Transient growth factor expression via mRNA in lipid nanoparticles promotes hepatocyte cell therapy in mice.

Primary human hepatocyte (PHH) transplantation is a promising alternative to liver transplantation, whereby liver function could be restored by partial repopulation of the diseased organ with healthy cells. However, currently PHH engraftment efficiency is low and benefits are not maintained long-term. Here we refine two male mouse models of human chronic and acute liver diseases to recapitulate compromised hepatocyte proliferation observed in nearly all human liver diseases by overexpression of p21 in hepatocytes. In these clinically relevant contexts, we demonstrate that transient, yet robust expression of human hepatocyte growth factor and epidermal growth factor in the liver via nucleoside-modified mRNA in lipid nanoparticles, whose safety was validated with mRNA-based COVID-19 vaccines, drastically improves PHH engraftment, reduces disease burden, and improves overall liver function. This strategy may overcome the critical barriers to clinical translation of cell therapies with primary or stem cell-derived hepatocytes for the treatment of liver diseases.

Transient growth factor expression via mRNA in lipid nanoparticles promotes hepatocyte cell therapy to treat murine liver diseases.

Primary human hepatocyte (PHH) transplantation is a promising alternative to liver transplantation, whereby liver function could be restored by partial repopulation of the diseased organ with healthy cells. However, currently PHH engraftment efficiency is low and benefits are not maintained long-term. Here we refine two mouse models of human chronic and acute liver diseases to recapitulate compromised hepatocyte proliferation observed in nearly all human liver diseases by overexpression of p21 in hepatocytes. In these clinically relevant contexts, we demonstrate that transient, yet robust expression of human hepatocyte growth factor and epidermal growth factor in the liver via nucleoside-modified mRNA in lipid nanoparticles, whose safety was validated with mRNA-based COVID-19 vaccines, drastically improves PHH engraftment, reduces disease burden, and improves overall liver function. This novel strategy may overcome the critical barriers to clinical translation of cell therapies with primary or stem cell-derived hepatocytes for the treatment of liver diseases.

VEGFA mRNA-LNP promotes biliary epithelial cell-to-hepatocyte conversion in acute and chronic liver diseases and reverses steatosis and fibrosis.

The liver is known for its remarkable regenerative ability through proliferation of hepatocytes. Yet, during chronic injury or severe hepatocyte death, proliferation of hepatocytes is exhausted. To overcome this hurdle, we propose vascular-endothelial-growth-factor A (VEGFA) as a therapeutic means to accelerate biliary epithelial-cell (BEC)-to-hepatocyte conversion. Investigation in zebrafish establishes that blocking VEGF receptors abrogates BEC-driven liver repair, while VEGFA overexpression promotes it. Delivery of VEGFA via nonintegrative and safe nucleoside-modified mRNA encapsulated into lipid nanoparticles (mRNA-LNPs) in acutely or chronically injured mouse livers induces robust BEC-to-hepatocyte conversion and elimination of steatosis and fibrosis. In human and murine diseased livers, we further identified VEGFA-receptor KDR-expressing BECs associated with KDR-expressing cell-derived hepatocytes. This work defines KDR-expressing cells, most likely being BECs, as facultative progenitors. This study reveals unexpected therapeutic benefits of VEGFA delivered via nucleoside-modified mRNA-LNP, whose safety is widely validated with COVID-19 vaccines, for harnessing BEC-driven repair to potentially treat liver diseases.

Growth Hormone Accelerates Recovery From Acetaminophen-Induced Murine Liver Injury.

Background and Aims: Acetaminophen (APAP) overdose is the leading cause of acute liver failure, with one available treatment, N-acetyl cysteine (NAC). Yet, NAC effectiveness diminishes about ten hours after APAP overdose, urging for therapeutic alternatives. This study addresses this need by deciphering a mechanism of sexual dimorphism in APAP-induced liver injury, and leveraging it to accelerate liver recovery via growth hormone (GH) treatment. GH secretory patterns, pulsatile in males and near-continuous in females, determine the sex bias in many liver metabolic functions. Here, we aim to establish GH as a novel therapy to treat APAP hepatotoxicity. Approach and Results: Our results demonstrate sex-dependent APAP toxicity, with females showing reduced liver cell death and faster recovery than males. Single-cell RNA sequencing analyses reveal that female hepatocytes have significantly greater levels of GH receptor expression and GH pathway activation compared to males. In harnessing this female-specific advantage, we demonstrate that a single injection of recombinant human GH protein accelerates liver recovery, promotes survival in males following sub-lethal dose of APAP, and is superior to standard-of-care NAC. Alternatively, slow-release delivery of human GH via the safe nonintegrative lipid nanoparticle-encapsulated nucleoside-modified mRNA (mRNA-LNP), a technology validated by widely used COVID-19 vaccines, rescues males from APAP-induced death that otherwise occurred in control mRNA-LNP-treated mice. Conclusions: Our study demonstrates a sexually dimorphic liver repair advantage in females following APAP overdose, leveraged by establishing GH as an alternative treatment, delivered either as recombinant protein or mRNA-LNP, to potentially prevent liver failure and liver transplant in APAP-overdosed patients.

VEGFA mRNA-LNP promotes biliary epithelial cell-to-hepatocyte conversion in acute and chronic liver diseases and reverses steatosis and fibrosis.

The liver is known for its remarkable regenerative ability through proliferation of hepatocytes. Yet, during chronic injury or severe hepatocyte death, proliferation of hepatocytes is exhausted. To overcome this hurdle, we propose vascular-endothelial-growth-factor A (VEGFA) as a therapeutic means to accelerate biliary epithelial cell (BEC)-to-hepatocyte conversion. Investigation in zebrafish establishes that blocking VEGF receptors abrogates BEC-driven liver repair, while VEGFA overexpression promotes it. Delivery of VEGFA via non-integrative and safe nucleoside-modified mRNA encapsulated into lipid-nanoparticles (mRNA-LNP) in acutely or chronically injured mouse livers induces robust BEC-to-hepatocyte conversion and reversion of steatosis and fibrosis. In human and murine diseased livers, we further identified VEGFA-receptor KDR-expressing BECs associated with KDR-expressing cell-derived hepatocytes. This defines KDR-expressing cells, most likely being BECs, as facultative progenitors. This study reveals novel therapeutic benefits of VEGFA delivered via nucleoside-modified mRNA-LNP, whose safety is widely validated with COVID-19 vaccines, for harnessing BEC-driven repair to potentially treat liver diseases. Highlights: Complementary mouse and zebrafish models of liver injury demonstrate the therapeutic impact of VEGFA-KDR axis activation to harness BEC-driven liver regeneration.VEGFA mRNA LNPs restore two key features of the chronic liver disease in humans such as steatosis and fibrosis.Identification in human cirrhotic ESLD livers of KDR-expressing BECs adjacent to clusters of KDR+ hepatocytes suggesting their BEC origin.KDR-expressing BECs may represent facultative adult progenitor cells, a unique BEC population that has yet been uncovered.

Influenza Vaccinations Among Privately and Publicly Insured Children With Asthma.

OBJECTIVE: Annual influenza vaccination rates for children remain well below the Healthy People 2030 target of 70%. We aimed to compare influenza vaccination rates for children with asthma by insurance type and to identify associated factors. METHODS: This cross-sectional study examined influenza vaccination rates for children with asthma by insurance type, age, year, and disease status using the Massachusetts All Payer Claims Database (2014-2018). We used multivariable logistic regression to estimate the probability of vaccination accounting for child and insurance characteristics. RESULTS: The sample included 317,596 child-year observations for children with asthma in 2015-18. Fewer than half of children with asthma received influenza vaccinations; 51.3% among privately insured and 45.1% among Medicaid insured. Risk modeling reduced, but did not eliminate, this gap; privately insured children were 3.7 percentage points (pp) more likely to receive an influenza vaccination than Medicaid-insured children (95% confidence interval [CI]: 2.9-4.5pp). Risk modeling also found persistent asthma was associated with more vaccinations (6.7pp higher; 95% CI: 6.2-7.2pp), as was younger age. The regression-adjusted probability of influenza vaccination in a non-office setting was 3.2pp higher in 2018 than 2015 (95% CI: 2.2-4.2pp), and significantly lower for children with Medicaid. CONCLUSIONS: Despite clear recommendations for annual influenza vaccinations for children with asthma, low rates persist, particularly for children with Medicaid. Offering vaccines in non-office settings such as retail pharmacies may reduce barriers, but we did not observe increased vaccination rates in the first years after this policy change.

Update on Hepatobiliary Plasticity.

The liver field has been debating for decades the contribution of the plasticity of the two epithelial compartments in the liver, hepatocytes and biliary epithelial cells (BECs), to derive each other as a repair mechanism. The hepatobiliary plasticity has been first observed in diseased human livers by the presence of biphenotypic cells expressing hepatocyte and BEC markers within bile ducts and regenerative nodules or budding from strings of proliferative BECs in septa. These observations are not surprising as hepatocytes and BECs derive from a common fetal progenitor, the hepatoblast, and, as such, they are expected to compensate for each other's loss in adults. To investigate the cell origin of regenerated cell compartments and associated molecular mechanisms, numerous murine and zebrafish models with ability to trace cell fates have been extensively developed. This short review summarizes the clinical and preclinical studies illustrating the hepatobiliary plasticity and its potential therapeutic application.

Content Analysis of Patient-Facing Information Related to Preeclampsia.

BACKGROUND: Previous research has shown pregnant people are not knowledgeable about preeclampsia, a significant cause of maternal morbidity and mortality. This lack of knowledge may impact their ability to report symptoms, comply with recommendations, and receive appropriate follow-up care. Pregnant people commonly seek information from sources outside their treating clinician, including pregnancy-specific books and online sources. We examined commonly used preeclampsia information sources to evaluate whether pregnant people are receiving up-to-date, guideline-based information. METHODS: We conducted a content analysis of preeclampsia-related information in top-ranking websites and bestselling pregnancy books. We used American College of Obstetricians and Gynecologists preeclampsia guidelines to construct a codebook, which we used to examine source content completeness and accuracy. For each source, we analyzed information about preeclampsia diagnosis, patient-reported symptoms, risk factors, prevention, treatment, and complications. RESULTS: Across 19 included sources (13 websites and 6 books), we found substantial variation in completeness and accuracy of preeclampsia information. We found high rates of mentions for preeclampsia symptoms. Risk factors were more commonly included in online sources than book sources. Most sources mentioned treatment options, including blood pressure medication and giving birth; however, one-third of online sources positively mentioned the nonrecommended treatment of bed rest. Prevention methods, including prenatal aspirin for high-risk pregnancies, and long-term complications of preeclampsia were infrequently mentioned. CONCLUSIONS: Varying rates of accuracy in patient-facing preeclampsia information mean there is substantial room for improvement in these sources. Ensuring pregnant people receive current guideline-based information is critical for improving outcomes and implementing shared decision-making.

c-Maf: The magic wand that turns on LSEC fate.

In this issue of Cell Stem Cell, Gómez-Salinero et al. (2022) identify c-Maf as a driver for murine liver sinusoidal endothelial cell (LSEC) fate and function during liver development, homeostasis, and repair. Similarly, c-Maf defines human LSECs, and its overexpression specializes generic HUVECs into functional induced-LSECs, potentiating regenerative therapeutics.

Differences and similarities among questionnaires to assess pain status in chronic widespread pain population: a quantitative analysis.

OBJECTIVE: In clinical practice, multiple questionnaires are often used as part of the diagnosis of chronic widespread pain. Body Surface Area (BSA), Visual Analogue Scale (VAS), Fibromyalgia Diagnostic Criteria (FDC) and Central Sensitization Inventory (CSI) have all been used as screening tools to assess pain status in individuals with widespread pain. However, substantial overlap can be observed among these commonly employed questionnaires. This study aimed to quantitatively determine the most independent and dependent clinical characteristics obtained through these questionnaires and to examine potential redundancies. METHODS: Seventy-nine participants with widespread pain, 61 females and 18 males, from a chronic pain outpatient clinic were recruited. The FDC, BSA, VAS and the CSI were measured for all participants. A principal component analysis (PCA) using a varimax rotation was used to determine which clinical measures represented separate constructs of widespread pain. This was followed by a regression analysis to assess redundancy between the constructs and related pain characteristics. RESULTS: The identified three-component PCA solution was characterized by (1) the FDC and CSI score, (2) the VAS score and (3) the BSA score. This indicates that the BSA and the VAS scores capture independent patient information. From the regression analysis, the FDC and CSI scores shared approximately 80% of the variance, indicative of substantial overlap between scores. CONCLUSION: Our findings demonstrated that BSA and VAS scores were independent clinical measures of widespread chronic pain, while the FDC and CSI scores were not independent, were highly correlated and provided redundant information. Clinicians should continue using both the BSA and VAS; however, either only FDC or CSI will be beneficial during clinical assessment of widespread chronic pain.

Transient yet Robust Expression of Proteins in the Mouse Liver via Intravenous Injection of Lipid Nanoparticle-encapsulated Nucleoside-modified mRNA.

With the recent availability of the SARS-CoV-2 mRNA-based vaccines, public attention has been drawn to this new technology and how it may be applied to other indications. Temporal activation of key hepatic regenerative pathways can induce liver regeneration, overcoming the lack of donor organs for liver transplantation and ineffectiveness of alternative treatments. Recombinant protein therapies and genetic therapies that target these pathways require frequent and repeated injections or, when integrated into the genome, may lead to deleterious effects. In contrast, nucleoside-modified mRNA encapsulated in lipid nanoparticles (mRNA-LNP) are non-integrative and induce transient yet robust expression of proteins that could serve as an ideal therapeutic tool to treat specific liver diseases. For instance, our recent publication in Nature Communications used mRNA-LNP to express hepatic mitogens, hepatocyte growth factor, and epidermal growth factor to induce liver regeneration following both acute and chronic liver injuries. Initial testing with firefly luciferase mRNA-LNP transfection and in vivo imaging confirmed specific hepatotropic delivery. In this protocol, we describe in detail the necessary steps to deliver mRNA-LNP to the murine liver and, following intravenous injection of eGFP mRNA-LNP, verify transfection efficiency using flow cytometry and liver cell specificity using immunofluorescence analyses. This procedure presents an unprecedented tool that can be customized with mRNA-LNP encoding any protein of interest to be expressed by virtually all hepatocytes, ~70% endothelial cells, and ~40% Kupffer cells for promoting liver function and/or regeneration. Graphic abstract: Experimental Design of mRNA-LNP IV Injection and Analysis of Liver Cell Specificity and Efficiency of Transfection (Created with BioRender.com).

A randomized double blinded placebo controlled study to evaluate motor unit abnormalities after experimentally induced sensitization using capsaicin.

Central sensitization is a condition that represents a cascade of neurological adaptations, resulting in an amplification of nociceptive responses from noxious and non-noxious stimuli. However, whether this abnormality translates into motor output and more specifically, ventral horn abnormalities, needs to be further explored. Twenty healthy participants aged 20-70 were randomly allocated to topical capsaicin or a placebo topical cream which was applied onto their left upper back to induce a transient state of sensitization. Visual analogue scale (VAS) ratings of pain intensity and brush allodynia score (BAS) were used to determine the presence of pain and secondary allodynia. Surface electromyography (sEMG) and intramuscular electromyography (iEMG) were used to record motor unit activity from the upper trapezius and infraspinatus muscles before and twenty minutes after application of capsaicin/placebo. Motor unit recruitment and variability were analyzed in the sEMG and iEMG, respectively. An independent t-test and Kruskal-Wallis H test were performed on the data. The sEMG results demonstrated a shift in the motor unit recruitment pattern in the upper trapezius muscle, while the iEMG showed a change in motor unit variability after application of capsaicin. These results suggest that capsaicin-induced central sensitization may cause changes in ventral horn excitability outside of the targeted spinal cord segment, affecting efferent pathway outputs. This preclinical evidence may provide some explanation for the influence of central sensitization on changes in movement patterns that occur in patients who have pain encouraging of further clinical investigation.Clinical Trials registration number: NCT04361149; date of registration: 24-Apr-2020.

Potential Role of MRI Imaging for Myofascial Pain: A Scoping Review for the Clinicians and Theoretical Considerations.

The most common cause of chronic musculoskeletal pain is chronic myofascial pain syndrome (MPS). MPS often presents with increased muscle stiffness, and the myofascial trigger point (MTrP). Imaging modalities have been used to identify the MTrP, but their role in the detection and diagnosis of MPS remains unclear. The purpose of this review was to identify evidence in literature for the use of imaging in the role of classifying and explaining the physiology of MTrPs. Since few imaging techniques have been performed on MTrPs, we explored the imaging techniques that can effectively image complex skeletal muscle microstructure, and how they could be used. As part of a scoping review, we conducted a systematic search from three medical databases (CINAHL, EMBASE and MEDLINE) from year to year to analyze past MTrP imaging, as well as analyzing imaging techniques performed on the microstructure of muscle. Previously, ultrasound has been used to differentiate active, latent MTrPs, but these studies do not adequately address their underlying anatomical structure. MRI remains the standard method of imaging skeletal muscle. The existing MRI literature suggests that the DTI technique can quantify muscle injury, strain, and structure. However, theoretically, HARDI and DKI techniques seem to provide more information for complex structural areas, although these modalities have a disadvantage of longer scan times and have not been widely used on skeletal muscle. Our review suggests that DTI is the most effective imaging modality that has been used to define the microstructure of muscle and hence, could be optimal to image the MTrP. HARDI and DKI are techniques with theoretical potential for analysis of muscle, which may provide more detailed information representative of finer muscle structural features. Future research utilizing MRI techniques to image muscle are necessary to provide a more robust means of imaging skeletal muscle and the MTrP.

Murine liver repair via transient activation of regenerative pathways in hepatocytes using lipid nanoparticle-complexed nucleoside-modified mRNA.

Induction of intrinsic liver regeneration is an unmet need that can be achieved by temporally activating key hepatocyte regenerative pathways. Here, we establish an efficient, safe, non-integrative method to transiently express hepatocyte-growth-factor (HGF) and epidermal-growth-factor (EGF) in hepatocytes via nucleoside-modified, lipid-nanoparticle-encapsulated mRNA (mRNA-LNP) delivery in mice. We confirm specific hepatotropism of mRNA-LNP via intravenous injection of firefly luciferase encoding mRNA-LNP, with protein expression lasting about 3 days. In the liver, virtually all hepatocytes are transfected along with a subpopulation of endothelial and Kupffer cells. In homeostasis, HGF mRNA-LNP efficiently induce hepatocyte proliferation. In a chronic liver injury mouse model recapitulating non-alcoholic fatty liver disease, injections of both HGF and EGF mRNA-LNP sharply reverse steatosis and accelerate restoration of liver function. Likewise, HGF and EGF mRNA-LNP accelerate liver regeneration after acetaminophen-induced acute liver injury with rapid return to baseline ALT levels. This study introduces mRNA-LNP as a potentially translatable safe therapeutic intervention to harness liver regeneration via controlled expression of endogenous mitogens in vivo.

Quantitative response of healthy muscle following the induction of capsaicin: an exploratory randomized controlled trial.

BACKGROUND: Myofascial pain syndrome (MPS) is a prevalent chronic pain disorder primarily characterized by myofascial trigger points (MTrPs). There is limited knowledge on the pathophysiology and mechanisms underlying MTrP and its development. Research has previously demonstrated the identification of MTrPs using ultrasound and vibration sonoelastography, although there is some contradictory evidence regarding if MTrPs present as hyper or hypoechoic regions. Electromyography (EMG) investigations of MTrP have demonstrated that MTrPs are usually located proximal to innervation zones where the peak surface EMG signals are obtained from. Central sensitization has been proposed as the primary mechanism underlying MTrP development. Central sensitization is associated with hyperexcitability of neuronal responses to normal or noxious stimuli. There is a need for a study that measures ultrasound image textural changes and motor unit activity responses in the muscle following sensitization. The purpose of this study is to determine whether sensitizing healthy muscle using capsaicin induces a regional change in image texture variables within the specific and surrounding muscles, as well as the motor unit frequency and amplitude changes that accompany them. This is an exploratory trial that aims to provide preliminary evidence on whether central sensitization is a direct cause of taut band and MTrP development. METHODS: Ethical approval was obtained from the University Health Network (UHN) Research Ethics Board. This proposed study is a single centered, factorial, randomized placebo-controlled trial with two independent variables, depth of capsaicin application and dose of capsaicin, for a total of six treatment arms and three control treatment groups. DISCUSSION: This will be the first study that assesses the B-mode ultrasound image texture of induced sensitized muscles and will provide more evidence on muscle motor unit activity and regional changes of central sensitization. Findings from this study may support one of few hypotheses proposed delineating the involvement of central sensitization in the development of trigger points. TRIAL REGISTRATION: National Institutes of Health ClinicalTrials.gov NCT03944889 . Registered on May 07, 2019.

Machine Learning Diagnostic Modeling for Classifying Fibromyalgia Using B-mode Ultrasound Images.

Fibromyalgia (FM) diagnosis remains a challenge for clinicians due to a lack of objective diagnostic tools. One proposed solution is the use of quantitative ultrasound (US) techniques, such as image texture analysis, which has demonstrated discriminatory capabilities with other chronic pain conditions. From this, we propose the use of image texture variables to construct and compare two machine learning models (support vector machine [SVM] and logistic regression) for differentiating between the trapezius muscle in healthy and FM patients. US videos of the right and left trapezius muscle were acquired from healthy (n = 51) participants and those with FM (n = 57). The videos were converted into 64,800 skeletal muscle regions of interest (ROIs) using MATLAB. The ROIs were filtered by an algorithm using the complex wavelet structural similarity index (CW-SSIM), which removed ROIs that were similar. Thirty-one texture variables were extracted from the ROIs, which were then used in nested cross-validation to construct SVM and elastic net regularized logistic regression models. The generalized performance accuracy of both models was estimated and confirmed with a final validation on a holdout test set. The predicted generalized performance accuracy of the SVM and logistic regression models was computed to be 83.9 ± 2.6% and 65.8 ± 1.7%, respectively. The models achieved accuracies of 84.1%, and 66.0% on the final holdout test set, validating performance estimates. Although both machine learning models differentiate between healthy trapezius muscle and that of patients with FM, only the SVM model demonstrated clinically relevant performance levels.

Endothelial cells instruct liver specification of embryonic stem cell-derived endoderm through endothelial VEGFR2 signaling and endoderm epigenetic modifications.

Liver organogenesis requires complex cell-cell interactions between hepatic endoderm cells and adjacent cell niches. Endothelial cells are key players for endoderm hepatic fate decision. We previously demonstrated that the endothelial cell niche promotes hepatic specification of mouse embryonic stem cell(ESC)-derived endoderm through dual repression of Wnt and Notch pathways in endoderm cells. In the present study, we dissected further the mechanisms by which endothelial cells trigger endoderm hepatic specification. Using our previously established in vitro mouse ESC system mimicking the early hepatic specification process, endoderm cells were purified and co-cultured with endothelial cells to induce hepatic specification. The comparison of transcriptome profiles between hepatic endoderm cells isolated from co-cultures and endoderm cells cultured alone revealed that VEGF signaling instructs hepatic specification of endoderm cells through endothelial VEGFR2 activation. Additionally, epigenetic mark inhibition assays upon co-cultures uncovered that histone acetylation and DNA methylation promote hepatic specification while histone methylation inhibits it. This study provides an efficient 2D platform modelling the endothelial cell niche crosstalk with endoderm, and reveals mechanisms by which endothelial cells promote hepatic specification of mouse ESC-derived endoderm cells through endothelial VEGFR2 activation and endoderm epigenetic modifications.

Adequacy of clinical trial evidence of metformin fixed-dose combinations for the treatment of type 2 diabetes mellitus in India.

There is growing national and international concern about the drug regulatory system in India. Parliamentary reports have highlighted the presence of high numbers of unapproved medicines and irrational combinations of both approved and unapproved drugs in the Indian market-place. Fixed-dose combinations (FDCs) are a peculiar feature of the Indian pharmaceutical landscape. Although metformin is a first-line treatment, FDCs for diabetes in India account for two-thirds of all diabetes medicine sales, and some have not been approved by the Central Drugs Standard Control Organization (CDSCO). This study examines the basis of efficacy and safety of top-selling metformin FDCs in India against four WHO criteria from clinical trials guidelines for the approval of FDCs. Data from a commercial drug sales database (PharmaTrac) were combined with searches through published literature, clinical trial registries, and published and unpublished trial websites of metformin FDCs in adults with type 2 diabetes mellitus. Five metformin FDCs in India from November 2011 to October 2012 accounted for 80% of all metformin FDC sales by value and volume. Although all five had obtained CDSCO approval, three had been sold and marketed prior to receiving this approval. Evaluation of published and unpublished clinical trials of these five FDCs found none provided robust evidence of safety and efficacy for the treatment of type 2 diabetes. Recommendations are made for publishing evidence that underpins drug approvals, marketing bans, greater transparency through updated clinical trials databases and legislative reform in order to prevent irrational FDCs from entering the market.