Genetic models of the ANP gene or the GCA receptor together with genomic medicine have solidified the concept that both cardiac hormones are fundamental for blood pressure homeostasis and when deficient or disrupted they may contribute to human hypertension. Advances in peptide engineering have led to novel peptide therapeutics including the ANP-analog MANP for human hypertension. Most importantly a first in human study of MANP in essential hypertension has demonstrated its unique properties of aldosterone suppression and blood pressure reduction. Physiology and pharmacology ultimately lead us to innovative peptide-based therapeutics to reduce the burden of cardiovascular disease.The heart serves as an endocrine organ producing the hormones atrial natriuretic peptide (ANP) and b-type natriuretic peptide (BNP) which  Hypertension still represents the leading cause of death worldwide. Importantly, ranking number two in cause of death is metabolic disease including diabetes and obesity both of which are predisposing and coexisting factors for hypertensive disease . Much prThe work of via a hormonal mechanism regulates blood pressure under physiologic conditions. Various studies involving additional genetic mice models reinforced the notion of cardiac hormones as blood pressure regulators. Further investigations also revealed that beyond blood pressure, GC-A protects the cardiomyocyte from hypertrophy  study, Cannone and colleagues extended studies on rs198389 focusing on a population with cardiovascular risk factors. Specifically, these investigators utilized the landmark St. Vincent Screening to Prevent Heart Failure Study (Stop-Heart Failure Study), which investigated the efficacy of using BNP based screening in combination with collaborative care between primary care physicians and cardiovascular specialists in the prevention of new onset heart failure and left ventricular dysfunction (LVD) . ImportaTherefore, Thus, this later study went beyond a simple biomarker investigation with relevance to precision medicine . The stuInvestigations of ANP and BNP since the discovery ANP by DeBold (1981) have resulted in thousands of publications elucidating mechanisms of action supporting therapeutic potential especially in hypertension . Centralin vivo study of MANP compared to ANP in normal canines, MANP demonstrated widespread superiority to ANP in reducing blood pressure, augmenting sodium excretion and inhibiting angiotensin II and aldosterone  design in multiple cohorts of hypertensive subjects. Three doses were employed. Natriuretic, aldosterone suppressing and BP lowering actions were assessed. Twelve patients were recruited. Inclusion criteria included (1) hypertensive subjects with SBP above 140 mm Hg; (2) on at least one anti-hypertensive medication; and (3) DBP equal to or in excess of 90 mm Hg. Exclusion criteria included (1) known hypersensitivity or allergy to MANP or its components, carperitide, other natriuretic peptides, or related compounds; (2) women who are pregnant or breast-feeding; (3) any disease or condition  which, in the opinion of the investigator, might compromise the hematologic, cardiovascular, pulmonary, renal, gastrointestinal, hepatic, or central nervous system; or other conditions that may interfere with the absorption, distribution, metabolism, or excretion of study drug, or would place the subject at increased risk; (4) the presence of abnormal laboratory values considered clinically significant by the Investigator; (5) positive screen for Hepatitis B , Hepatitis C , or HIV (anti-HIV 1/2); (6) received an investigational drug within a 30-day period of Visit 1; (7) consumption of alcohol within 48 h prior to dose administration or during any in-patient period; (8) a positive urine drug screen including ethanol, cocaine, THC, barbiturates, amphetamines, benzodiazepines, and opiates unless, in the opinion of the Investigator, a positive finding is as a result of a legitimate medical prescription for a valid medical condition; (9) a history (within the last 2 years) of alcohol abuse, illicit drug use, significant mental illness, physical dependence to any opioid, or any history of drug abuse or addiction; (10) a history of difficulty with donating blood or donated blood or blood products within 45 days prior to enrollment; (11) clinically significant new illness in the 1 month before screening in the opinion of the Investigator; (12) history of severe allergies ; (13) history of coronary artery disease; (14) history of cerebrovascular disease; (15) history of epilepsy or other seizure disorder; (16) history of syncope; (17) history of organ transplantation; and (18) malignancy within 5 years of the screening visit .The first in human study of MANP was recently reported and performed in subjects with essential hypertension bypassing normal volunteers based upon MANP’s efficacy and safety in multiple studies in normal, hypertensive and heart failure canines and rodent models . The goavia GC-A/cGMP. In this first in human study, investigators reported a dose response to MANP in natriuresis during the 4 h after SQ MANP. While the duration of aldosterone reduction was variable among the treatment cohorts, a signal for a reduction and off-response by 24 h was clearly observed, thus, extending to the human the aldosterone suppressing actions of GC-A activation previously reported by in vitro and in animal studies , angiotensin A1 (AT1) receptor blockers and calcium channel blockers. Thus, data will be available on the blood pressure reducing actions of MANP on top of conventional anti-hypertensive agents. A new expanded clinical trial has been initiated in 40 patients with resistant hypertension (20 Caucasians and 20 African Americans) which is placebo controlled and with multiple ascending doses. MANP is administered once daily for 5 days.The current delivery method for MANP is by subcutaneous injection once daily. As advances in peptide therapeutics in cardiovascular and metabolic disease accelerate, SQ delivery is emerging as with GLP-1 agonists for diabetes and PKC9 inhibitors for hypercholesterolemia. Nonetheless, research and development of MANP also includes the development of oral delivery of MANP. Here, advances in oral delivery of peptides such as BNP and GLP-1 agonists have also been reported supporting the feasibility of this exciting delivery strategy for peptides such as MANP .It should be stated that several questions remain to be addressed as MANP studies continue. These include establishing the ability of MANP to chronically reduce blood pressure and activate the cGMP pathway thus not having drug tolerance. In addition, the role of gender in responsiveness to MANP also needs to be established as sex-based differences in circulating ANP and BNP exist with plasma ANP and BNP being higher in females. Further, the potential modulating action of ANP and BNP genetic variants, which are associated with variations in circulating levels of the two hormones, should also be investigated in regard to blood pressure response to MANP. While ANP as an intravenous drug for heart failure has proved successful in Japan, the use of BNP (nesiritide) and urodilatin (ularitide) in heart failure has been disappointing. This may be related to excessively high doses which are hypotensive as well as their use only as acute drugs as opposed to chronic therapy at a lower dosage. Indeed, hypertension may be the most physiologic target for natriuretic peptide therapy based upon the evidence of a relative deficiency of ANP and BNP in human hypertension and blood pressure reduction being the signature pharmacologic action of MANP.Another drug breakthrough that involves the natriuretic peptides system by augmenting their circulating levels is sacubitril/valsartan. Sacubitril inhibits the enzyme neprilysin (NEP) which is widely expressed in the human body and degrades ANP and BNP. Valsartan serves as an AT1 blocker. Sacubitril/valsartan was approved for heart failure based upon an improvement in mortality in the pivotal PARADIGM HF trial . RuilopeSince the discovery of ANP by Adolfo DeBold that established the heart as an endocrine organ, the field of natriuretic peptides is again accelerating. It is not unusual that it has taken four decades to also establish the therapeutic potential natriuretic peptides and testing in humans. That may be the usual cycle of drug development. This includes designer peptides such as MANP and novel small molecules as sacubitril/valsartan which may inhibit the breakdown of endogenous ANP and BNP and boost their circulating levels . PerhapsThe original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.Both authors conceived the outline of the manuscript, wrote the first and final draft, contributed to the article, and approved the submitted version. JB designed the figures. VC finalized the references.Mayo Clinic has licensed MANP to E-STAR BIO and JB is the inventor of MANP. The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Cellular membranes can adopt a plethora of complex and beautiful shapes, most of which are believed to have evolved for a particular physiological reason. The closely entangled relationship between membrane morphology and cellular physiology is strikingly seen in membrane trafficking pathways. During clathrin-mediated endocytosis, for example, over the course of a minute, a patch of the more or less flat plasma membrane is remodeled into a highly curved clathrin-coated vesicle. Such vesicles are internalized by the cell to degrade or recycle plasma membrane receptors or to take up extracellular ligands. Other, steadier, membrane morphologies can be observed in organellar membranes like the endoplasmic reticulum or mitochondria. In the case of mitochondria, which are double membrane-bound, ubiquitous organelles of eukaryotic cells, especially the mitochondrial inner membrane displays an intricated ultrastructure. It is highly folded and consequently has a much larger surface than the mitochondrial outer membrane. It can adopt different shapes in response to cellular demands and changes of the inner membrane morphology often accompany severe diseases, including neurodegenerative- and metabolic diseases and cancer. In recent years, progress was made in the identification of molecules that are important for the aforementioned membrane remodeling events. In this review, we will sum up recent results and discuss the main players of membrane remodeling processes that lead to the mitochondrial inner membrane ultrastructure and in clathrin-mediated endocytosis. We will compare differences and similarities between the molecular mechanisms that peripheral and integral membrane proteins use to deform membranes. Cells of many higher eukaryotic organisms have developed mechanisms to take up nutrients, cell surface receptors, and other molecules. The arguably best studied uptake process is called clathrin-mediated endocytosis (CME)  are found in various proteins and peptides involved in a wide range of membrane-based processes . Binding to PIP2 leads to folding of the N-terminus into an AH referred to as α0. Basic residues on the polar side of the AH stabilize PIP2 binding and position the hydrophobic side that consequently inserts into the inner leaflet of the plasma membrane. Membrane binding of ENTH was shown to lead to membrane deformation  complexes. The understanding that the inner mitochondrial membrane is not just randomly folded but is structurally highly regulated started with the investigation of the OXPHOS complex V, also known as FThe fungal mitochondrial genome maintenance protein (Mgm1), in animals known as optical atrophy 1 protein (OPA1), is a large GTPase of the dynamin superfamily that is localized to the MIM. Members of this protein family are involved in various membrane trafficking and membrane remodeling processes, and Mgm1/OPA1 is one of the central elements in remodeling of the MIM (Ferguson and De Camilli Mature Mgm1 is anchored into the lipid bilayer of the MIM via a single transmembrane domain and exposes a large soluble part into the IMS. The IMS domain of Mgm1/OPA1 comprises four domains: GTPase (G-domain), bundle signaling element (BSE), stalk, and membrane-interacting paddle  or cardiolipin (CL) , phosphatidylethanolamine (PE), phosphatidylserine PS, phosphatidylinositol (PI), and phosphatidic acid (PA)  resides at mitochondrial cristae junctions. This multi-subunit protein complex is found in all eukaryotic organisms containing mitochondria (Muñoz-Gómez et al. Mic10 is the smallest, but the most abundant component of the MICOS complex. Studies on Mic10’s involvement in the maintenance of the MIM morphology began from the investigation of mitochondrial ultrastructure of the knock-out cells, as well as from overexpression experiments (Hoppins et al. According to the secondary structure prediction, the 10 kDa protein consists of two transmembrane helices. The first helix exhibits typical parameters of standard transmembrane domain, while the second helix appears to be several amino acid residues longer. Both helices contain conserved sequence motifs comprising consecutive glycine residues (Alkhaja et al. As glycine-rich protein motives are crucial for helix–helix interaction and transmembrane segments packing within lipid bilayers (Russ and Engelman Mic60 is the largest and evolutionary, the oldest component of the MICOS complex (Muñoz-Gómez et al. Secondary structure prediction suggests that the mature Mic60 protein consists of single N-terminal transmembrane domain, large central coiled coil domain and C-terminal signature mitofilin domain. Based on the results of proteinase K treatment of yeast mitoplasts, the N-terminal transmembrane domain of Mic60 is anchored into the MIM, while the coiled coil and the mitofilin domains are exposed to the mitochondrial IMS (Rabl et al. In vitro reconstitution studies using recombinantly purified full-length Mic60 revealed its membrane bending activity (Tarasenko et al. According to liposome co-sedimentation, as well as liposome flotation assays, Mic60’s ability to bind and remodel lipid membranes seems to be independent of its transmembrane domain (Tarasenko et al. Using secondary structure prediction tools, it was identified that the Mic60 protein contains two predicted AH, designated LBS1 and LBS2, in the region between its coiled coil and mitofilin domains (Hessenberger et al. Taking all together, Mic60 uses a different mechanism to bind and remodel the mitochondrial inner membrane in comparison to Mic10. In fact, this protein, while residing permanently in the MIM, used a similar membrane molding mechanism as proteins found in membrane trafficking events like CME. AH are known to sense and induce positive membrane curvature. They insert into the lipid bilayer via intercalation into one membrane leaflet competing for the space with the lipid head groups and eventually such asymmetrical insertion leads to curvature of the membrane. Usually, such curved membrane is additionally stabilized by rigid protein scaffold that supports the form of the underlying membrane e.g., BAR proteins or endophilins. Long, rod-shaped coiled coil domain of Mic60 together with amphipathic nature of its LBS1might employ similar mechanisms to shape the mitochondrial inner membrane at the CJ sites.As expected for a dynamic process like CME, where membrane remodeling takes place on a timescale of a few seconds, membrane curvature inducing molecules are only transiently attached and are recruited, and most probably released in a spatiotemporal regulated fashion. For a process of this importance, it is also not surprising that many membrane shaping proteins with partially overlapping function are found. In addition to scaffolding and shallow insertion of proteins on membranes, molecular crowding is likely contributing to the morphological maturation of clathrin-coated vesicles. In the future, it will be important to study cooperativity of protein–protein and protein–membrane interactions in CME. As for studying the physiological role of specific lipids, these experiments are much more complex and rely on continuous exciting progresses in model membrane systems and structural investigation of biological samples.The morphology of organellar membranes like mitochondria or the endoplasmic reticulum is, although able to dynamically respond to different cellular needs, steadier than what is observed in membrane trafficking events. Consequently, the few organelle shaping proteins that have been identified so far are, in most cases, integral membrane proteins, that reside constantly within the membrane. Due to their physical–chemical properties, in vitro investigations of these hydrophobic molecules are more demanding than for soluble proteins. A common mechanism by which membrane proteins induce curvature seems to be sterically driven. Here, the molecules cover different surface areas on each side of the membrane and, as such, adopt a wedge-like topology. Interestingly, integral membrane proteins that carry AH at their soluble domains, with which they mold membranes, have also been identified. Shallow insertion of protein moieties seems to be a general theme in the remodeling of biological membranes. Prospective studies will surely identify more membrane bending proteins and molecular mechanisms. It will be exciting to see more similarities and differences between soluble and integral membrane proteins and their effect on membrane morphology uncovered.


Bone diseases are the leading causes of disability and severely compromised quality of life. Neuropeptide Y (NPY) is a multifunctional neuropeptide that participates in various physiological and pathological processes and exists in both the nerve system and bone tissue. In bone tissue, it actively participates in bone metabolism and disease progression through its receptors. Previous studies have focused on the opposite effects of NPY on bone formation and resorption through paracrine modes. In this review, we present a brief overview of the progress made in this research field in recent times in order to provide reference for further understanding the regulatory mechanism of bone physiology and pathological metabolism. The mammalian skeleton is a vital organ formed by several bone types, and it is also the place for hematopoiesis and mineral storage, with powerful self-repair ability and mineralized extracellular matrix. The traditional view of factors affecting bone metabolism such as endocrine, paracrine, and mechanical stimulation has long been discussed. Recent findings reported that bone tissue  was abundantly innervated by autonomic nerve terminals, which is one of the key factors regulating bone metabolism and remodeling through direct or indirect manner , 2, makiWhen neuropeptide Y (NPY) was first discovered in 1983, the awareness of its function in energy balance, obesity, and bone metabolism has gradually increased . As a 36In this review, we focus on the effects of NPY on bone metabolism in some physiological and pathological states. The aims of this article are to review the regulatory effects and to achieve a comprehensive understanding of NPY on bone metabolism.Bone remodeling involves mineralized bone removal by osteoclasts followed by bone matrix formation through osteoblasts that subsequently become mineralized . It is avia five G-protein-coupled receptor subtypes named Y1R, Y2R, Y4R, Y5R, and Y6R, of which Y1R and Y2R modulate bone mass at differing sites and through different ways  and osterix are considered as master transcription factors in osteogenic differentiation and they control bone formation . Zhang eParadoxically, several studies have reported that NPY acts as a promoting factor in the process of bone formation and fracture repair. Liu et al. found that low doses of NPY stimulate BMSC osteogenic differentiation and mineralization while a high NPY concentration had the opposite effect . In pati2+ concentration, changed the gut microflora community composition, and improved bone mass in OVX rats  in the presence of macrophage colony-stimulating factor (M-CSF). When the receptor activator of nuclear factor-kappa B ligand (RANKL) binds RANK on the surface of osteoclast precursors, osteoclastogenesis is immediately triggered. CFU-M is further differentiated into mononucleated osteoclasts and subsequently fused to multinucleated osteoclasts, then fully matured upon a cognate interaction with osteoblasts . Wu et aough Y1R .in-vitro osteoclast differentiation induction, which means that central NPY inhibited osteoclastogenesis through Y2R  from the bone marrow to the peripheral blood and ameliorated low bone density in an ovariectomy-induced osteoporosis mouse model by reducing osteoclast number . Seldeenough Y2R .In our study, osteoclast number and activity seem not be significantly influenced by bone-specific deficiency of NPY in young and aged mice . Matic eIn addition to participating in bone metabolism through affecting bone turnover, NPY may also affect bone mass through other ways. Blood vessels play an irreplaceable important role in the metabolic balance of bones. Several studies have confirmed that NPY-immunoreactive fibers were predominantly localized alongside with blood vessel walls in bone; moreover, Y1R, Y2R, and Y5R were confirmed to be expressed on endothelial cells (ECs), providing a material basis for the vasoregulatory role of NPY in addition to directly regulating bone tissue cells . It has in vivo for OVX rats could improve bone microstructure and decrease bone microdamage, and this may be achieved via the cAMP/PKA/CREB signaling pathway (Osteoporosis (OP) is a common skeletal disorder characterized by compromised bone mass and degraded bone microarchitecture, often resulting in fragility fractures and severely compromised quality of life in elderly people. Increasing age and postmenopausal state are proven to be associated with this condition. Zhang et al. reported that ovariectomy induced NPY upregulation in bone tissue after constructing a model of OP in adult female mouse. γ‐Oryzanol (ORZ), a functional substance extracted from rice bran, alleviated the severity of postmenopausal and senile OP through the autonomic nervous system by inhibiting osteocytic-NPY secretion . In gluc pathway . Also, N pathway . Above a−/−) and osteoblastic-specific Y1R knockout mice to characterize whether Y1R plays a role in fracture healing. The fracture healing process was delayed in the global deletion of Y1R in mice, and this delay is independent from osteoblast-specific Y1R. In Y1R-specific deficient mice, delayed endochondral fracture healing seems to be the result of impaired inflammatory response and cartilage removal since Y1R is widely expressed in neuronal but also in non-neuronal cells, such as immune cells  patients, autonomic nervous system dysfunction and low BMD are intertwined with some mood disorders such as depression, fatigue, and migraine . Higher Previous studies have verified that NPY is widely present in the brain and bone tissue and strongly influences bone metabolism through direct and indirect manner. In addition to directly regulating bone formation and resorption, NPY may also participate in bone metabolism by affecting gut microbiota and blood vessel formation. Furthermore, NPY has also been reported to play an intermediary role in autonomic nerve regulation on bone metabolism. As a substance synthesized by multiple places, it will be a challenge to clearly clarify the role of NPY on bone turnover and elucidate the pathophysiology of common bone diseases mentioned above. Also, whether NPY derived from sympathetic nerve endings and osteocytes has different physiological effects remains to be explored. Even though previous studies have shown that NPY participates in bone metabolism, especially in the bone formation process and BMSC fate decision, the effect of NPY on osteoclastogenesis and mood disorder is not fully understood.In spite of NPY being mostly expressed in the central nervous system, the role of NPY secreted by surrounding tissues, organs, and cell types in bone metabolism and cell signal transduction may be an important future research consideration. Future research on NPY and its receptors will be beneficial for new drug development and identifying new treatments for bone diseases.YZ conceived and designed the manuscript. Q-CC wrote the paper. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Obesity continues to be a global health issue. According to the World Health Organization (WHO), its prevalence has increased almost 3-fold between 1975 and 2016; thirteen percent of the adult population worldwide was obese in 2016 . It is tGlucagon-like peptide-1 (GLP1) is a peptide hormone produced by the L cells of the small intestine when the cells detected food. By binding to its receptor on the pancreatic β-cells, GLP1 stimulates insulin secretion in a glucose-dependent manner. GLP1 also exerts the following effects: suppresses pancreatic glucagon release as well as hepatic glucose production, promotes the proliferation of pancreatic β-cells, prevents apoptosis of β-cells, and delays gastric emptying . These pCurrently, Liraglutide is the only FDA-approved GLP1 receptor agonist for weight loss treatment in non-diabetic patients. Semaglutide was added to the list by FDA recently for chronic weight management in obese or overweight adults with at least one weight-related condition such as hypertension, diabetes, or hypercholesterolemia . HoweverA cohort study reported that individuals who are obese and overweight had a 20% reduction in the GLP1 response to oral glucose compared with subjects who have normal weight . While tThe appetite-suppressant effect of GLP1 is well known. Briefly, two sets of neuronal circuitry are present within the arcuate nucleus (ARC) of the hypothalamus. These neuronal circuits signal to the paraventricular nucleus and modulate feeding behavior. The neuronal circuit that expresses neuropeptide Y (NPY) and agouti-related peptide (AgRP) stimulates food intake, while the ARC neuron that expresses pro-opiomelanocortin (POMC) and cocaine- and amphetamine-regulated transcript (CART) reduces food intake. GLP1 suppresses the expression of NPY and AgRP, while increases the activity of POMC and CART . BesidesThe concept that gut plays a critical role in obesity has been widely described. Most of these studies involved rodent treated with high fat diet (HFD), as the DIO model presents metabolic status that is closest to the human condition. Two months of HFD feeding increases the sensitivity of the gut to luminal nutrients, but the increased luminal sensitivity may not have stimulated GLP1 secretion . The posin vitro study conducted subsequently by the same group of researchers, shows that the increased GLP1 in the cell culture media as induced by methane, was not dose-dependent. It is therefore not clear if the enhanced GLP1 secretion in DIO mice (in vivo study beyond 14 weeks and measuring relevant parameters could confirm the methane effect on GLP1 secretion. In short, what is clearer at the moment is that HFD induces changes in the gut microbiome, and that causes inflammation and adiposity.Human studies reported the presence of inflammatory proteins in the systemic circulation of obese patients, which correlates positively with body weight , 34. TheDIO mice , which wResearch over the past two decades has established that GLP1 acts along the gut-brain axis, producing anti-obesity effects. But obesity negatively influences the effect of GLP1 centrally and peripherally through the appetite-regulating centre and the gut, respectively. A treatment regime that targets on both axes instead of one axis may therefore be more beneficial. This could mean increasing the appetite-suppressant effect of GLP1 and increasing or keeping the intestinal production of GLP1, both of which are achieved by improving the systemic bioavailability of GLP1. The bioavailability of GLP1 can be improved by developing GLP1 receptor agonists, which are resistant to enzyme dipeptidyl peptidase IV degradation. This approach has been widely adopted and most of the agonists have been approved for use in diabetic treatment . LiragluMany researchers have turned to dietary intervention, hoping that the approach would increase GLP1 secretion. Highly soluble and fermentable fibers, through the production of short chain fatty acids, promote GLP1 secretion . FructooDietary strategies are effective in reducing body weight. The effect is prolonged if the strategies are combined with behavioral counseling and ongoing support . High prVarious plant-based food, which have weight and fat mass reduction effects , 52, andCurrently available therapy has not been effective in curbing the upward trend of obesity prevalence, which should prompt us to reconsider the traditional approach of obesity treatment. The outcomes of various clinical trials suggest that adopting a holistic approach in weight management may be more effective than focusing on one specific strategy involving either drug or non-drug treatment. The holistic approach may comprise of a pharmacological therapy, implemented alongside a dietary intervention. Liraglutide and Semaglutide are promising weight reduction pharmacological agents. The treatment should be accompanied by behavioral therapy and on-going support for dietary changes. Long-term adherence to dietary intervention such as Mediterranean diet and low carbohydrate diet may help to produce a sustainable weight loss, while the consumption of food or herbal products with antioxidant and anti-inflammatory properties synergizes the effect of GLP1 receptor agonist and improves the overall health.The author confirms being the sole contributor of this work and has approved it for publication.The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Prostate cancer, after lung cancer, is the leading cause of death among men. The incidence rate of prostate cancer varies worldwide between regions and population groups. Its incidence increases with age and is more likely to occur in older men. Although the pathophysiological mechanisms and the etiological factors of prostate cancer development are still poorly understood, there are several factors associated with the risk of developing the disease such as age, family history, lifestyle-related factors , and testosterone levels. Cannabinoids are an emerging class of pharmacological molecules that may exert their therapeutic effect against different cancers, including those from the prostate. Several studies have shown that various agonists are able to target cannabinoid receptors exhibited on prostate cancer cells. This affects the release of pro-inflammatory cytokines and has impacts on cell proliferation, cell apoptosis and necroptosis; consequently leading to the development of cancerous cells.Prostate cancer is the second most frequently occurring cancer diagnosed among males. Recent preclinical evidence implicates cannabinoids as powerful regulators of cell growth and differentiation. In this review, we focused on studies that demonstrated anticancer effects of cannabinoids and their possible mechanisms of action in prostate cancer. Besides the palliative effects of cannabinoids, research from the past two decades has demonstrated their promising potential as antitumor agents in a wide variety of cancers. This analysis may provide pharmacological insights into the selection of specific cannabinoids for the development of antitumor drugs for the treatment of prostate cancer. The incidence rate of prostate cancer varies worldwide between regions and population groups. According to the global cancer statistics in 2020, 1,414,259 new cases of prostate cancer were registered globally, representing 7.3% of all cancers in men [Treatment options for prostate cancer include radical prostatectomy , radiatiThe correlation between androgen receptors and the production of dihydrotestosterone (DHT) in the prostate is shown in Cannabis sativa (C.sativa) plant. Cannabis has been known by numerous names such as hemp, hashish, bhang, and weed. C. sativa is one of the oldest cultivated plants and has been used for thousands of years, not only for its nutritional value but also for medicinal and textile applications [Plants have been used as traditional medicines for the treatment of numerous diseases. Recently, bioactive natural products are being used as an alternative therapy for prostate cancer, that offer fewer side effects compared to the conventional treatment of various prostate cancer cell lines such as cannabinoids obtained from the ications ,11. It hications . Since t9-THC) as a psychoactive component of cannabis (phytocannabinoids) and the major non-intoxicating compound cannabidiol (CBD); (b) endocannabinoids including arachidonoyl ethanolamide (anandamide) and 2-arachidonoylglycerol (2-AG), which were found to be products of lipid membrane precursors in humans and animals, forming the endocannabinoid system (ECS) as a critical neuromodulatory system; and (c) synthetic molecules that mimic the structure of either plant or mammalian cannabinoids such as (R)-methanandamide (MET), WIN-55, and JWH-133 [Cannabinoids are classified into three different categories based on their origin: (a) plant-originated cannabinoids that are found in the marijuana plant such as ∆9-tetrahydrocannabinol  cannabinoids and SCs has continued. Studies are now being conducted on the potential efficacy of cannabinoids, both natural and synthetic, as anticancer agents and their possible mechanisms of action [Synthetic cannabinoids (SCs) generate similar psychoactive and physiological effects to that of ∆o et al.  reportedf action ,20,21.The term “Entourage effect” was first introduced by Mechoulam and his group in 1988 . It refe9-THC in both in vitro and in vivo models of breast cancer. Furthermore, LaVigne et al. [Several studies have further supported this mechanism and demonstrated that all the compounds present in the cannabis plant work together to produce the therapeutic effect ,27. Blase et al.  reportedCannabinoids are a new emerging class of pharmacological molecules that can exert their therapeutic effect against different tumors including prostatic cancer ,30,31,32The pharmacological effect of cannabinoids is mediated through the stimulation of G protein-coupled receptors (GPCRs) called cannabinoid receptor type 1 (CB1) and cannabinoid receptor type 2 (CB2) . CB1 recReviewing the literature concerning the pharmacodynamics of a wide range of different cannabinoids that have been researched over recent years, it is far less clear what and how their pharmacological effect is mediated through the convoluted ECS . As discMacaca fascicularis [Cannabinoid receptors are well known to form heteromers with a wide range of different receptors . For exaicularis . Nowadayicularis .Heteromerisation has attracted the attention of scientists in recent years as a prominent potential target in cancer treatment amongst other chronic illnesses. Heteromers exhibit unique physicochemical properties associated with allosteric modulations. Such characteristics support those hybrid molecules to function as viable target sites and strategic platforms for different cannabinoid ligands treating a wide range of chronic diseases, including prostate cancer. Unlike the primary functional forms of plain monomeric receptors, they can lead to various and exemplary pharmacological and cellular functionalities that might influence the antitumor activity of the ECS depending on the combined properties of the complex heteromer and the applied pharmacological agent ,52,53. PAlong with heteromerization as one of the most successfully targeted platforms for selective therapeutic agent innovation, the allostery of ligands is becoming another attractive area to explore cannabinoid-based drug discovery for various medical conditions. Hence, switching from the traditional approach of designing a highly conserved orthostatic agonist/antagonist mimicking the endogenous cannabinoids, e.g., AEA and 2-AG, to flexible allosteric ligands offers a new class of selective therapeutic molecules. These new molecules modulate GPCRs through allosteric sites, resulting in specific heteromeric alterations manifested as a positive/negative allosteric modulator, leading to fine-tuned cellular signaling without undesirable adverse reactions, which has gained tremendous interest during the last decade .As mentioned above, the two cannabinoid receptors possess distinctive molecular structures. They are an integral part of the GPCR system, which undergoes configurational transformation following the binary interactions of the CB1/2 with different agonists, and the association of various intracellular signaling proteins associated with the distinctive complex of the G protein ,56. ThisMoreover, cannabinoid receptors can transduce cell signaling through non-G proteins pathways such as β-arrestins, as illustrated in The activation of cannabinoid receptors CB1 and CB2 results in an inhibition of adenylyl cyclase resulting in reduced cAMP levels in most tissues . NeverthThe inhibitory effect of cannabinoids on cell proliferation has been known since 1974. Dixit et al.  observedIn recent years, attention has been drawn to the pharmacological potential of cannabis on prostate cancer. Several experimental studies have demonstrated and evaluated the effects of various cannabinoids in prostate tissue and prostate cancer cells, which contain both CB1 and CB2 receptors ,70,71,72Nithipatikom et al.  tested tAdministration of cannabinoid receptor agonists such as MET and WIN-55,212 to the culture of LNCaP and PC3 cell lines showed a significant decline of their growth. This reduction in cell viability was associated with inhibition of Akt and activation of ERK ,37,81. F9-THC triggered cytosolic ceramide accumulation leading to the activation of ER stress response, resulting in the initiation of factor 2α phosphorylation followed by the autophagy process. Furthermore, they suggested that autophagy resulted from the inhibition of the Akt/mTOR complex 1 (mTORC1) axis, which is mediated via the tribbles homolog 3-dependent signaling (TRIB3) [Salazar et al.  found th (TRIB3)    found th (TRIB3) .Increased ceramide levels followed by an enhancement in ER stress triggered the activation of the caspase cascade leading to apoptosis . Several2−), hydroxyl radical (OH.), and hydrogen peroxide (H2O2). The high reactivity of these species is due to the presence of unpaired electrons whereby they can generate new reactive molecules or so-called free radicals, interacting with other biological molecules and leading to cellular injury [Apoptosis can be also induced by ROS. Oxidative stress is a phenomenon generated by an imbalance between the overproduction of ROS in the cell and the under-detoxification of these reactive molecules throughout the cell’s antioxidant capacity ,88. ROS r injury ,90. Moder injury . Neverthr injury ,89,92,93r injury . TargetiMassi et al.  investigThe underlying molecular mechanisms of apoptosis that are induced by oxidative stress via cannabinoid receptors have been thoroughly investigated. De Petrocellis et al.  tested tDando et al.  used metCannabinoids can be potential regulators of cell motility. Manipulation of cannabinoid receptor activity in different cell lines resulted in changes of cell motility ,98. It hNithipatikom et al.  investigWith respect to cancer, clinical trials demonstrating the effectiveness of cannabinoids are limited. A study in Australia is a phase I/II double blind trial assessing the effect of medicinal cannabis on the quality of life and symptom management in advanced cancer (ACTRN12619001534178) . A randoThe significant knowledge of anti-tumor and palliative characteristics of cannabinoids acquired by the research community in the past few years has increased the utilization of these molecules as favorable candidates for cancer treatment. Although, the use of cannabis in medicine is restricted due to its psychoactive effects, cannabinoid-based treatments that lack the unwanted side effects are sought after. The lack of knowledge of how cannabis exerts its anticancer effects makes it even more complicated to find what is the best way these compounds can be used without these psychoactive effects. However, there is enough evidence in the literature stating the ability of cannabinoids to induce cell death by various pathways in prostate cancer, but there is still more research needed to be undertaken to understand their mechanism ,105. Alt


Pollen, the plant male gametophyte, is a multinucleated structure that must develop into a pollen tube to penetrate through the style of the pistil and fertilize the female gametophyte. The development of the pollen tube is regulated by a molecular dialogue mediated by the ovary through the female pistil . Pollen Pollen tube extension proceeds by exocytotic activity of Golgi-derived vesicles at the pollen tip, resulting in a polarized tube membrane extension . These vPlant Physiology, Rui and colleagues (2021) show the COG complex interacts with two SYP3 (SYntaxin of Plants family 3) proteins, called SYP31 and SYP32. The interactions appear to be mediated by COG subunit 3, which bound SYP31 and SYP32 in pull-down assays both in vivo and in vitro, as well as in yeast two-hybrid and by split luciferase assays. Moreover, the absence of SYP31 and SYP32 disrupted COG association with the Golgi apparatus and led to its diffuse localization into the cytoplasm, thus showing SYP3s are essential for addressing the COG complex to the Golgi.In this issue of Arabidopsis thaliana) proteins SYP31 and SYP32 belong to the SNARE (Soluble Nsf-Associated Protein REceptor) protein family, and therefore likely mediate the fusion of vesicles to their target membrane (SYP31 overexpression inhibits anterograde trafficking in tobacco (Nicotiana tabacum) protoplasts . However, it was not possible to obtain double mutants after crossing the heterozygous syp31 line with any heterozygous syp32 line. This, together with the rescue of the mutant phenotype with SYP31- or SYP32-overexpression, confirmed that SYP31 and SYP32 are redundant and that their function is essential for pollen development.To assess the importance of SYP31 and SYP32 for pollen viability and development, the authors self-pollinated one heterozygous syp31 syp32 double mutants as early as after the first mitotic division (bicellular pollen stage), when SYP31 and SYP32 expression begins. Interestingly, these severe phenotypes appeared much earlier than the abnormal phenotype of the cog mutant, which is restricted to pollen tube growth (A follow-up on pollen grain maturation revealed pollen degeneration, including DNA degradation, in e growth . This susyp31 syp32 double mutants, which indicate deficient vesicle trafficking, both to the ER and to the plasma membrane (syp31 syp32 double mutant pollen grains was unevenly distributed, with empty patches on the pollen grain surface, suggesting defects in cellulose and cell-wall related enzymes trafficking to the plasma membrane. The morphology of the Golgi apparatus was also deeply affected (In agreement with this hypothesis, Rui and colleagues observed a relocation of ER proteins in the vacuole, as well as cytoplasmic pectin accumulation points in membrane . Moreoveaffected , with onIn conclusion, the Qa-type SNARE proteins SYP31 and SYP32 are essential for ensuring correct membrane transport in Arabidopsis pollen grains. Their functions are partly due to their role in mediating COG interaction with the Golgi and therefore for correct anterograde vesicle trafficking. However, SYP3s seem to orchestrate both anterograde and retrograde vesicle trafficking, likely by interacting with many other vesicle trafficking signaling components which remain to be identified.


The Bill & Melinda Gates Foundation supported respiratory syncytial virus (RSV) mortality surveillance studies in several low- and middle-income countries to address the striking gap in community mortality burden data from these geographies. The compelling findings generated from these studies reveal a high unmeasured burden of community RSV mortality, particularly among infants aged <6 months who are the target population for RSV immunization products currently in late-stage clinical development. These findings should inform revised global RSV mortality estimates and inform policy decisions on RSV vaccine financing and prioritization at the global and national levels. Respiratory syncytial virus (RSV) is the most common cause of severe lower respiratory tract infection (LRTI) in the first 6 months of life. The overwhelming burden of death occurs in low- and middle-income countries (LMICs) . HoweverBetween 2011 and 2013, a Bill & Melinda Gates Foundation–supported study in low-income neighborhoods of Buenos Aires documented striking seasonal peaks in all-cause LRTI mortality among infants in the community that consistently coincided with seasonal spikes of in-hospital RSV infant deaths, suggesting an undetected and unmeasured out-of-facility burden of RSV deaths among vulnerable communities . These oUnlike many other respiratory pathogens, detection of RSV in the nasopharynx is highly likely to indicate RSV illness. In a systematic review and analysis of 18 592 acute lower respiratory infection cases in young children across 23 studies, Shi and colleagues demonstrated that detection of RSV was significantly more common in children with acute LRTI than asymptomatic controls , with an attributable fraction among exposed of 90% .In the Pneumonia Program at the Gates Foundation, we leveraged these important findings to support RSV community mortality surveillance studies in Argentina, India, Pakistan, and Zambia, representing diverse LMIC geographies. All sites focused on the collection of nasopharyngeal (NP) swabs from recently deceased infants to assess the presence of RSV by polymerase chain reaction. Two of the studies, in Zambia and Argentina, enrolled deceased infants at the morgue, while the teams in India and Pakistan each implemented active surveillance to prospectively identify infant deaths within a defined community. The studies in Zambia and Pakistan focused on infants aged <6 months, while those in Argentina and India included those aged <5 years and <2 years, respectively. Common to all 4 studies was a goal of generating evidence on the burden of RSV among community deaths in young infants (aged <6 months) in LMICs.Murphy et al, CID 2021, current issue). Investigators in Pakistan used community-based active mortality surveillance for infants aged <6 months in 4 low-income settlement areas in peri-urban Karachi . Half of all deaths occurred in the community, though RSV detection during the surveillance period was lower (3.7%).In Lusaka, Zambia, the study team made use of local regulations mandating medical certification of all deaths before release for burial, thus enabling the capture of the majority (>80%) of deaths in the study catchment area. Community deaths accounted for 70% of infants enrolled. Over the course of 3 years, nearly 800 infants (aged <6 months) who died in the community were enrolled on presentation to the morgue; RSV was detected in nearly 8% . This is notably higher than the inflation factor of 2.2 applied to adjust RSV LRTI mortality in children aged <5 years in LMICs to account for increased unmeasured mortality in community settings in currently published global RSV mortality estimates [In central India, the study was conducted in Melghat, a rural area with high baseline infant mortality (>50/1000 live births), where the team used prospective active weekly surveillance for LRTIs and deaths across 93 villages. RSV was detected in 7.3% of annual post-neonatal mortality, and 80% of RSV deaths occurred in the community. In India, their active surveillance platform enabled calculation of a population-based estimate of an RSV LRTI mortality rate and indicated that mortality was 7- to 10-fold higher in the community compared with the hospital . Among 63 community deaths of children aged <5 years, RSV was detected in 12 (19%). All RSV-positive deaths were in infants aged <6 months , and RSV was deemed to be in the causal chain of death in 11 (91%).In Argentina, investigators partnered with the judicial morgue serving impoverished districts surrounding Buenos Aires to evaluate RSV in community deaths among children aged <5 years. Early study data from 2016–2017 indicated detection of RSV in almost 10% of these children, mostly among infants aged <12 months [Mazur et al, CID 2021, current issue). This is important because it is exactly this younger infant age group that stands to benefit most from RSV immunization products, where the highest efficacy is likely to be seen in the first 90 days of life. These community findings have important implications for the potential impact that RSV prophylaxis may have on mortality and are important to drive immunization policy.In all 4 settings, infants aged 0–3 months accounted for most RSV community deaths. In fact, relative younger age at death was consistently associated with community RSV deaths across RSV GOLD’s 38 country comparison of community vs hospital deaths .In addition to these 4 studies, all reported in this supplement, the CHAMPS platform permitted assessment of multiple etiologies of illness and death among deceased infants and children using molecular and pathologic evaluation of biological and MITS specimens from multiple organs, as well as assignment of the causal chain for each death using a standardized process. Although the majority of those evaluated in CHAMPS were facility deaths, these comprehensive data from sites in 7 African and South Asian countries provide additional important insights into the role of RSV in early infant mortality. Most notably, RSV was detected in 12% of deaths among infants aged 28 days–6 months and was determined to be in the causal chain of 6.5% of all deaths in this age group , RSV was determined to be in the causal chain in roughly 50% of RSV-positive deaths, and very few deaths were deemed to be due to only RSV. This may be, in part, a reflection of greater burden of additional infectious disease etiologies, including human immunodeficiency virus and malaria, in the higher mortality settings that characterize CHAMPS sites, as well as the greater complexity of facility-based deaths with multiple comorbidities. Nevertheless, it is important to recognize that RSV can increase risk for both bacterial coinfection as well as for subsequent LRTI episodes and thus serve as a triggering event for a poor outcome . FurtherMeasuring community mortality is a significant and challenging undertaking. Critically important to the success of all 4 studies was the importance of thoughtfully engaging the community to identify, enroll, and collect specimens from recently deceased infants in a timely fashion. While the nature of this engagement differed in each setting, the need to develop culturally sensitive approaches to meeting with families soon after the death of their child was universal. Investigators developed locally appropriate strategies that considered religious and cultural beliefs and practices to provide grief counseling to family members at the time of enrollment and beyond. Consistent engagement with local leaders was similarly critical to ensuring community acceptance of surveillance efforts.The compelling data generated from these surveillance efforts have revealed a high and previously unmeasured burden of community RSV mortality among young infants in LMICs. These findings are critical to enabling a more accurate understanding of the preventable burden of RSV deaths and should be incorporated into revised global RSV mortality estimates. RSV maternal vaccines and infant monoclonal antibody prophylaxis are currently in phase 3 trials and, if found to be efficacious, could be licensed within the next few years. It is imperative that decisions on global immunization financing as well country-level prioritization on vaccine preventable diseases be informed by estimates that reflect the complete burden of RSV.


Metastasis accounts for more than 90% of cancer related mortality, thus the most pressing need in the field of oncology today is the ability to accurately predict future onset of metastatic disease, ideally at the time of initial diagnosis. As opposed to current practice, what would be desirable is that prognostic, biomarker-based detection of metastatic propensity and heightened risk of cancer recurrence be performed long before overt metastasis has set in. Without such timely information it will be impossible to formulate a rational therapeutic treatment plan to favorably alter the trajectory of disease progression. In order to help inform rational selection of targeted therapeutics, any recurrence/metastasis risk prediction strategy must occur with the paired identification of novel prognostic biomarkers and their underlying molecular regulatory mechanisms that help drive cancer recurrence/metastasis (i.e. recurrence biomarkers). Traditional clinical factors alone (such as TNM staging criteria) are no longer adequately prognostic for this purpose in the current molecular era. FOXC1 is a pivotal transcription factor that has been functionally implicated to drive cancer metastasis and has been demonstrated to be an independent predictor of heightened metastatic risk, at the time of initial diagnosis. In this review, we present our viewpoints on the master regulatory role that FOXC1 plays in mediating cancer stem cell traits that include cellular plasticity, partial EMT, treatment resistance, cancer invasion and cancer migration during cancer progression and metastasis. We also highlight potential therapeutic strategies to target cancers that are, or have evolved to become, “transcriptionally addicted” to FOXC1. The potential role of FOXC1 expression status in predicting the efficacy of these identified therapeutic approaches merits evaluation in clinical trials. Cancer screening strategies have proven to be successful in decreasing cancer-specific mortality in a variety of cancers –4. HowevTraditional clinical factors alone (such as TNM staging criteria) are no longer prognostically adequate for this purpose and we are in need of new prognostic biomarkers that are superior in their ability to predict cancer recurrence and metastasis risk . Absent In recent years, several excellent review articles on the role of the Forkhead box C1 (FOXC1) Transcription Factor (TF) in cancer have been published, highlighting the increasing recognition of its importance as a clinically useful biomarker and potential therapeutic target –15. The The earliest reports on the biologic role and function of the FOXC1 TF implicated it in abnormal pathologic conditions like glaucoma –23, congThe functional relevance and prognostic significance of FOXC1 in cancer was first reported in breast cancer in 2010 . Since t expression status further correlated with increased incidence of brain and lung metastases and decreased metastasis-free survival in patients without lymph node involvement  molecular subtype, the most aggressive subtype of breast cancer , 128–131olvement . More reolvement . A cliniolvement . The assThe prognostic significance of FOXC1 in lung cancer was first reported in 2013  and lateFOXC1 expression status has now also been shown to be a poor prognostic indicator in multiple gastrointestinal cancers including esophageal cancer, gastric cancer, liver cancer, pancreatic cancer and colon cancer. Two independent published reports provided initial insight into the potential prognostic significance of FOXC1 expression in esophageal squamous cell carcinoma with regard to overall survival. However, univariate and multivariate hazard ratio values were not available in the published reports , 51. OneA fairly large investigation of patients diagnosed with hepatocellular carcinoma (HCC) demonstrated that FOXC1 expression was a powerful, statistically significant prognostic indicator of worse overall survival as well as recurrence free survival, independent of other clinical variables . This reAmongst the gastrointestinal cancers, the evidence in support of the prognostic utility of FOXC1 is perhaps most robust in colon cancer. Three independent studies have confirmed the fact that FOXC1 expression status can in fact predict worse overall survival in colon cancer –49. One Within head and neck cancers, some functional and mechanistic studies have been published implicating FOXC1 in the biology of these tumors. However, investigations examining the prognostic role of FOXC1 are not yet available. A preliminary report on tongue cancer did demonstrate FOXC1 to be a statistically significant predictor of worse overall survival . HoweverIn contrast, data regarding the prognostic impact of FOXC1 expression in cervical cancer is more developed. Two independent studies have reported that FOXC1 possesses prognostic value with regard to predicting shorter overall survival , 46. MorIt is important to note, however, that in two very specific cancer types, elevated FOXC1 expression, in contradistinction to the above evident trend, proved to be a predictor of favorable prognosis. These departures from the norm are unexpected exceptions to the seemingly apparent rule that is supported by the overwhelming majority of research findings in support of FOXC1 expression being an adverse prognostic indicator. Thus, while the preponderance of evidence supports elevated expression of FOXC1 being an accurate predictor of adverse clinical outcomes, the two notable exceptions include ovarian cancer and Luminal B molecular subtype of breast cancer where elevated expression of FOXC1 predicts good prognosis.vs. III-IV) as well as pathologic subtypes from benign to borderline and malignant tumors trending towards good prognosis. However, it should be noted that the sample size was small for cystadenocarcinomas (n=40) and pathologic subtype (n=80). Although nuclear and cytoplasmic FOXC1 staining was observed in cell lines, the above conclusion was based solely upon cytoplasmic FOXC1 as FOXC1 was not detected in the nucleus of the ovarian tumors.In the case of ovarian cancer, a retrospective study  demonstrThe second report on FOXC1 expression in Luminal B breast cancer  looks atThese highly exceptional findings merit further investigation into the context-specific subcellular localization of FOXC1, and factors that control and determine its post-translational stability and degradation. While initial insight into this aspect of FOXC1 regulation was provided in two published investigations of FOXC1 protein release, stability and degradation , 135, itin vitro and in vivo models The insight into FOXC1 as a prognostic predictor of cancer progression and metastasis was accompanied by a growing body of work that also demonstrated its functional importance as a molecular driver of these processes utilizing both By virtue of being a transcription factor, and being a central hub gene controlling a network of hundreds of genes, it is not surprising that upregulation of FOXC1 in cancer does cast wide influence on a number of biologic processes critical for tumor survival and propagation. While this includes proliferation as reported in a number of studies, what has been a hallmark feature of FOXC1+ status is that it appears to predominantly be responsible for a pro-metastatic phenotype. This is evident in Several molecular mechanisms have also been elucidated in this regard. These include induction of partial EMT that promotes enhanced cancer cell migration (discussed below), increased production of matrix metalloproteinases that promotes cancer cell invasion  enables cancer cells to depart from the primary tumor, invade surrounding tissue, and disseminate to distant organs. Several excellent reviews have implicated FOXC1’s role in EMT. After multiple initial reports that linked FOXC1 with EMT in various cancers, some recent reports have further refined our understanding that FOXC1 is in fact associated with a partial EMT phenotype comprising of hybrid E/M cells  and these mesenchymal CTCs were associated with disease progression . Similarotherapy . In thisotherapy , 146. Thotherapy . AdditioIn an independent study performed by Blanpain and colleagues that utilized a model of squamous cell carcinoma, the clonal population of tumor cells which maintain a hybrid E/M state, has been demonstrated to possess greater metastatic potential than either complete E polarized or complete M polarized cancer cells . In an HTumor cells are surrounded by a heterogeneous and complex tumor microenvironment (TME). The TME consists of a tumor specific extracellular matrix, which recruits an abundance of non-cancer cells including epithelial cells, endothelial cells, mesenchymal cells, immune cells and fibroblasts, all of which interact with the primary tumor cell contributing to tumor progression and metastasis. TME-tumor signaling actively secretes chemokines, cytokines, growth factors, and other metabolites to create a dynamic changing environment , 151. A via a PI3K/AKT-HIF1α-driven mechanism, FOXC1 in turn transcriptionally upregulates the cognate receptor of IL-8 which is CXCR1, thereby establishing a self-sustaining positive feedback loop. With regard to a different cytokine, FOXC1 overexpression robustly increased NFκB-driven luciferase activity in breast cancer MDA-MB-231 and MCF-7 cells (via 1) direct stimulation of signaling pathways that promote cancer cell growth, metastasis, and angiogenesis; 2) indirect effects, including the recruitment of CXCR4/CXCR7-positive cancer cells to CXCL12-expressing organs.Akin to the above general trend, crosstalk between FOXC1+ cancer cells and other cells of the TME has also been a prominently noted feature. Upregulated FOXC1 in tumor cells induces production and release of cytokines, chemokines and growth factors which mediates recruitment of stromal cells to the TME . Consequ-7 cells . Upregul-7 cells . Pharmac-7 cells . CXCL12 FGFR1 is a proven transcriptional target of FOXC1 in breast cancer, following its own transcriptional upregulation by TGFβ pathway activation . FGFR1 iFOXC1 has been shown to play a critical role in the development and progression of multiple malignancies. Aberrant FOXC1 expression is involved in diverse tumorigenic processes, such as abnormal cell proliferation, cancer stem cell maintenance, cancer migration, and angiogenesis. However, although FOXC1 overexpression often drives aggressive traits in a wide array of human carcinomas, the mechanisms of FOXC1 deregulation that influence the oncogenic and metastagenic processes seem specific to each tumor setting. Here we present an overview of FOXC1 and its correlation with various signal transduction pathways that have been reported in various cancers and highlight how the signal transduction-FOXC1 connection may provide an effective modality to therapeutically intervene and block the aggressive progression of FOXC1+ cancers.via the phosphoinositide 3-kinase/AKT signaling and HIF 1α. FOXC1 transcriptionally activated CXCR1 and CCL2, which promoted inflammation, invasion and metastasis  signaling (FOXC1 induces cancer stem cell (CSC) properties in BLBC cells ignaling   . IGF-1R via Smad2 and Smad3 transcription factors. Hopkins and colleagues demonstrated that FOXC1 expression was activated during TGF-β1-mediated EMT events through the binding of Smad3 proteins to a region in the FOXC1 promoter, ~800 bp upstream of the transcription start site  are a hyper active subset of enhancers, that recruit transcription factors, cofactors, and chromatin regulators to drive abundant expression of some significant genes  in the cancer context. FOXC1 is an SE-associated transcription factor that contributes to invasion, migration and metastasis in TNBC , 89. Youin vitro  and inhibited NSCLC cell tumorigenicity in vivo (Cancer stem cells (CSCs), are a small population of cancer cells that recapitulate most normal stem cell traits, but also play an essential role in tumor initiation, maintenance, progression, metastasis, drug resistance to anti-cancer drugs and metastatic recurrence. Recent studies have indicated that FOXC1, which is associated with a wide variety of cancers, is also strongly associated with mediating CSC activity . On the  in vivo . Battula in vivo . In thusChemotherapeutic drug resistance is a well-established cancer stem cell property. Mullan and colleagues were the first to show that FOXC1 acts as a mediator of drug resistance to a chemotherapeutic drug regimen comprising 5-fluorouracil/epirubicin/mitomycin C (FEM) as well as to docetaxel  Figure . OxaliplTherapeutic resistance in cancer also includes resistance to endocrine therapy with anti-estrogen drugs. In breast cancer, Cui and colleagues demonstrated that ectopic FOXC1 expression in ERα-positive MCF7 luminal breast cancer cells greatly diminished the effects of growth-stimulatory B-estradiol and growth-inhibitory antiestrogen treatment with Tamoxifen and Fulvestrant . Furtherde novo DNA methylase 3B (DNMT3B) expression, which resulted in the decrease of cysteine levels and increase reactive oxygen species (ROS) levels  expression by binding directly to the promoter regions of the FBP1 gene and negatively regulating its transcription.Metabolic reprogramming is another essential hallmark of cancer . Specifi) levels . In humaIt is evident from the above narrative that multiple signaling pathways converge upon and regulate the FOXC1 transcription factor. FOXC1, in turn, influences and coordinates multiple biological processes, again utilizing a variety of downstream signaling pathways, by which FOXC1 pro-metastatic cancers participate in the maturation of the aggressive migratory phenotype leading to metastasis. In some cases, self-sustaining positive-feedback loops are created which make their targeted interruption particularly attractive therapeutic strategies to test in the clinic. While further mechanistic elucidation of the upstream regulators and downstream mediators of FOXC1 activity are required, certain preliminary therapeutic strategies have begun to emerge based on the foundational investigations that have thus far been completed. Below we describe some of these potential strategies which hold promise and merit further consideration during the clinical trial design process.in vitro, and significantly reduced tumor growth and metastatic dissemination to the lungs in vivo. Further evidence to support such an approach was obtained in another preclinical study wherein the Wnt signaling pathway (a CSC-associated pathway) was therapeutically inhibited in an in vitro FOXC1+ cancer model  is currently underway to examine the efficacy of Ixazomib in combination with Carboplatin in previously treated advanced TNBC , Ixazomib is an oral drug with a superior toxicity profile, making it an ideal candidate for evaluating therapeutic efficacy against FOXC1+ pro-metastatic cancers by targeting the NFĸB signaling pathway ced TNBC . Marizomnografts . Marizomin vivo mouse model of BLBC  or dual inhibition of both pathways is often fraught with severely limiting toxicity issues. One potential reason is that the bulk of the preclinical data that provided the initial rationale for these trials was obtained in models of BLBC. However, at the time of patient recruitment for these trials, the TNBC criterion was employed instead. As we have learned time and time again, the BLBC and TNBC definitions are definitely not synonymous and have a widely variable degree of overlap. The assumption that TNBC status is a close enough approximation and adequate surrogate equivalent of BLBC status is known to be incorrect. This might very well be an important factor contributing to potential miscalculation of the therapeutic efficacy in these trials as BLBC or high FOXC1+ status was not employed for patient enrichment, nor to assess potential companion diagnostic utility  where, based on traditional clinical factors, a heightened recurrence risk would have been least suspected. The fact that FOXC1 expression can now be measured quite easily using an inexpensive, clinically validated and commercially available assay  means thWhile identification of elevated metastasis risk is important, it serves another purpose than the one delineated above. Not only would it allow for potentially life-extending systemic treatment (chemotherapy) to be administered to identified high risk patients, it would also allow for their targeted enrollment in the latest clinical trials. Both of these FOXC1 biomarker status-driven interventions have the potential to exert a significant decrease in overall cancer related morbidity and mortality. Beyond the above outlined points of clinical utility, however, sufficient evidence has now accrued to suggest that FOXC1 expression status might also find use to predict the efficacy of certain classes of targeted therapeutics in oncology practice. We have presented some such potentially therapeutic rationales in this review with the intention of spurring debate and stimulating discussion amongst the global oncology community. This of course will need many years of focused effort to first identify the most promising therapeutic approaches to target FOXC1+ pro-metastatic cancers, followed by carefully conducted clinical trials to test and prove their effectiveness. But if successful, such practice-changing advances hold the promise of diminishing the devastating prognostic impact of metastasis, currently the single largest contributor to cancer-related mortality, and extend the lives of cancer survivors.PR and TaR conceptualized the study. PR, TaR and TeR performed critical literature review, wrote the manuscript, critically reviewed and edited the manuscript and were collectively responsible for all manuscript content.PR, TaR, and TeR are employees of and have equity in Onconostic Technologies, Inc. and 3N Diagnostics, Ltd. PR is a named inventor on patents related to the role of FOXC1 in cancer.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Pantothenate kinase-associated neurodegeneration (PKAN) is a rare disorder associated with brain iron accumulation caused by a recessive mutation in pantothenate kinase 2 gene (PANK2). We present a case of an 11 year-old boy presenting extrapyramidal signs and developmental regression. T2-weighted images showed the classic eye of the tiger sign seen in pantothenate kinase-associated neurodegeneration. Neurodegeneration with brain iron accumulation (NBIA) is a group of diseases characterized by an abnormal accumulation of iron in the basal ganglia leading to various progressive disorders of movement and development. There are ten main types, including pantothenate kinase-associated neurodegeneration (PKAN), which is the most common form . The claAn 11 year-old boy from a consanguineous marriage with no significant past medical history presents at the age of 9 psychomotor delay, upper limb tremor, and frequent falls. There was no family history of neurological diseases. Clinical examination revealed dysarthria, spastic paraparesis with rigidity, and dystonic movements of the upper member. Laboratory investigations and EEG were normal.An initial head CT showed bilateral calcifications of the globus pallidus .A brain MRI was performed, revealing diffuse bilateral and symmetric hypointensity of both the globus pallidus with relatively central hyperintensity in T2-weighted images . These aPKAN formerly known as Hallervorden-Spatz disease is a neurodegenerative brain disorder characterized by brain iron accumulation that is caused by a recessive mutation in the pantothenate kinase 2 gene (PANK2).The PANK2 is necessary for the production of coenzyme A in mitochondria. Abnormal PANK2 function will lead to an accumulation of N-pantothenyl cysteine and free cysteine. Cysteine is a potent iron chelator and can lead to secondary iron accumulation that will exacerbate neuronal injury by inducing oxidative stress [The amount of residual enzyme activity influences the disease severity; patients with 2 null mutations (resulting in absent PANK2 enzyme) consistently have the earlier disease onset and later disease onset in individuals with mutations that allow residual enzyme activity. Globally, the 1561G>A missense mutation is the most common cause of PKAN, and homozygotes have classic PKAN [In the classical form, the first manifestations appear in infancy, between 3 and 4 years old, and progress rapidly , 3. The Atypical PKAN is characterized by later onset (age > 10 years), motor involvement tends to be less severe, but cognitive decline and psychiatric alterations are predominant traits. Speech difficulties , psychiatric problems, behavioral difficulties, or frontotemporal-like dementia early in their disease are common presenting features.Extrapyramidal dysfunction, corticospinal tract signs, and retinitis pigmentosa are also described, but they are less prevalent than in the classic form , 4, 5.On MRI, eye of the tiger sign is the typical manifestation. T2-weighted sequences show hypointensity of the globus pallidus with a central hyperintensity, reflecting an excessive accumulation of iron with gliosis.Hypointensity of the substantia nigra may be present in some cases. Eye of the tiger sign is not pathognomonic of PKAN although the majority of individuals carry PANK2 mutations , 6.The caliber and intensity of the eye are predominant in early symptomatic patients. As the disease progresses, pallidal hypointensity increases .This sign has also been reported in other diseases  and should be interpreted correctly in the clinical context. PANK2 genetic testing is the gold standard for confirming the diagnosis , 6.In our case, molecular analysis of the PANK2 gene revealed a missense homozygous mutation in exon 2.CT is not used in routine for patients suspected of PKAN. Thus, the incidence of basal ganglion calcifications may be underestimated . Eye of In our country, PKAN was first described in a 10-year-old girl born from first-degree consanguineous parents with PANK2 homozygous deletion. She had a typical PKAN phenotype presentation with extrapyramidal signs and developmental regression at the age of 7. Brain MRI performed at the age of 9 revealed the classic eye of the tiger sign. Molecular analysis identified a rare homozygous deletion in PANK2 exon 3 [Our patient is the first case in Morocco demonstrating both eye of the tiger sign on MRI and bilateral basal ganglia calcifications on CT.The natural history of classical PKAN is a nonuniform progression with intervals of stability interspersed with periods of neurological deterioration leading to early adulthood death.Atypical PKAN is less aggressive than the classic disease, and most individuals remain ambulant until adulthood. There is presently no cure. Current management strategies focus on medical and surgical symptom palliation to improve the quality of life.


Hereditary cancer syndromes are inherited pathogenic genetic variants that significantly increase the risk of developing cancer. When individuals become aware of their increased probability of having cancer, the whole family is affected by this new reality and needs to adjust. However, adjustment to hereditary cancer syndromes has been mainly studied at an individual level, and research about familial adjustment remains dispersed and disorganized. To overcome this gap, this review aims to understand how families adjust to genetic testing and risk management, and to what extent the family’s adjustment influences the psychological response and risk management behaviors of mutation carriers. We conducted searches on the PubMed/Med Line, PsycInfo, SCOPUS, and Google Scholar databases and used the Mixed Methods Appraisal Tool (MMAT-v2018) to assess the methodological quality of each selected study. Thirty studies met the inclusion criteria. Most results highlighted the interdependent nature of adjustment of pathogenic variant carriers and their families. The way carriers adjust to the syndrome is highly dependent on family functioning and related to how family members react to the new genetic information, particularly partners and siblings. Couples who share their worries and communicate openly about cancer risk present a better long-term adjustment than couples who use protective buffering  or emotional distancing. Parents need help dealing with disclosing genetic information to their children. These findings reinforce the importance of adopting a family-centered approach in the context of genetic counseling and the necessity of involving family members in research. Nearly five to ten percent of all cancers are hereditary and caused by genetic pathogenic variants that significantly increase the lifetime risk of cancer when compared to the general population . To idenTherefore, the aftermath of a positive genetic testing result commonly represents a critical event for the whole family . Not onlPrevious systematic reviews have already summarized the existing knowledge concerning the psychological adjustment of the applicant to genetic testing and counseling ,26,27,28Intending to address this gap, the present systematic review organizes current knowledge on the topic of familial adjustment to hereditary cancer syndromes in a rigorous and replicable way. Our objective is to review findings from existing research investigating the psychological adjustment of the families of pre-symptomatic pathogenic variant carriers to an increased susceptibility to hereditary cancer. Specifically, we focused on the period following a positive genetic test, including the long-term adjustment to personalized prevention programs to understand: (1) how do families adjust to a positive genetic test result of a pre-symptomatic member and his/her risk management behaviors? (2) to what extent does the family influence the psychological adjustment and risk management behaviors of the pre-symptomatic pathogenic variant carrier?This systematic review was registered with PROSPERO (ID CRD42020142200). Data extraction, critical appraisal, and qualitative synthesis followed established systematic review and qualitative synthesis methods  and the We conducted searches in the following databases: PubMed/MEDLINE, Scopus, PsycInfo, and Google Scholar between 28 June to 2 July 2019, which were updated between the 13 and 17 January 2020, and again between 23 June and 2 July 2021 and complemented with hand-searching. The search strategies combined key terms and Medical Search Headings (MESH) terms for the concepts of inherited cancer risk, unaffected pathogenic variant carrier, emotional distress, family psychosocial adjustment and risk management behavior, and personalized preventive programs, following the PICO elements . BooleanWe only included articles that: (1) were original studies; (2) described the psychosocial adjustment of the family system of pre-symptomatic pathogenic variant carriers to inherited cancer risk test and/or personalized preventive programs; (3) investigated at least one measure of psychological adjustment  to genetic testing results or risk management behaviors of carrier’s family members; (4) investigated family barriers and facilitators of decision making and adherence to personalized preventive programs/risk management behaviors. Studies targeting families of either mixed populations (affected/unaffected) or currently unaffected individuals previously diagnosed with cancer were included. We did not restrict publications based on the study design, ethnicity, or year of publication. We opted only to include studies on pre-symptomatic pathogenic variant carriers to better capture the experience of being and living with a pathogenic variant carrier, which may be different from that of being and living with a hereditary cancer patient. The methodological choice of not restricting studies by design methods aimed to aggregate all existing knowledge about the psychological adjustment of families to inherited cancer risk. Articles were excluded if: (1) pre-symptomatic pathogenic variants carriers were under the age of 18 years; (2) considered families were made up only of individuals not yet tested for inherited cancer risk; (3) considered families of affected (cancer patients) were solely pathogenic variant carriers; (4) only biomedical outcomes were considered; (5) only the psychosocial adjustment of pre-symptomatic pathogenic variant carriers was investigated, but not one of their family members.Following the search and exclusion of duplicates, two reviewers (authors GP and VB) independently screened the eligibility of the articles first by title and abstract, and the full text according to the inclusion criteria. Authors CS and PMM resolved disagreements. Authors PG and CS conducted a separate search by independently screening reference lists of previous systematic reviews ,28,30,35n = 8) [n = 1) [n = 5) [n = 1) [n = 2) [Searches of electronic databases identified 1338 reports, and searches by hand identified 16 articles. From the 1354 total records found, 21 were duplicates, and 1286 records were excluded based on information from the title and abstract. The remaining 47 articles were evaluated for inclusion by reviewing their full text and resulted in the exclusion of 17 records for the following reasons: (1) considered only the psychosocial adjustment of unaffected pathogenic variant carriers, but not of their families (n = 8) ,43,44,45 [n = 1) ; (3) did [n = 5) ,49,50,51 [n = 1) ; (5) stu [n = 2) ,54. Duri [n = 2) ,56,57. RThe mixed-methods appraisal tool (MMAT-v2018)  was usedAuthors GP and VB independently extracted the following data from included studies: first author and year of publication; country; aim of the study; study design; study setting; study outcome; follow-up points, sample size, age of applicants; gender of the applicants; clinical status of the family member; the role of the member in the family; the family role of the pathogenic variant carriers; type of pathogenic variant/cancer; psychological/behavioral outcome(s) and measure; results; retention; barriers and facilitators. The results of selected studies were summarized to provide a qualitative synthesis of the impact that genetic risk test positive results and risk management decisions have on the family psychosocial adjustment, and how the family influences the psychological response and risk management behaviors of both the pre-symptomatic pathogenic variant carrier and family members themselves.n = 5) [The distribution of MMAT-v2018 scores was generally high across study designs, with 22 out of 30 73.3%) studies meeting 100% of quality criteria and eight records having 80% of quality criteria met see . All quan = 5) ,59,60,61n = 5) ,63,64 an.3% studin = 5) ,75,76. On = 5) . No quann = 17) were conducted in the USA, while five studies were carried out in the Netherlands [We identified 30 studies meeting inclusion criteria. Most of them  predicted more depressive symptomatology in pathogenic variant carriers six months after the test result . Cancer-Four studies included in this review focused specifically on the adjustment dynamics within sibships ,74,75,78Eleven studies focused specifically on the adjustment process to test results regarding couples ,72,82,83Six studies reported on the impact of the pathogenic variant in the partner and the couple’s relationship ,68,82,83Findings from two studies suggest that approaching cancer risk as a team may benefit the couple’s adjustment ,69. StudSix studies ,60,62,64Two studies found that risk management decisions had a significant impact on the process of adaptation to life with increased cancer risk, both for the pathogenic variant carriers and for their partners ,64. StudRegarding the risk management of unaffected non-carriers, those that belong to BRCA1/2 families report overscreening behaviors, even though these were not recommended . These oThree studies reported on the experience of being a parent in a family identified with hereditary cancer syndromes ,76,77. TDifficulties in discussing cancer risk with their children also affected pathogenic variant carriers significantly . In a stOne study reported that some adult descendants of HBOC carriers viewed the likelihood of being a pathogenic variant carrier as an intense experience, eliciting internal conflicts and emotional pain . AlthougA summary of all the factors we found that affect adjustment to hereditary cancer syndromes within the family system can be found in This systematic review intended to synthesize how families with pathogenic variant carriers unaffected by cancer adjust to genetic testing results and life-saving risk management programs offered in the context of cancer genetic counseling. Taken together, the findings reveal a complex panorama where the systemic nature of family adjustment is evident. The diagnosis of a hereditary cancer syndrome in an unaffected individual works as a collective stressor, affecting not only the applicant but also their family. As posited by Daly , the herReviewed studies also highlighted that pathogenic variant carriers may adjust differently to positive genetic testing results, depending on the history and presence of the syndrome in the nuclear and extended family . These fFamily functioning and adjustment to a positive genetic test result and risk management behaviors of a pathogenic variant carrier also influenced the psychological adjustment and decision-making of new carriers. Some family dynamics  before and after genetic testing were associated with individuals’ depression , higher Although the literature is scarce, our review suggests that pathogenic variant carriers followed the example of their family members, deciding in terms of genetic testing and risk-management options. Family members might positively influence individual decisions and risk management behavior of those carrying a pathogenic variant, functioning as role models, and through sharing first-hand experiences or as interlocutors thus support the decision-making process. Testimonials and active encouragement from family members might also provide valuable information to help potential applicants decide whether to undergo genetic testing and other preventive measures ,90. NeveSupport received from family members was recognized by both non-carriers and carriers as important in dealing with the challenges posed by their condition. Emotional support in families with hereditary cancer, in which multiple members simultaneously provide and receive support from relatives, was characterized as a communal coping process ,91, as pMost of the selected studies focused on the psychological adjustment of families with inherited hereditary breast and ovarian cancer risk and HNPCC risk. Only one study explored the emotional response and role of the family in the decision-making process of positive mutation carriers of other syndromes . This may reflect a gap in the literature on the topic, but also that our search entailed a syntax that was not broad enough to encompass other hereditary cancer syndromes. Other medical terms may have been used in studies relating to other hereditary cancer syndromes. Also, not restricting studies by study design helped aggregate existing knowledge but prevented a meta-analytical approach that could yield a more precise estimate of the strength of the findings.Our review identified four major research gaps. First, there is scant knowledge about the psychological adaptation of children growing up in families with a known hereditary cancer syndrome. This calls for further research that explores how hereditary cancer affects the psychological development and health behaviors of children and adolescents, and how it should be managed to minimize negative impacts. A second major gap concerns the lack of research focused on the long-term adjustment phases of risk management. Little is known about how best to involve family members at critical turning points in risk-reduction decision-making and their adjustment to the consequences of those decisions over time. In this process, it is important to consider the limits and possible contraindications of family involvement. Such knowledge is critical to empowering the family to use its resources for mutual support.A third gap concerns the lack of cancer genetic health literacy interventions for both applicants and their relatives on the following topics: the multiple psychological impacts of hereditary cancer risk on applicants, their biological relatives, and partners; the expected impact on relationships and how to overcome it; the importance of family self-supportive dynamics , and the common difficulties associated with it . Health literacy targeting close relatives from whom the carrier expects support, such as partners, might be more important in families without a clinical history of familial cancer, given the possible devaluation of the syndrome and the carrier’s distress. Fourth and lastly, there is a lack of translational research towards family-centered genetic cancer risk care. The reviewed role of the family in the individual adjustment of pathogenic variant carriers calls for a debate about balancing the right to self-determination and individual privacy  with family-centered interventions. Hereditary cancer syndrome is a clinical condition in which several members of the same family are at different stages of diagnosis/risk management through interdependent adjustment processes and communal coping. Thus, interventions that target the family system and not just its members individually should be considered, to better address the family’s psychosocial and clinical needs. The few existing interventions, for example, Multifamily Discussion Groups (MFDG) , have shResearch should further focus on the multiple interrelated adjustment processes  to betteTo our knowledge, our review is the first to describe family member adjustments to genetic testing and risk management, and its influence on the psychological response and risk management decisions and behaviors of pathogenic variant carriers. Our findings provide valuable evidence regarding the interdependent adjustment processes of family members to the increased risk of developing cancer. Three family members emerged as particularly important for adult pathogenic variant carriers’ coping with the inherited genetic pathogenic variant: siblings, romantic partners, and children. In fact, hereditary cancer syndrome is a familial health condition, and a positive genetic testing result in a person might have implications for their close and extended relatives. Their relationships change, and pre-existing family functioning may affect not only the adjustment of the newly diagnosed pathogenic variant carrier over time but may also subsequently affect the adherence to genetic testing and the health behavior of other relatives. It is therefore important to educate and support healthcare professionals in identifying key strengths and weaknesses in individual and family adjustments for the provision of family-centered care.