays a role in early GnRH neuronal development and during vaginal opening. The phenotype of Gas6 KO mice suggests that TAMs function in a ligand-dependent and independent manner to control GnRH neuron development to modulate normal reproductive function.

Pancreatic tumor cell secreted CCN1/Cyr61 promotes endothelial cell migration and aberrant neovascularization.

The complex signaling networks between cancer cells and adjacent endothelial cells make it challenging to unravel how cancer cells send extracellular messages to promote aberrant vascularization or tumor angiogenesis. Here, in vitro and in vivo models show that pancreatic cancer cell generated unique microenvironments can underlie endothelial cell migration and tumor angiogenesis. Mechanistically, we find that pancreatic cancer cell secreted CCN1/Cyr61 matricellular protein rewires the microenvironment to promote endothelial cell migration and tumor angiogenesis. This event can be overcome by Sonic Hedgehog (SHh) antibody treatment. Collectively, these studies identify a novel CCN1 signaling program in pancreatic cancer cells which activates SHh through autocrine-paracrine circuits to promote endothelial cell migration and tumor angiogenesis and suggests that CCN1 signaling of pancreatic cancer cells is vital for the regulation of tumor angiogenesis. Thus CCN1 signaling could be an ideal target for tumor vascular disruption in pancreatic cancer.

Exposure of neonatal rats to the endocrine disrupter Bisphenol A affects ontogenic expression pattern of testicular steroid receptors and their coregulators.

In light of the adverse reports of Bisphenol A (BPA) on reproduction and considering the pivotal role played by the steroid receptors (SRs) and their coregulators in male reproduction, it was of interest to decipher the influence that BPA may have on their expression pattern during critical 'windows' of development. Male rats were injected with 2.4 µg per pup per day of BPA from postnatal days (PND) 1-5 and controls received vehicle. During development, the testicular expression pattern of SRs (AR, ERß and ERα), coactivators (SRC-1, SRC-2 and SRC-3) and corepressors (NCoR and SMRT) in BPA-exposed rats were compared. A significant decrease in the expression of SRs was seen in the BPA group. SRC-1 showed a significant decrease, whereas SRC-2 and SRC-3 showed a significant increase in the protein expression whereas corepressor expression remained unaltered in the BPA-exposed groups. Such impairments in the expression pattern can be a putative mechanism of adversities on fertility as a result of BPA exposure.

Helex device closure of multiple atrial septal defects.

OBJECTIVES: To describe the effectiveness of the Helex Septal Occluder (HSO) to close multiple atrial septal defects (mASDs). Background : Limited information is available describing closure of mASDs with the HSO. METHODS: A total of 28 patients who underwent closure of mASDs with the HSO were identified by retrospective review of our catheterization database between 2001 and 2012. Procedural details and follow up information were collected. RESULTS: Median age was 19.2 years, median weight 48 kg, with 10 (36%) patients weighing <25 kg. Indication for closure was RV enlargement (RVE) in all patients and additionally neurologic events occurred in 3/28 (11%). Median stop-flow diameter for the largest ASD was 14 (4-23) mm. One HSO was implanted in 21/28 (75%), 2 in 6/28 (21%), and 3 in 1/28 (4%). One embolization and one transient arrhythmia occurred with no sequelae. Immediate residual shunt was absent in 5/28 (18%), trivial in 15/28 (54%), small in 6/28 (21%), and moderate in 2/28 (7%). Of the 25 patients with ≥6 months follow-up (median 53 months), residual shunt was absent in 13/25 (52%), trivial in 5/25 (20%), and small in 7/25 (28%). RVE resolved in all but one patient with no other associated lesions and ≥6 months of follow-up. No patient with prior neurological event had recurrence at last follow-up. CONCLUSIONS: We conclude that closure of mASDs with ≥1 HSO is effective with a low complication rate. The ability of HSO devices to overlap or sandwich each other may facilitate safe implantation of multiple devices in smaller patients.

AXL and MET crosstalk to promote gonadotropin releasing hormone (GnRH) neuronal cell migration and survival.

The membrane tyrosine kinase receptors, AXL and MET, are implicated in GnRH neuron migration and/or survival. We hypothesized that the receptors with their ligands, GAS6 and HGF, respectively may cross-talk in GnRH neuronal function. In NLT GnRH neuronal cells, MET co-immunoprecipitated with AXL, although HGF or GAS6 did not transphosphorylate AXL or MET, respectively. Co-expression of a kinase dead AXL blocked HGF activation of MET and indirectly AKT and p38MAPK. Silencing of AXL decreased HGF's ability to phosphorylate MET and activate AXL's downstream effectors, p38MAPK and AKT. HGF/MET signaling modulated neuron migration dependent and independent of AXL co-expression and p38MAPK. Conversely, AXL's control of GnRH neuronal survival was dependent on HGF/MET signaling. Together, these data support that the importance of membrane tyrosine kinase receptor crosstalk to regulate neuronal cell-specific developmental functions.

Predictive factors in patients with advanced and metastatic squamous cell carcinoma of the head and neck: a study based on SWOG protocol S0420.

To evaluate the prognostic values of different protein expression in the progression of squamous cell carcinoma of the head and neck (SCCHN) patients, we conducted immunohistochemical (IHC) analysis in tissue samples of different patients enrolled on SWOG protocol S0420. S0420 was a phase II trial to evaluate the efficacy and safety of single-agent sorafenib in chemotherapy-naïve patients with metastatic or recurrent SCCHN. The primary end point was response probability, i.e., confirmed complete (CR) and partial response (PR). Sorafenib was administered orally at 400 mg twice daily on a continuous basis in 28-day cycles to eligible patients. Responses were evaluated according to RECIST (Response Evaluation Criteria in Solid Tumors) criteria. IHC analysis was performed for various markers and data were analyzed statistically. IHC data were obtained from 19 patients enrolled on S0420. There was a high frequency of cases with expression of the angiogenesis markers SMA, HIF-1α, Raf-1, VEGF and VEGF-R. None of the markers were significantly associated with response. Negative HER-2 status was associated with longer progression-free survival (PFS), P=0.04. Negative NRP-1 status was associated with longer overall survival (OS), P=0.04. There were no other significant associations. An almost universal overexpression of angiogenesis markers in the samples analyzed supports the evaluation of angiogenesis inhibition as a potential target for therapy. High levels of NRP-1 and HER-2 in SCCHN samples appear to be associated with decreased survival and earlier progression of disease, respectively, in SCCHN patients and may represent targets for therapy.

Personal FM systems for children with autism spectrum disorders (ASD) and/or attention-deficit hyperactivity disorder (ADHD): an initial investigation.

UNLABELLED: The goal of this initial investigation was to examine the potential benefit of a frequency modulation (FM) system for 11 children diagnosed with autism spectrum disorders (ASD), attention-deficit hyperactivity disorder (ADHD), or both disorders through measures of speech recognition performance in noise, observed classroom behavior, and teacher-rated educational risk and listening behaviors. Use of the FM system resulted in significant average improvements in speech recognition in noise for the children with ASD and ADHD as well as large effect sizes. When compared to typically functioning peers, children with ASD and ADHD had significantly poorer average speech recognition performance in noise without the FM system but comparable average performance when the FM system was used. Similarly, classroom observations yielded a significant increase in on-task behaviors and large effect sizes when the FM system was in use during two separate trial periods. Although teacher ratings on questionnaires showed no significant improvement in the average level of educational risk of participants, they did indicate significant improvement in average listening behaviors during two trial periods with the FM system. Given the significantly better speech recognition in noise, increased on-task behaviors, and improved teacher ratings of listening behaviors with the FM system, these devices may be a viable option for children who have ASD and ADHD in the classroom. However, an individual evaluation including audiological testing and a functional evaluation in the child's primary learning environment will be necessary to determine the benefit of an FM system for a particular student. LEARNING OUTCOMES: 1. The reader will be able to describe the potential benefit of FM systems for children with ASD and/or ADHD. 2. The reader will be able to identify on-task versus off-task listening behaviors in children with ASD and/or ADHD. 3. The reader will be able to explain the components of a successful pre-fit education program that may be necessary prior to fitting an FM system in children with ASD.

Unusual cause of myocardial ischemia in an infant.

Abnormal coronary artery anatomy should be ruled out in any patient with myocardial dysfunction and an abnormal electrocardiogram. The reported etiologies of infantile myocardial ischemia are abnormalities of coronary arteries, perinatal asphyxia, umbilical catheterization, and myocarditis. Generalized arterial calcification of infancy, although rare, should be considered in the differential diagnosis of infantile myocardial ischemia if coronary artery origin is found to be normal on echocardiography.

The matricellular protein CCN1/Cyr61 is a critical regulator of Sonic Hedgehog in pancreatic carcinogenesis.

CCN1 is a matricellular protein and a member of the CCN family of growth factors. CCN1 is associated with the development of various cancers including pancreatic ductal adenocarcinoma (PDAC). Our recent studies found that CCN1 plays a critical role in pancreatic carcinogenesis through the induction of EMT and stemness. CCN1 mRNA and protein were detected in the early precursor lesions, and their expression intensified with disease progression. However, biochemical activity and the molecular targets of CCN1 in pancreatic cancer cells are unknown. Here we show that CCN1 regulates the Sonic Hedgehog (SHh) signaling pathway, which is associated with the PDAC progression and poor prognosis. SHh regulation by CCN1 in pancreatic cancer cells is mediated through the active Notch-1. Notably, active Notch-1is recruited by CCN1 in these cells via the inhibition of proteasomal degradation results in stabilization of the receptor. We find that CCN1-induced activation of SHh signaling might be necessary for CCN1-dependent in vitro pancreatic cancer cell migration and tumorigenicity of the side population of pancreatic cancer cells (cancer stem cells) in a xenograft in nude mice. Moreover, the functional role of CCN1 could be mediated through the interaction with the αvß3 integrin receptor. These extensive studies propose that targeting CCN1 can provide a new treatment option for patients with pancreatic cancer since blocking CCN1 simultaneously blocks two critical pathways (i.e. SHh and Notch1) associated with the development of the disease as well as drug resistance.

Rapid-onset hypogonadism secondary to crizotinib use in men with metastatic nonsmall cell lung cancer.

BACKGROUND: The objective of this study was to document the differences in testosterone (T) levels between crizotinib-treated and noncrizotinib-treated patients with metastatic nonsmall cell lung cancer (NSCLC). METHODS: Testosterone levels were measured in 19 men with metastatic NSCLC who received crizotinib and in 19 men with metastatic NSCLC who did not receive crizotinib. Clinical characteristics of the patients were compared, and additional hormone assays were performed as appropriate. Two patients who began crizotinib and 4 patients who had dose interruptions or who stopped crizotinib therapy had serial hormone measurements, permitting the documentation of dynamic hormone changes on and off crizotinib treatment. RESULTS: Total T levels were low (<241 ng/dL) in 19 of 19 (100%) crizotinib-treated men and in 6 of 19 men (32%) with metastatic NSCLC who did not receive crizotinib (mean T levels, 131 ng/dL and 311 ng/dL, respectively; P = .0002). Only 1 in 5 patients who had anaplastic lymphoma kinase (ALK) gene rearrangements and had not yet received crizotinib had low T. The initiation of crizotinib in 2 patients who had previously normal T levels was associated with a rapid decreases in T and in luteinizing hormone and follicle stimulating hormone levels within 14 to 21 days. Discontinuation of crizotinib led to increases back to normal T levels. CONCLUSIONS: Crizotinib therapy caused rapid suppression of T levels in men. The current results indicated that the site of action must include a central (hypothalamic or pituitary) effect, but additional direct testicular effects could not be excluded. Further work is required to assess the correlation between low T levels and crizotinib side effects as well as the exact molecular mechanism and site of drug toxicity.