Since the etiology of numerous human diseases has been associated with protein aggregation, and it has become a field of increasing importance in the biopharmaceutical industry, the biophysical characterization of protein misfolded states and their aggregation mechanisms continues to receive increased attention. Mass spectrometry (MS) has firmly established itself as a powerful analytical tool capable of both detection and characterization of proteins at all levels of structure. Given inherent advantages of biological MS, including high sensitivity, rapid timescales of analysis, and the ability to distinguish individual components from complex mixtures with unrivalled specificity, it has found widespread use in the study of protein aggregation, importantly, where traditional structural biology approaches are often not amenable. The present review aims to provide a brief overview of selected MS-based approaches that can provide a range of biophysical descriptors associated with protein conformation and the aggregation pathway. Recent examples highlight where this technology has provided unique structural and mechanistic understanding of protein aggregation.Protein aggregation is now recognized as a generic and significant component of the protein energy landscape. Occurring through a complex and dynamic pathway of structural interconversion, the assembly of misfolded proteins to form soluble oligomers and insoluble aggregates remains a challenging topic of study, both  Proteins rely on a high degree of conformational flexibility to fold into their native states and facilitate the extensive interactions with a wide variety of binding partners that underlie biological function. Reaching and maintaining such native structures is enabled by a downhill-folding pathway on a free-energy surface. However, the near cancellation of enthalpic and entropic contributions to free energy during the folding process, and the low energy barriers to structural interconversion, mean that the flexibility required for a protein to function often places them at the brink of stability . ProteinProtein aggregation refers to the multistep process by which abnormal association of proteins gives rise to larger aggregate structures, which tend to be insoluble. This process commences with the self-assembly of misfolded monomers to produce a range of intermediates termed soluble-misfolded oligomers. Further assembly from such early oligomeric states leads to formation of insoluble structures in the forms of amorphous aggregates or amyloid fibrils with ordered β-sheet conformations ,8 Figur. This caGiven increasing recognition of the importance of protein aggregation in biology, significant research effort has been focused on enabling detailed structural and mechanistic understanding of the aggregation pathway. However, this is a particularly complex and challenging task given the heterogeneous and dynamic nature of the aggregation process. Many biophysical methods have been employed for the study of protein misfolding and aggregation . Importam/z) [From the most fundamental perspective, an MS experiment involves the conversion of molecules into gas-phase ions, where they are separated in a mass analyzer according to their mass-to-charge ratios (m/z) . For prom/z) . A numbem/z) . Perhapsm/z) .m/z as a basis of protein identification from biological samples. The measurement of molecular weight for intact proteins and their assemblies can enable the identification of stoichiometry to provide the number of monomeric units in an oligomeric structure. Furthermore, as native ESI-MS allows for preservation of noncovalent-binding interactions, the association of small-molecule cofactors, ligands, or chaperones with aggregating proteins may also be observed. Beyond simple molecular weight measurement however, extensive structural information can be provided through a range of coupled techniques, with key examples further described below. On this basis, MS has emerged as a highly sensitive technique for protein aggregation studies, offering complementary insights into the onset of amorphous aggregation and particularly amyloid fibril formation, including monomer and oligomer structural characterization, aggregation mechanisms, and inhibitor screening.In the context of protein aggregation, if the protein/oligomer is enzymatically digested before MS analysis, then the resulting peptides can be separated according to their m/z or a single species has different conformational states, multidimensional separation is critical. Ion mobility (IM) coupled to MS provides a means for further gas-phase separation of ions based on the rate at which they migrate through a neutral carrier gas under the influence of an external electric field. Importantly, the mobility of an ion can be related to rotationally averaged collision cross-section (CCS), and therefore can be used to infer information regarding biomolecular structure [While native MS serves as a key method for the detection of intact protein assemblies, in cases where oligomeric complexes may overlap at the same tructure . IM-MS tn or ‘product-ion’) spectra, which contain rich structural information [Tandem MS allows selected ions to be isolated based on a first MS or IM analysis step, then submitted to activation and/or fragmentation. The resulting products are subsequently analysed to give MS/MS  coupled with MS provides an additional method to characterize 3D conformations of proteins and identify protein-binding interfaces. In a basic CX-MS workflow, bifunctional small-molecule cross-linking reagents are used to covalently link two functional sites on the protein, typically with some defined reactivity toward specific amino acids. Since the links are made by a spacer with distinct length, localizing these linkage sites using proteomics methods provides an upper distance constraint to probe tertiary and quaternary structures . As CX-Min situ are based on limited proteolysis (LiP) and stability profiling. LiP-MS identifies structural changes in different cellular states or under different conditions by quantifying the relative sensitivity of proteins to a given protease through structure-dependent proteolytic patterns [The advantages of MS that enable investigation of complex protein mixtures mean that it is increasingly being exploited to study structural and biophysical properties of proteins at a whole proteome scale. Of note, two additional MS approaches that probe proteome structural alterations patterns ,34. In cpatterns ,36. In tDue to their dynamic, polydisperse, and polymorphic nature, aggregating proteins are particularly challenging to characterize using traditional structural biology methods. Here, recent studies are selected from among the many of this expansive field, to showcase the unique ability of MS in characterizing the underlying assembly pathway for a broad range of amyloidogenic and amorphously aggregating proteins. Furthermore, these studies outline the potential of MS as a tool for the screening of aggregation inhibitors and quality control monitoring.The mechanisms by which proteins fold and misfold in solution remain the subject of significant study and debate. Disruption of native state(s) can occur because of many influences, and the early conformational changes from native structure have traditionally monitored in solution using a variety of techniques. However, IM-MS has proven invaluable to investigate the conformational landscape of many proteins ,38 and tA range of IM-MS studies have focused on early conformational changes of amyloidogenic proteins, showing a structural reorganization from extended structures of the intrinsically disordered proteins clinically implicated in neurodegenerative disorders to more compact states which precede aggregation ,43. SimiPulse-labeling HDX-MS also enables direct monitoring of rapid events such as the earliest stages of protein misfolding, with high structural and temporal resolution. For example, this technique has provided enhanced understanding of the mechanism of aggregation for the clinically important proteins amyloid-β, α-synuclein, and tau . By moni105–115), describing oligomeric states and aggregation over time. For example, the NNQQNY peptide was shown to form extended, likely β-sheet-rich structures, for oligomers beyond octamers, representing a distinct transition toward the amyloid pathway from that point [Growing evidence suggests that the soluble oligomers on pathway to large aggregates are primary contributors to cell toxicity, particularly in the context of the diverse range of amyloid disorders . Once agat point . In compat point . Amyloidat point . One sugat point .Kinetic-pulsed variations of HDX have also been used to characterize early amyloidogenic intermediates , enablinm/z and charge [Given extensive characterization by X-ray crystallography, nuclear magnetic spectroscopy and, more recently, cryoelectron microscopy, the structure of mature amyloid fibrils is largely well understood. Nevertheless, while gas-phase analysis of amyloid fibrils is particularly challenging due to their enormous size, MS is still able to further probe the properties of such mature assemblies. For example, molecular-weight determination is the most direct way of determining the aggregation state of a protein. Recent applications of charge-detection MS, a single-molecule method where the mass of each ion is directly determined from individual measurements of d charge , demonstd charge ,60. Thisd charge ,62. As td charge .A range of MS-based methods are able to observe and interrogate binding interactions of protein–ligand complexes . Often, While much focus in the protein aggregation field has been directed toward amyloid aggregation due to impacts in human health, amorphous aggregation is also of significant note in a biopharmaceutical context. Here, aggregates must be strictly controlled as they are not only less efficacious but resultant particulates may cause serious immunological responses when administered . Liquid Although there are many different approaches to study protein aggregation, MS has firmly established itself as critical to this pursuit based on the diversity of information afforded. It is clear from the proliferation of protein aggregation studies reported in recent decades, which employ MS methodologies that it can reveal many previously hidden aspects of protein misfolding and aggregation across the entire structural landscape. Importantly, a key strength of MS is its ability to further combine with orthogonal techniques. Continued advancements in MS technology and an ongoing drive toward integrated structural biology, where multiple methods combine to contribute to a deep understanding of protein structure and dynamics, will lead to a rich knowledge of the fundamental factors that modulate aggregation. We might ultimately see MS-driven technologies routinely describe protein aggregation states across the whole proteome and under a wide diversity of conditions, providing further opportunities to intervene to the advantage of human health and biotechnology.Protein aggregation occurs via a complex, multistep pathway involving aberrant assembly of misfolded proteins.MS offers unique advantages for the biophysical analysis of heterogeneous and dynamic processes involved in protein misfolding and aggregation.Utilizing MS-based detection with various experimental approaches, information regarding structural changes, stability, self-assembly, and other molecular interactions and dynamics can be captured along the entire protein aggregation pathway.MS-based understanding of protein aggregation can have broad and impact across areas of health and biotechnology.


Human cytomegalovirus (HCMV) remains an essential global concern due to its distinct life cycle, mutations and latency. As HCMV is a herpesvirus, it establishes a lifelong persistence in the host through a chronic state of infection. Immunocompromised individuals are at risk of significant morbidity and mortality from the virus. Until now, no effective vaccine has been developed to combat HCMV infection. Only a few antivirals targeting the different stages of the virus lifecycle and viral enzymes are licensed to manage the infection. Therefore, there is an urgent need to find alternate strategies to combat the infection and manage drug resistance. This review will provide an insight into the clinical and preclinical antiviral approaches, including HCMV antiviral drugs and nucleic acid-based therapeutics. Human cytomegalovirus (HCMV) is an ubiquitous virus either existing quiescently in healthy persons or causing severe disease in immunocompromised patients and congenitally infected newborns. To date, eight herpesviruses have been identified that are known to have humans as primary hosts. According to ICTV , they have been named as Human herpesvirus 1–8   and sharHCMV is a large DNA virus that belongs to the genus of Cytomegalovirus and the betaherpesvirinae subfamily . The virThe large DNA virus is arranged in a class E structure, with two unique regions (unique long UL and unique short US). These two regions are both flanked by a pair of inverted repeats . The TRL and TRS share a region of a few hundred bps which is called “a sequence”; the other regions of the repeats are usually referred to as “b sequence” and “c sequence”. The genome exists as an equimolar mixture of four genomic isomers by inversion of UL and US regions . There aFurthermore, the emergence of drug resistance and dose-limiting toxicities limit anti-viral treatments for HCMV infections and diseases. In addition, there are no approved vaccines or immunoglobulins available to prevent congenital HCMV infection. However, intravenous immunoglobulin (IGIV) preparation such as Cytogam have been used to treat certain HCMV infection, particularly in immunocompromised patients such as transplant recipients . The useCurrently, only a few antiviral drugs are approved for HCMV infection .Ganciclovir (GCV) or its prodrug valganciclovir (VGCV), an acyclic nucleoside analog, is the preferred antiviral agent for the treatment of confirmed HCMV disease. In 1987, Erice et al. first reported that ganciclovir showed antiviral activity against HCMV in patients with depressed immunity and bone marrow and organ transplant recipients . PhosphoAmong nucleotide analogs, cidofovir (CDV), which is administered intravenously, was the first to receive FDA approval. It is mostly used to treat HCMV infection in patients with AIDS and in transplant recipients. CDV is a monophosphate nucleotide analog . ThroughFoscarnet is an inorganic pyrophosphate with a broad spectrum of activity. The drug works by binding reversibly near the pyrophosphate-binding site of viral DNA poly-merase pUL54 and prevents the cleavage of pyrophosphate from the deoxynucleotide triphosphate, in turn halting chain elongation. Two important uses of foscarnet are in the early stage following hematopoietic stem cell transplant due to comparatively less suppression of bone marrow than GCV; and in treating GCV-resistant HCMV infection in transplant recipients and HIV/AIDS patients. It is available as an IV formulation and like GCV, it is also used intravitreously for treatment of HCMV retinitis.Letermovir is FDA approved for prophylaxis of HCMV infection and disease in adult HCMV-seropositive recipients . It is pMaribavir is a potent, selective, orally bioavailable benzimidazole riboside that is active against CMV infection in humans. This drug works by inhibiting the UL97 kinase enzyme, which plays a critical role in replication by phosphorylating proteins in-volved in the assembly of the viral capsid . MaribavRecently, a few neuroactive mood stabilizers are reported to be potent antivirals against HCMV. Valproate is a widely prescribed anti-epileptic drug employed to treat multiple psychiatric and neurological diseases. A study by Ornaghi et al., first reported that Valpromide (VPD) and Valnoctamide (VCD) can inhibit both human and murine CMV both in vitro or in vivo . The stuLeflunomide is a pyrimidine synthesis inhibitor belonging to the DMARD (disease-modifying antirheumatic drug) class of drugs originally indicated for treating rheumatoid arthritis. It is cheap and easily available and shown to have anti-HCMV properties both in vitro and in vivo. It also has been used effectively for HCMV which has reactivated after renal transplant  as well A critical issue with all of these treatments is that HCMV can develop resistance to antiviral drugs, especially in immunocompromised individuals, including AIDS patients and those who have received prolonged antiviral treatment. In several cases of HCMV treated with GCV/VGCM, antiviral drug resistance has also been reported. Mutation in the viral protein kinase gene (UL97) and the DNA polymerase gene (UL54) confers resistance to GCV/VGCV  57,58,5,558,59. HCMV is a complex virus that has evolved many mechanisms to evade the immune system, making it challenging to develop an effective vaccine. In order to prevent infection and its associated costs, it has been considered the highest priority to develop a vaccine against CMV. HCMV vaccines were first developed in the 1970s when AD169 and Towne strains were attenuated to function as active immunity boosters ,61. The Despite these challenges, various vaccine candidates are currently under clinical trial, including live-attenuated vaccines, subunit vaccines, viral vector vaccines, chimeric peptide vaccines, vaccines based on enveloped virus-like particles, plasmid-based DNA vaccines, RNA-based vaccines and peptide vaccines . Current′-untranslated region (3′-UTR) of the mRNA molecules, inhibiting their translation into protein. An 18–25 nt synthetic siRNA/miRNA strand could specifically bind to a target region and cause gene silencing . Th. Th89]. Antisense molecules are promising gene-targeting technologies for the treatment of various viral infections. Fomivirsen, an antisense oligonucleotide-based drug, was developed to treat HCMV in AIDS patients with . This syAll the oligonucleotide-based antiviral approaches against HCMV are still at pre-clinical stage. There are several challenges that must be addressed before they can be considered for use in the clinical setting.HCMV infection is a significant global health problem, especially in immunocompromised individuals and congenitally infected infants. The virus can remain latent in the body, making it challenging to eliminate and increasing the risk of reoccurrence. Currently, there is no licensed vaccine against human cytomegalovirus (HCMV). Despite ongoing research and development efforts, developing a safe and effective vaccine for HCMV has been challenging due to the virus’s complex biology and ability to evade the immune system. This renders the role of antivirals critical in the management of HCMV infection. Unfortunately, the efficacy of licensed antiviral drugs against HCMV is also limited due to side effects, cross-resistance and other factors. It is challenging to develop a safe and effective antiviral against HCMV. In such times, using oligonucleotides to inhibit viral replication could be a desirable strategy. A major advantage of oligonucleotides is that they are designed rationally and specifically bind to target nucleic acid sequences or proteins. Recent advancements in bioinformatics and genome sequence technologies improved the in silico assessment of off-target interaction . Such bi


The number of patients with cancer requiring intensive care unit admission isincreasing around the world. The improvement in the pathophysiologicalunderstanding of this group of patients, as well as the increasingly better andmore targeted treatment options for their underlying disease, has led to asignificant increase in their survival over the past three decades. Within theorganizational concepts, it is necessary to know what adds value in the care ofcritical oncohematological patients. Practices in medicine that do not benefitpatients and possibly cause harm are called low-value practices, whilehigh-value practices are defined as high-quality care at relatively low cost. Inthis article, we discuss ten domains with high-value evidence in the care ofcancer patients: (1) intensive care unit admission policies; (2) intensive careunit organization; (3) etiological investigation of hypoxemia; (4) management ofacute respiratory failure; (5) management of febrile neutropenia; (6) urgentchemotherapy treatment in critically ill patients; (7) patient and familyexperience; (8) palliative care; (9) care of intensive care unit staff; and (10)long-term impact of critical disease on the cancer population. The disclosure ofsuch policies is expected to have the potential to change health care standards.We understand that it is a lengthy process, and initiatives such as this paperare one of the first steps in raising awareness and beginning a discussion abouthigh-value care in various health scenarios. Data suggest that 25% to 30% ofthe beds in ICUs are occupied by cancer patients and that cancer characteristics arenot associated with worse outcomes in the short term.Cancer mortality has decreased in the past three decades;,4 Within organizational concepts, it is necessary to know whatadds value in caring for critical oncohematological patients; thus, the term“high-value care” was coined.The evolving knowledge of critically ill patients with cancer has introduced newconcepts in patient management and ICU admission policies. To identify whatthese practices are, researchers examine “positive deviants” .,8 The application ofthese practices has already been successfully applied in child nutrition andgestational care.,10High-value care practices are defined as high quality of care at relatively lowcost. Cliff et al. demonstrated that the disclosure of suchitems has the potential to change patterns of health activity.It is known, however, that the mere publication of items with scientific evidence oflow-value care reduction alone will not be effective. Change will come from abroader discussion of culture, the benefit of economic savings in cutting low-valuecare, and the involvement of sectors of society so that knowledge of these topicswill not be limited to the medical community.,14 The above statement has arationale, but although a cross-sectional study by Park et al. found that 13low-value services were similarly present in two different models, practices such asunnecessary cancer screening and antibiotics for upper respiratory infection were asprevalent in a fee-for-service model as in a fee-for performance model. This suggests that the model isone of the topics that should be addressed to cut low-value care costs, along withhealth service costs, education, protocol development and better career plans forhealth care workers.Decreasing low-value care has the potential for cost savings; therefore, discussionof the best payment models is of paramount importance. It is logical to think thatthe fee-for-service model, in which payment is based on the procedure executed,stimulates low-value care practices when compared to other models such asfee-for-performance.We know that it is a lengthy process, and in this article, through the collaborationof seven experts on oncologic critical care, we list 10 domains with evidence ofhigh value in the care of patients with cancer .,16 but it is known that delayedadmission and oxygen therapy use are associated with highermortality, whilerapid ICU transfer is associated with better outcomes. Therefore, timely ICUadmission is a measure of high-value care. Lengline et al. demonstrated that inpatients with acute myeloid leukemia and a high risk of tumor lysis, directadmission to the ICU, even if there was no organ dysfunction, was associatedwith better outcomes.Applying objective criteria, such as the modified early warning score (MEWS),can be a strategy to improve ICU admission at the right time.Intensive care unit admission of patients with cancer is frequentlydelayed,,21,22It is of paramount importance to realize that cancer-related characteristics donot predict short-term outcomes. Actually, the severity of organ dysfunction atICU admission and performance status are more sensitive prognostic predictors.,24 Lecuyer et al., in a pioneering study about TLT,demonstrated that the behavior of organ dysfunction up to the fifth day of ICUadmission accurately predicted ICU mortality in nonbedridden patients withcancer. In patients requiring mechanical ventilation, vasopressors or renalreplacement therapy, 3 days of treatment was enough to predict ICUmortality.In cases of uncertainties related to the benefit of ICU “full code care”, atime-limited trial (TLT) is a high-value care practice.The question of the TLT duration may have some peculiarities. Shrime et al.showed that a TLT of up to 4 days for patients with solid tumors and more severeorgan dysfunction could discriminate 30-day survival. However, Shrime et al.concluded that for patients with hematological malignancies or with lower levelsof organ dysfunction, a longer TLT duration, up to 2 weeks, isrecommended.Regardless of the nuances of the evolution of each patient, it is important thatwe have a clear objective for admitting patients with cancer to the ICU. We knowthat some symptom control conditions require resources that the ward cannotprovide; therefore, some patients can be admitted for symptom care.In conclusion, points of ICU admission high-value care are as follows: admissionas soon as organ dysfunction is identified; breaking the paradigm that cancercharacteristics alone are prognostic factors; and TLT use in cases withuncertainties about the short-term evolution. Additionally, Soares et al. showed that thepresence of clinical pharmacists in ICUs was associated with increased odds ofsurvival in critically ill patients with cancer.Intensive care unit practices and team organization have an impact on the outcomeof patients with cancer. Hawari et al. demonstrated that implementing ahigh-intensity ICU staffing model was associated with a decrease in mortality by15% to 60% in critically ill patients with cancer. The results showed thatalthough most oncologists and intensivists were in agreement about the goals oftreatment, there were significant differences in how they approached themanagement of these two hypothetical cases. Intensivists favored the withdrawalof life support measures for patients with breast cancer more than oncologistsdid . The results of thisstudy suggested that oncologists tend to focus on cancer characteristics, whileintensivists focus on multiple organ failure. Regular meetings betweenoncologists and intensivists can reduce potential conflicts regarding theintensive care of patients with cancer. Therefore, this integration is very important and isthe rationale behind why daily rounds are associated with reduced hospitalmortality and a more efficient use of resources in critically ill patients withcancer.Effective communication and therapeutic planning are essential in an oncologicalICU because there may be points of care that are unknown by the ICU team, suchas chemotherapy support, particularities of cancer care and alignment of patientand family expectations. However, oncologists and intensivists have differentknowledge, and conflicts may arise about the proper management of thesepatients. A survey was conducted in Brazil among oncologists and intensivists at2 academic cancer centers on the management of hypothetical patients withdifferent types of cancer (metastatic pancreatic cancer and metastatic breastcancer) who developed septic shock and multiple organ failure. Their study suggested thatpatients treated in ICUs combining expert intensivist coverage, a dedicatedpharmacist and nurses with greater autonomy had the best outcomes.In a retrospective analysis of 129,680 admissions in 93 ICUs, Zampieri et al.characterized the ICUs into three “phenotypes” based on three organizationalcharacteristics: degree of nursing autonomy , presence of a dedicated clinical pharmacist, and presence ofintensivists with certification 24 hours/7 days a week.Notably, the number of available protocols for the prevention of care-relatedinfections was also significantly different in ICUs with better performance,with a higher average in ICUs with better performance.In conclusion, high-intensity ICU staffing models, with the presence ofboard-certified intensivists 24 hours/7 days a week, a dedicated clinicalpharmacist, daily rounds in the ICU with the presence of an oncologist, and ahigher degree of nursing autonomy in domains preestablished by protocols, arepart of what we currently interpret as positive deviants of high-value care. Cancerpatients with hypoxemic respiratory failure require immediate empiricaltreatment, but it is also of paramount importance to reach the cause ofrespiratory failure, since patients without an etiological diagnosis have ahigher mortality.The main cause of ICU admission of patients with cancer is hypoxemic respiratoryfailure.,35 Studies usinga systematic approach and a robust diagnostic arsenal have shown thatapproximately two-thirds of the causes of hypoxemia are infectious, whileone-third are divided between noninfectious or indeterminable causes Direct mnemonic described in the study by Schnell et al. isa relevant aid. Imagingtests are indispensable because they narrow the diagnostic hypotheses but areinsufficient because they usually do not allow etiological diagnosis. To collectsamples of lower airway secretions, it is possible to use an invasive approach,via bronchoscopy, or a noninvasive approach. Although the invasive approachseems more productive, there is at least one randomized study showing that thenoninvasive strategy is equally effective when compared to the use ofbronchoscopy with bronchoalveolar lavage.The systematic approach begins with clinical investigation, and the,39 In patients without thrombocytopenia and outsidepositive pressure ventilation, the safety of transbronchial biopsy is greater,and the diagnostic gain is high for diseases with peribronchial involvement. Asthe diagnostic gain for infectious diseases is greater for bronchoalveolarlavage than for lung biopsy, there is no point in replacing bronchoalveolarlavage with transbronchial biopsy, but they can be consideredcomplementary. Ina study with non-HIV immunocompromised patients, the combined diagnostic gain ofbronchoalveolar lavage with transbronchial biopsy was greater than the gain ofbronchoalveolar lavage alone.Finally, when empirical treatment does not yield a positive result and anoninvasive or minimally invasive approach does not allow etiological diagnosis,an invasive approach, which consists of transbronchial or open lung biopsy,should be considered. Due to the high risk of pneumothorax or bleeding,transbronchial biopsy is usually contraindicated in patients withthrombocytopenia or with positive pressure ventilation. The mortality of an open lungbiopsy is not higher than that of most elective surgeries, occurs inapproximately 2% of cases, and is almost exclusively caused by bleeding andhemothorax. The incidence and occurrence of severe bleeding are associated withcoagulation disorders, especially thrombocytopenia. Compared to bronchoalveolar lavage, open lungbiopsy has a higher gain for noninfectious diagnoses but a lower gain forinfectious diagnoses with a higher probability of complications.Open lung biopsy is an exception for etiological diagnosis. The ideal time toindicate an open biopsy is uncertain, but biopsies performed immediately afterthe onset of hypoxemia are not superior to noninvasive or minimally invasivemethods, while biopsies performed more than 10 days after recognition of thepulmonary infiltrate do not decrease in-hospital mortality.,44 The mostcommon etiology of ARF in these patients is lung infection, which representsapproximately 65% of patients with acute respiratory distress syndrome(ARDS). The riskof ARF is higher in patients with hematological malignancies than in those withsolid tumors, especially in patients with neutropenia and those undergoing bonemarrow transplantation.,47 Patients with cancer andfebrile neutropenia may develop a distinct form of ARDS that occurs during theneutrophil recovery phase in association with the administration of granulocytecolony stimulating factor (G-CSF).,49 Lung canceris the most frequent tumor associated with respiratory complications amongpatients with solid tumors. Other frequent causes of ARF are drug-related pulmonarytoxicity, radiation and clinical situations frequently presented in patientswith cancer, such as chronic obstructive pulmonary disease, cardiogenicpulmonary edema, diffuse alveolar hemorrhage and acute lung injury associatedwith transfusion (TRALI). In patients affected by acute leukemia, ARF may becaused by leukemic pulmonary infiltration, leukostasis and pneumopathy secondaryto tumor lysis.Acute respiratory failure (ARF) is the main cause of unplanned ICU admissionamong patients with cancer.  A multicenter, prospective, observational studyevaluated 1,611 immunosuppressed patients  with ARF admitted to 68 ICUs between 2015 and 2016. Among the1,611 patients analyzed, 596 patients (37%) were intubated at ICU admission orin the emergency room, and of these, 52% died. A total of 915 (56.8%) patientsreceived noninvasive support first, such as standard oxygen therapy, high-flownasal cannula (HFNC), noninvasive ventilation (NIV) or combination NIV plusHFNC. Approximately 40% of these patients required tracheal intubation duringhospitalization. Approximately 85% of patients who failed noninvasive strategiesand required IMV died.The hospital mortality of patients with cancer and ARF is approximately 50%,depending on the etiology, severity, need for invasive mechanical ventilation(IMV) and associated organ dysfunction.,54,55,56 In 2001,Hilbert et al. compared the use of NIV with standard oxygen therapy in arandomized controlled trial that included 54 immunosuppressed patients  with fever, pulmonary infiltrate and hypoxemic ARF. Patientsin the NIV group had a lower intubation rate (46% versus 77%)and hospital mortality compared to the group with standard oxygen therapy (50%versus 81%). Recent studies did not confirm previous reports of earlyNIV benefits compared to oxygen therapy. ,58 Amulticenter, randomized, prospective study conducted in 28 hospitals in Franceand Belgium evaluated the best ventilatory strategy (NIV versusoxygen mask) in 374 immunosuppressed patients with hypoxemic ARF. Neither 28-daymortality nor the intubation rate differed between the groups.The best initial strategy for the ventilatory management of oncological patientswith ARF still raises doubts and uncertainties. Previous studies have suggestedthat early NIV could improve survival and reduce the incidence of intubation andIMV.,62,63,64 Some identified factorsassociated with NIV failure in previous studies are a high respiratory rate, thetime interval between ICU admission and NIV initiation, the need for vasopressoror renal replacement therapy and the ratio of partial pressure of oxygen andfraction of inspired oxygen (PaO2/FiO2) <200.,62,65 Although there are no specific data available onpatients with cancer, NIV can be considered in patients with cardiogenicpulmonary edema or exacerbation of chronic obstructive pulmonary disease withrespiratory acidosis.However, intubation should not be delayed if NIV does not provide immediateimprovement of ARF.Noninvasive ventilation failure is more frequent in patients with cancer than inthe general population and is associated with more complications related tointubation and worse outcomes.versus 52%, p = 0.04) and higher survival at 28days. Tworandomized studies that evaluated HFNC in immunosuppressed patients wererecently published.,69 The HIGH study was amulticenter, prospective study that randomized 778 immunosuppressed patientswith ARF into two groups: HFNC and oxygen therapy. The primary outcome was28-day mortality. Mortality between groups was similar (35.6%versus 36.1%), as was the percentage of patients whorequired IMV (38.1% versus 43.8%). The FLORALI-IM study published in 2022evaluated the use of HFNC versus NIV intercalated with HFNC in299 immunosuppressed patients with ARF. There was no difference in 28-daymortality, intubation or IMV use. There are no studies demonstrating the superiority of HFNCtherapy over NIV or standard oxygen therapy in patients with cancer and ARF.However, hospital mortality in patients with cancer has been decreasing over theyears, probably due to better ARF approaches, including etiologicalinvestigation, early ICU admission and protective ventilation.High-flow nasal cannula appears to be a promising therapy for patients withcancer. In a retrospective cohort of 178 patients with ARF, 76 (43%) receivedNIV plus HFNC, 74 (42%) received NIV plus standard oxygen therapy, 20 (11%)received only HFNC and 8 (4%) received only standard oxygen therapy. Thecombination of NIV and HFNC was associated with lower mortality rates  in cases of suspected intravascular catheter infection, bloodculture with gram-positive bacteria in identification, hemodynamic instability,soft tissue or skin infections or mucositis grade > l.,74In patients without gram-positive bacteria, empirical antibiotic therapy shouldbe discontinued within 48 to 72 hours. In patients colonized or suspected withvancomycin-resistant enterococcus, empirical treatment should be guided by thefollowing criteria: hematological or solid tumors colonized byvancomycin-resistant enterococcus associated with clinical stability;persistence of fever and neutropenia with the use of carbapenem plus vancomycinfor more than 72 hours; or clinical instability. The recommendations of theguidelines of the International Disease Society of America (IDSA) suggest theuse of caspofungin (70mg on the first day followed by 50mg/day),) andamphotericin B liposomal as the second choice (3 - 5mg/kg/day). The third optionin stable patients without prior imidazole use is fluconazole . Ifaspergillosis is a strong clinical hypothesis or the patient is usingechinocandins for prophylaxis, liposomal amphotericin B should be the firstchoice. Micafunginand anidulafungin have not been adequately tested in patients with febrileneutropenia; however, they can be used as an alternative in the absence ofcaspofungin since the spectrum and antifungal activity of the two agents aresimilar.Usually, fungi are not the initial cause of fever in neutropenic patients, andinitial empirical coverage is indicated only in patients with invasive fungaldisease ; after ≥ 4 days ofpersistent fever, if the radiological pattern is compatible with fungal disease;or if febrile neutropenia persists for more than 7 days. The decision on antimicrobial maintenance should be reviewed within 72 and 96hours .Granulocytic stimulating agents can be used in patients without response toantimicrobial treatment in the presence of sepsis or septic shock; outside thissituation, the use should be individualized. There is no benefit of specific protectivemeasures to neutropenic patients, such as masks, gloves or aprons, but allpatients should be subjected to standard precautions, such as hand hygiene(washing hands with soap and water for 2 minutes); appropriate use of personalprotective equipment; respiratory hygiene; careful handling of materials,equipment, clothing and food utensils; environmental hygiene; prevention ofaccidents with sharps and biological materials; safe practice in the preparationand administration of medications; exclusion of plants and flowers in rooms orunits of neutropenic patients; and contraindication of rectal manipulation inneutropenic patients . Three meta-analyses did notrecommend the use of specific diets for neutropenic patients as a measure forthe prevention of infection; however, a well-cooked diet without raw food wassuggested for all neutropenic patients.Regarding prevention, the most effective measure is hand hygiene, especiallyduring the management of neutropenic patients. In patients with solidtumors, chemotherapy in the ICU is often indicated in involvement, such as acuteliver failure secondary to liver metastases, malignant tumor obstructions causedby tumors of the gastrointestinal tract and acute respiratory failure secondaryto bronchial obstruction in lung tumors. However, with the exception of patientswith small cell lung carcinomas and some germ-cell tumors associated withcomplications, the use of emergency chemotherapy in the ICU for solid tumors isassociated with higher mortality.,83,84,85The use of chemotherapy in patients admitted to the ICU is an exception. Fewtumors have a rapid response to chemotherapy to the point of reversal of organdysfunction caused by the cancer itself. The studies that have evaluated the useof chemotherapy in the ICU were small, observational, retrospective, and singlecenter. In general, chemotherapy in the ICU is acceptable in patients withhematological malignancies with complications leading to organ dysfunction, suchas hyperviscosity syndrome, leukostasis and blast crisis.The World Health Organization (WHO) considers that patient experience involvesthe perspectives of patients, families, and communities, considering them asparticipants and beneficiaries. andsimple measures such as time devoted to communication, with proactiveend-of-life conferences and a printed copy of the unit’s guidelines, can helpreduce such symptoms.Interactions between intensivists, other physicians and family members are othermeans of caring for such symptoms, with minimal costs but significant impact.Family meetings are the cornerstone of facilitating open communication, adheringto the care plan and minimizing distress between family members and health careproviders.,89Relatives of patients who experience a critical illness have more symptomsassociated with anxiety, depression and posttraumatic stress than patientsthemselves,versus 1.4 hours) did not reduce delirium; however, therewas a decrease in anxiety and depression among family members. Regardless of the outcome,the preparation of the multidisciplinary team to treat these families isimperative.Although the time of interaction is an important factor, with a rationale forbetter disease understanding, participation in the decision-making process andinteraction with the ICU staff, extending visiting hours remains controversialin the literature. The ICU Visits Group Investigators showed that an extendedvisiting hours policy compared to a more restrictive one  generates stress andburnout in up to half of the professionals.From the organizational perspective, it is vital to guarantee that an ICU is wellstaffed and resourced but also that it is a resilient ICU and therefore capableof adequately responding to daily challenges as well as unexpected situations.,103 Although the short-termmortality of ICU patients with cancer has substantially improved over the lastfew decades,,105 patient-centered outcomesand long-term survival remain suboptimal. Patients with leukemia requiring ICU admission,relapsed or refractory disease, secondary leukemia, or multiorgan failuretherapy were independently associated with 1-year mortality. These factors associatedwith adverse long-term outcomes are similar to those observed in patients withsolid tumors.Post-intensive care syndrome as well as other long-term debilitating conditionsare often described in survivors of severe and prolonged criticalillness. Inpatients with hematologic malignancies, poor quality of life associated withimpaired physical and mental health issues was observed at the 3-and 12-monthfollow-ups.Additionally, the challenges of ICU survivors are not restricted to theirquality of life but also to the impact on the potential continuity of cancertreatment. Loss of functional capacity is often described in ICU survivors,especially those who experience prolonged ICU stays and mechanical ventilation.The fact that up to 80% of cancer patients discharged from the ICU may notreceive optimal chemotherapy should alert clinicians and oncologists and guidethem to engage early in the discussion of the goals of care, considering thatICU discharge will not guarantee cancer survival. (15) In a study of patientswith lung cancer treated in the ICU, 38% of survivors had to change theirinitially planned treatment regimen.Regarding the quality of life of survivors, a study demonstrated that among thebaseline poor quality of life risk factors, previous health-related quality oflife and performance status were associated with better outcomes at the 18-monthfollow-up.Among the priorities to ensure high-value care for ICU cancer patients,intensivists and oncologists should consider the expected quality of life ofsurvivors as well as their ability to remain eligible for highly effectiveanticancer therapies. In addition, patients with cancer should be considered forrehabilitation after critical illness. Although specific evidence related tocancer patients is missing, physical and nutritional rehabilitation has beensuccessfully tested in general ICU survivors.,112,113 This isof utmost importance in ICU survivors and may be even more relevant for patientswith cancer, as the loss of functional capacity after ICU admission may be adetermining factor for eligibility for anticancer therapies.A combination of strategies aiming to improve muscle strength and endurance,respiratory function, nutrition and cognitive status is currently proposed tocounteract the effects of postintensive care syndrome (PICS).There are “positive deviances” in the care of critically ill patients with cancerthat have the potential to generate high-value care from patient admission toend-of-life care considering the experiences of patients and their families.The implementation of such policies is feasible for most centers that care for thispopulation. Even for facilities with scarce resources, applying resources that addvalue is imperative.Initiatives such as this paper are one of the first steps in raising awareness andbeginning a discussion about high-value care in various health scenarios.


Increased GABA Transmission to GnRH Neurons After Intrahippocampal Kainic Acid Injection in Mice Is Sex-Specific and Associated With Estrous Cycle DisruptionNeurobiol Dis. 2022;172:105822. doi:10.1016/j.nbd.2022.105822Ingram RJ, Leverton LK, Daniels VC, Li J, Christian-Hinman CA. A receptor activation is depolarizing in adult GnRH neurons. Therefore, here we tested the hypothesis that increased GnRH neuron firing in IHKA mice is associated with increased GABAergic drive to GnRH neurons. When ionotropic glutamate receptors (iGluRs) were blocked to isolate GABAergic postsynaptic currents (PSCs), no differences in PSC frequency were seen between GnRH neurons from control and IHKA diestrous females. In the absence of iGluR blockade, however, GABA PSC frequency was increased in GnRH neurons from IHKA females with disrupted estrous cycles, but not saline-injected controls nor IHKA females without estrous cycle disruption. GABA PSC amplitude was also increased in IHKA females with disrupted estrous cycles. These findings suggest the presence of an iGluR-dependent increase in feed-forward GABAergic transmission to GnRH neurons specific to IHKA females with comorbid cycle disruption. In males, GABA PSC frequency and amplitude were unchanged but PSC duration was reduced. Together, these findings suggest that increased GABA transmission helps drive elevated firing in IHKA females on diestrus and indicate the presence of a sex-specific hypothalamic mechanism underlying reproductive endocrine dysfunction in IHKA mice.Patients with epilepsy develop reproductive endocrine comorbidities at a rate higher than that of the general population. Clinical studies have identified disrupted luteinizing hormone (LH) release patterns in patients of both sexes, suggesting potential epilepsy-associated changes in hypothalamic gonadotropin-releasing hormone (GnRH) neuron function. In previous work, we found that GnRH neuron firing is increased in diestrous females and males in the intrahippocampal kainic acid (IHKA) mouse model of temporal lobe epilepsy. Notably, GABA Approximately 60% of women with TLE, not using anti-seizure drugs (ASDs), suffer comorbidities such as menstrual disorders and polycystic ovary syndrome. In men, semen abnormalities and low serum testosterone levels are common. Susceptibility to comorbid disturbances is influenced by the location of the epileptic focus, seizure laterality, and the influence of ASDs. Significantly, the relationship between epileptic activity and reproductive endocrine dysfunction is bidirectional; seizure activity can induce dysfunction of the hypothalamic–pituitary–gonadal (HPG) axis, altering serum levels of gonadal hormones, which can further exacerbate seizure activity. It is unsurprising, therefore, that the development and impact of reproductive endocrine disorders in epilepsy is significantly impacted by biological sex.Epilepsy, particularly temporal lobe epilepsy (TLE), is associated with reproductive endocrine disorders in both men and women. Previous work in TLE mouse models found altered activity of GnRH neurons to be associated with reproductive disruption in the form of prolonged estrous cycles. Importantly, estrous stage had a differential effect on GnRH activity in irregularly cycling mice; GnRH neurons were hyperexcitable in diestrus and hypoexcitable, or silent, on estrous. This led the authors to propose that GnRH neuron hyperexcitability in diestrus is a possible driver of comorbid reproductive dysfunction. Here, Ingram and colleagues aimed to investigate the circuit mechanisms underlying GnRH neuron hyperexcitability.The menstrual cycle, or the equivalent rodent estrous cycle, is controlled by the hypothalamic regulation of gonadotropin secretion from the pituitary gland by gonadotropin-releasing hormone (GnRH) neurons. Gonadotropin-releasing hormone neurons are the central output pathway in the neural control of the HPG axis and are likely mediators of epilepsy-associated reproductive dysfunction.2 They studied this in the well-established intrahippocampal kainic acid (IHKA) mouse model of TLE. Unilateral IHKA injection results in chronic epilepsy and hippocampal sclerosis following status epilepticus (SE) with histological features of neuronal loss and reactive gliosis. Status epilepticus was confirmed by video monitoring of behavioral seizures (Racine ≥ 3) in the 5 hours immediately after injection. Chronic epilepsy was not directly established, instead the authors refer to their previous study where 100% of mice were found to develop spontaneous recurrent seizures 1 month after injection. Cycle monitoring was performed before, and then 1 month after, injection. Importantly, as in humans, not all mice developed reproductive comorbidities, allowing for the neural differences associated with prolonged estrous cycle length to be dissociated from the effects of epilepsy. The female mice were thus stratified into 3 experimental groups: “KA-regular” (4/5 day cycle), “KA-long” (≥7 day cycle), and “saline” controls.Ingram and colleagues investigated GABAergic signaling in the hypothalamus as the driver of GnRH neuron hyperexcitability because GABA is the dominant fast synaptic input received by GnRH neurons and paradoxically has an excitatory depolarizing effect in adults similar to the GABA shift seen in immature neurons. The authors observed an increased frequency and amplitude of spontaneous GABAergic inputs onto GnRH neurons in the KA-long group compared to KA-regular and saline groups. The duration of events was also greater for KA-long neurons compared to controls, overall revealing that enhanced GABAergic drive differentiates KA-long and KA-regular mice. Importantly, these differences were not so evident in the presence of blockers of ionotropic glutamatergic transmission. This suggests that hyperexcitability is generated upstream of the GABAergic afferents projecting onto GnRH neurons and that the increased GABAergic drive is potentially due to an increased activity of upstream glutamatergic neurons.Two months after injection, the authors performed electrophysiological recordings of genetically labeled GnRH neurons in acute hypothalamic slices of mice during diestrus. Reproductive endocrine abnormalities in males were not investigated, which may have drawn out some individual differences. However, testosterone levels were not previously found to be altered for males in this model.Next, the authors wanted to see whether this effect was also seen in male KA mice. They found no differences in the amplitude or frequency of GABAergic inputs, only observing a slight decrease in event duration. Their working model is that upstream anteroventral periventricular kisspeptin neurons receive increased glutamatergic inputs which results in a feedforward enhancement of GABAergic transmission onto GnRH neurons. The authors acknowledge, however, that the HPG is regulated by multiple feedback loops and the model they propose may not be the cause of estrous cycle disruption. Future studies could directly test the causal role of increased GABAergic drive by selecting inhibiting the GABAergic neurons and assessing its effect on estrous cycle disruption. Another potential follow-up study could assess the impact of sex hormone feedback on the development of GnRH hyperexcitability. A previous study using the IHKA model found altered levels of E2 and P4 in mice with disrupted cycles, which could influence GABAergic transmission given that these neurons express estrogen receptors, and progesterone metabolites have a powerful antiepileptic effect on the brain.Overall, the authors uncover a sex-specific mechanism for GnRH hyperexcitability in female mice, relating to increased GABAergic drive that is related to estrous cycle disruption. In the IHKA model, neuronal loss and circuit reorganization may extend further than the hippocampus, such that it could be worthwhile to perform histological examinations of connected structures. The hippocampus and hypothalamus are synaptically connected, such that the hyperexcitability of the epileptic hippocampus may be driving increased GABAergic signaling in the hypothalamus. Understanding the mechanisms underlying increased GABAergic signaling in vulnerable mice will be important for the development of targeted interventions, especially because some ASDs can further exacerbate endocrine disruption. Whether novel anti-epileptic treatments, such as gene therapy, targeting hippocampal hyperexcitability can positively impact upon reproductive endocrine comorbidities is an important question, which could shed light on the permanence of such alterations and whether adjunct therapies are needed.The important question remains as to why some epileptic mice develop reproductive comorbidities while others do not. The development of reproductive comorbidity does not appear to correlate with seizure burden or the degree of hippocampal sclerosis, suggesting that changes secondary to the primary hippocampal lesion are at play.Overall, this study is an important demonstration of sex differences in the development of comorbidities in epilepsy. Stratifying males and females, as well as taking in consideration the estrous cycle stage, can reveal specific mechanisms that may otherwise be masked by variability. The management of reproductive comorbidities through a sex-specific lens is important for seizure control and quality of life, given the potent feedback that peripheral sex hormones can exert on the brain. Research that examines, rather than excludes female variability, promises to deliver more effective treatments.Jenna C. Carpenter, PhD and Gabriele Lignani, PhDDepartment of Clinical and Experimental Epilepsy, Queen Square Institute of Neurology, University College London, London


This article summarizes previous and recent research on the fundamental role of arterial smooth muscle cells (SMCs) as drivers of initial and, along with macrophages, later stages of human atherosclerosis.Studies using human tissues and SMC lineage-tracing mice have reinforced earlier observations that SMCs drive initial atherogenesis in humans and contribute a multitude of phenotypes including foam cell formation hitherto attributed primarily to macrophages in atherosclerosis.Arterial smooth muscle cells (SMCs) are the primary cell type in human pre-atherosclerotic intima and are responsible for the retention of lipoproteins that drive the development of atherosclerosis. Despite this, images of atherogenesis still depict the process as initially devoid of SMCs, primarily macrophage driven, and indicate only relatively minor roles such as fibrous cap formation to intimal SMCs. This review summarizes historical and recent observations regarding the importance of SMCs in the formation of a pre-atherosclerotic intima, initial and later foam cell formation, and the phenotypic changes that give rise to multiple different roles for SMCs in human and mouse lesions. Potential SMC-specific therapies in atherosclerosis are presented. Atherosclerosis is the underlying illness resulting in more mortality than any other cause internationally . DespiteRudolf Virchow, in his cellular pathology lectures in 1858, provided an image of what he termed an early sclerotic plate from human aorta, showing spindle-shaped cells containing lipid droplets and “fatty degeneration” in the deep intima  of known CAD-associated variants identified in genome-wide association studies show statistically significant expression quantitative trait locus (eQTL) or splicing QTL effects in vascular SMCs •. A furtLIPA, encoding lysosomal acid lipase (LAL), the sole lysosomal hydrolase capable of hydrolyzing lipoprotein-derived and intracellular cholesteryl esters, when compared to macrophages [One altered intimal SMC phenotype is the SMC foam cell, which, rather than being “macrophage-like,” exhibits distinctly different functional and phenotypic features compared to macrophage foam cells. We originally reported that intimal SMCs exhibit markedly lower expression of the key cholesterol exporter ABCA1 when compared to medial SMCs and proposed they should therefore constitute a large fraction of plaque foam cells . Immunohrophages •. Due torophages •. As sucrophages •, 31•, trophages .A huge volume of work over the last 30 years has delineated the roles of macrophages in the initiation and progression of atherosclerosis. This work has by necessity been done primarily in mouse models, which as indicated require an extreme inflammatory, macrophage-driven state to allow atherogenesis. Macrophages participate in numerous pathways within the plaque including inflammation, apoptosis, necroptosis, phagocytosis, efferocytosis of dying and dead cells, clonal hematopoiesis, cholesterol efflux, and plaque regression , 55, 56.As indicated above, the role of macrophages in human atherosclerosis appears to be more important in intermediate to advanced stage lesions •, where The fact that SMCs are the cell type contributing most to atherosclerotic lesions makes them a clear target to reduce the development and induce the regression of plaque. The migration of SMCs from the media to form the pre-atherosclerotic intima (DIT) occurs in all humans in fetal and early neonatal life, making it a challenging target for therapeutic intervention. Inhibiting the binding of apoB-containing lipoproteins to SMC-generated proteoglycans, for example by administering a vaccine to generate antibodies that block this interaction, represents the exciting possibility of using immunization to block lipoprotein retention in the artery wall lifelong for atherosclerosis prevention , 63. TheAt this point, we are well beyond considering atherosclerosis to be a primarily macrophage-driven process, though it continues to be represented as such in many prominent reviews of this disease. Previous and recent data indicate that atherogenesis in humans is SMC-driven, based on the presence of pre-atherosclerotic diffuse intimal thickening composed almost entirely of SMCs and their secreted proteoglycans that drive initial lipoprotein retention in the artery wall. Evidence indicates SMCs represent the first foam cells formed in the lesion and contribute the majority of cholesterol-laden foam cells through later stages of atherosclerosis. Recent findings using macrophage-specific cell markers, SMC lineage-tracing mice, and GWAS CAD loci indicate a much larger role for SMCs in predicting lesion fate and overall CAD risk than previously known. Further studies will delineate the biologic itinerary of intimal SMCs and SMC foam cells and the suitability of SMCs to become a unique and novel therapeutic target to advance the prevention and treatment of ischemic atherosclerotic vascular disease.


Transcription of the mitochondrial genome is essential for the maintenance of oxidative phosphorylation (OXPHOS) and other functions directly related to this unique genome. Considerable evidence suggests that mitochondrial transcription is dysregulated in cancer and cancer metastasis and contributes significantly to cancer cell metabolism. Recently, inhibitors of the mitochondrial DNA-dependent RNA polymerase (POLRMT) were identified as potentially attractive new anti-cancer compounds. These molecules  inactivate cancer cell metabolism through reduced transcription of mitochondrially-encoded OXPHOS subunits such as ND1-5 (Complex I) and COI-IV (Complex IV). Studies from our lab have discovered small molecule regulators of the mitochondrial matrix caseinolytic protease (ClpP) as probable inhibitors of mitochondrial transcription. These compounds activate ClpP proteolysis and lead to the rapid depletion of POLRMT and other matrix proteins, resulting in inhibition of mitochondrial transcription and growth arrest. Herein we present a comparison of POLRMT inhibition and ClpP activation, both conceptually and experimentally, and evaluate the results of these treatments on mitochondrial transcription, inhibition of OXPHOS, and ultimately cancer cell growth. We discuss the potential for targeting mitochondrial transcription as a cancer cell vulnerability. There is growing interest in the role of mitochondria in cancer due to their involvement in cellular metabolism, apoptotic regulation, redox balance maintenance, and activation of the integrated stress response and related immune responses ,4,5,6,7.Inhibition of mitochondrial transcription has recently been pursued as a novel anti-cancer approach due to the significant effects observed on cancer cell viability ,20. WhilAnother evolving approach to targeting mitochondrial transcription includes the recently discovered small molecule activators of the mitochondrial protease ClpP (herein referred to as ClpP agonists). These compounds include the imipridones ONC201 and ONC206 , the more potent TR compounds, and other recently discovered molecules ,35,36,37Given the emergence of these potential strategies for targeting mtDNA or mitochondrial transcription in cancer, the objective of this paper is to summarize current studies exploring these approaches. We specifically compare and contrast known results of both POLRMT inhibition and depletion and provide initial data comparing these interventions in a common cell model. Lastly, we discuss the promise of inhibiting mitochondrial transcription as a cancer cell vulnerability with an emphasis on identifying promising new therapies for the treatment of various cancers.POLRMT, the only RNA polymerase present in the mitochondria, transcribes and facilitates the replication of mtDNA . POLRMT γ (POLG) and other replication factors are recruited , Twinkle mtDNA helicase (TWNK), etc.) to replicate the mtDNA  were first introduced in clinical trials for Hepatitis C as a strategy to inhibit the HCVRNA-dependent RNA polymerase . An off-Recently, a novel class of mitochondria-targeting drugs were discovered based on their highly unusual property of activating the mitochondrial protease ClpP. For in-depth reviews on pharmacological ClpP activation see Wong and Houry (2019) and Wedam et al. ,62. ONC2Though ClpP activation results in the widespread loss of mitochondrial matrix proteins , identifPOLRMT protein was shown to be strongly reduced in triple-negative breast cancer (TNBC) cells following ClpP activation by TR-57 and TR-107 ,38. ImmuClpP agonists and POLRMT inhibitors have been shown to impair cellular proliferation in multiple cancer cell lines and models ,71,77,78One of the unusual properties of ClpP agonists is that they appear to adversely affect cancer cells but have little or no effect on normal cells. Several groups have tested ClpP agonists on immortalized cell lines  or primary cells and observed no significant effects on cell proliferation, viability, or apoptosis as compared to the cancer cell models ,68,75,84Both ClpP agonists and POLRMT inhibitors induce major metabolic alterations in cancer cells, most prominently observed with the inhibition of OXPHOS ,75,78,83Another common metabolic alteration observed following ClpP activation or POLRMT inhibition is AMPK activation ,75. GreeAs an expected consequence of OXPHOS inhibition, ClpP agonists have been shown to cause a compensatory increase in glycolysis ,75,78. AThese observations also suggest that cancer cells that are more reliant on OXPHOS for proliferation will respond better to ClpP agonists or POLRMT inhibitors. This includes cancer types and cancer populations known to be more OXPHOS-dependent, including cancer stem cells , chemothAn important similarity between ClpP agonists and POLRMT inhibitors is the corresponding reduction in mtDNA content, despite observations that the mtDNA copy number appears to change at different rates. IMT1 treatment reportedly took ~96 h to reduce the mtDNA copy number to ~25% of its original number , whereasImportantly, another potential mechanism leading to a reduction in mtDNA is through loss of POLRMT protein or activity. POLRMT is critical to mtDNA replication as a primase , so with0 cells) are resistant to ONC201 [0 cells have not been reported. Comparing p0 cells to ClpP agonist- or POLRMT inhibitor-treated cells may help establish if the loss of mtDNA is a critical and common anti-cancer response to these treatments.While it is logical that mtDNA depletion impairs OXPHOS due to loss of required mtDNA templates to produce OXPHOS subunits, it remains to be established whether depletion of mtDNA is required for the growth inhibition observed by these compounds . To be ro ONC201 , the effIt has been shown that POLRMT and TFAM protein expression decreases after treatment with ClpP agonists, but it has not been established that mitochondrial transcription is inhibited. We predicted that mitochondrial transcription would be impaired by ClpP agonists due to their ability to deplete POLRMT, TFAM, and TEFM proteins which are all essential for mitochondrial transcription . Both TRFrom the combined analysis of these studies, there are multiple similarities in the mechanism of action of ClpP agonists and POLRMT inhibitors. This includes depletion of mtDNA, inhibition of mitochondrial transcription, impairment of OXPHOS, activation of AMPK, metabolic remodeling, and ultimately, inhibition of cancer cell proliferation. From these observations, we propose that the similarities in mechanism between ClpP agonists and POLRMT inhibitors in part stems from the inhibition of mitochondrial transcription observed with both drug classes. While mitochondrial transcription may be impaired by different mechanisms, exploring the similarities and differences between ClpP agonists and POLRMT inhibitors may ultimately help understand the mechanism of action of both drug classes and evaluate the feasibility of targeting this mitochondrial process in cancer cells.50 of TR-57 in HEK293T cells is ~15 nM after 72 h, whereas the POLRMT inhibitor IMT1 has an IC50 of ~190 nM after 120 h of treatment , while all the ClpP agonists tested were highly effective at much lower concentrations  . ImportaThough ClpP agonists and POLRMT inhibitors effectively reduce cancer cell proliferation, much remains to be discovered about the mechanism of action of these compounds. An outstanding question is why ClpP agonists and POLRMT inhibitors are effective inhibitors of cancer cell growth, but not normal cell growth. As mentioned above, clinical trial data report ClpP agonists to be well tolerated with minimal side effects in cancer patients ,65, furtLimitations to the current studies include a lack of studies directly comparing ClpP agonists and POLRMT inhibitors. While this review aimed to emphasize the surprising similarities between these anti-cancer approaches, how ClpP agonists and POLRMT inhibitors compare to other mitochondria-targeting drugs also remains to be determined. This is important because there are many cytotoxic effects to other mitochondria targeting drugs , so it iIn summary, targeting mitochondrial transcription is a novel and potentially promising approach to disrupt cancer cell metabolism. Both ClpP agonists and POLRMT inhibitors have intriguing anti-cancer properties that will continue to be investigated as these compounds advance in clinical trials. In addition to emphasizing the importance of mitochondrial transcription in mitochondrial function, elucidating the mechanism of action of ClpP agonists and POLRMT inhibitors could lead to broader discoveries about cancer and how to better treat this prolific disease.


Among many types of wearable sensors, MOFs-based wearable sensors have recently been explored in both commercialization and research. There has been much effort in various aspects of the development of MOF-based wearable sensors including but not limited to miniaturization, size control, safety, improvements in conformal and flexible features, improvements in the analytical performance and long-term storage of these devices. Recent progress in the design and deployment of MOFs-based wearable sensors are covered in this paper, as are the remaining obstacles and prospects. This work also highlights the enormous potential for synergistic effects of MOFs used in combination with other nanomaterials for healthcare applications and raise attention toward the economic aspect and market diffusion of MOFs-based wearable sensors.  •Review article on metal organic framework (MOFs)-bsed wearable sensors.•The advantages of using MOFs as sensing platform and their healthcare applications are discussed.•The use of MOFs combinations with other materials in the production of electrochemical sensors is also discussed.•MOF-basedsensors manufacturing, achievements, market diffusion and related limitations are described in depth.•Challenges and prospects using MOFs-based wearable sensors have been proposed. The market demand for these technologies has rapidly expanded from $20 billion in 2016 to $80 billion in 2022 .In addition to sensing of various physiological variations, wearable sensors should be able to detect biomarkers specific to diseases at very early stages of their emergence, then it can help accurate diagnosis of diseases like DNA damage , Alzheim2 g−1) . SE. SE2, endiabetes a and b wdiabetes c shows tdiabetes d. From tdiabetes e. The va4.3in-situ detection of radial artery pulse signals  growing Cu3(HHTP)2 on the surface of ZnO thin film, a novel composite based on ZnO and MOFs (Cu3(HHTP)2) has been generated to prepare type-II heterojunction UV photodetector . The MOF4.4Additionally, in the past 10 years, efforts have been made to increase the sensitivity of wearable pressure sensors so they can be used in a wider range of medical applications. In order to achieve fast and accurate health monitoring, wearable pressure sensors must be produced with high sensitivity, wide sensing range, excellent reproducibility and/or reusability, and with few restrictions and drawbacks related to small mechanical deformations such as physical motions of fingers. Recently, various skin wearable sensors have b een developed, however, often limited by air and moisture, which restrict their long-term applicability and make it too difficult to obtain tight adhesiveness to the skin ,61.In this regard, a sandwiched C-MOF/PANIF@PU-based wearable pressure sensor has been fabricated . C-MOF aUnfortunately, there aren't many research reporting MOF-based wearable sweat sensors in the literature so far. However, a number of research that were published in this field and offered some answers that ought to be taken into account in the near future to enhance MOFs' potential future utility in creating more precise wearable sensors. Sweat is a readily available, non-invasive fluid that can be drawn from a person's body during any physical activity, as well as on demand using thermal heating or a chemical induction process known as iontophoresis ,64.Sweat analysis is more advantageous than blood diagnosis. However, this method has a number of issues and restrictions that make it difficult to create wearable sweat sensors based on MOF. Four significant issues need to be resolved: (i) the inability to control sweat production rates; (ii) the impact of health circumstances, such as pregnancy, on sweat production; (iii) the limitations of iontophoresis; and (iv) the stretchable wearable heater that makes diagnosis uncomfortable and annoying. By regulating sweat generation and enhancing its storage and transportation, efforts should be made to develop MOF-based wearable sweat sensors .For instance, because patients frequently detest such non-invasive approaches, producing perspiration through physical activity is not a desirable technique. Since excessive sweating makes it uncomfortable to take showers six times a day for a diabetic person. Accurate glucose detection requires diabetic patients to produce sweat during six times each day from breakfast to dinner ,66. EvenAdditionally, once perspiration starts with any physical activity, glucose concentrations decrease over time in sweat because it is quickly ejected from the sweat gland. The diluting effect brought on by an increase in sweat rate during an extensive exercise, is thought to be responsible for the lower levels of sweat glucose ,68. In aDespite similar patterns, different body areas sweat at different rates, which, as a result of the dilution effect, because varying glucose concentrations at any given time.In conclusion, it is highly unlikely that sweat glucose monitoring that is accurate, real-time, continuous, and achieved through exercise sweat will be the best course of action, especially for diabetic persons who suffer from other serious diseases such as blood pressure and heart related disease a and b. Iontophoresis is a well-known technique for health monitoring and therapeutic medicines, particularly for sedentary individuals . It work2 may be reached by carefully constructing the iontophoresis electrodes and improve sweat stimulation of the medication in the synthesized films. It was possible to extract enough perspiration for an accurate in-situ glucose analysis without endangering the individuals' epidermis or making them uncomfortable [To collect local sweat effectively, the system should have embedded iontophoresis capabilities. The sensing area is sealed to collect sweat using an iontophoresis technique with the sweat-rate sensor mounted on the positive electrode . Additiofortable .Iontophoresis is essential in addressing the issues of an aging population and diabetic people with health issues and a lack of access to healthcare thanks to these benefits. While the chemical makeup of workout sweat is significant for uses in physiology and athletics, chemical iontophoresis-generated sweat is more beneficial for medical diagnostics ,74. Due Iontophoresis, however, might be more appropriate for a single application as opposed to ongoing monitoring, as the repetitive administration of iontophoretic current at the same location could be damaging to the underlying skin. To protect the skin from operational damage and irritation, the wearable sensors should be prepared taking into account the current density and iontophoresis time. As a result of repetitive sweat induction of iontophoresis, the pH of the epidermis will vary due to ionic buildup at the sweat-sampling site. In order to minimize burns brought on by unfavorable pH effects, safeguard the epidermis, and solve this problem, buffered gel coatings may be utilized ,76. The However, if the stretchable wearable heater is made of a soft, thin, solid-layer patch, it will be difficult to wear and will not allow for adequate airflow. The solution to this issue is the use of super hydrophobic materials, which can introduce the sweat into the sensing system and keep the skin dry during the sweating process. This might be a useful method in the field of for non-invasive biosensing of glucose, however it requires power. The use of this method for in-situ sweat glucose monitoring has not been documented in any studies .The market for electrochemical biosensors is being driven by diagnostic and health monitoring applications, notably the growing need for wearable sensors ,80. The MOF-based wearable sensors frequently include a biological component in the sensing matrix, such as enzymes and antibodies, which require special storage conditions. Although specific characteristics of commercially marketed glucometers' excellent storage stability are covered by patent, the capillary chamber where the enzyme is maintained along with mediators and stabilizers appears to play a significant role in it . DespiteIt should be noted that approaches based on enzymatic inhibition can produce false positive results since various substances impede enzyme activity, reducing sensor selectivity and lowering its sensitivity . The resNon-enzymatic biosensors based on biomimetic materials are the subject of significant research because they have the potential to overcome recurring stability difficulties, paving the path for mass production and commercialization of viable sensors . In addiOne of the major technological challenges is the gap between the innovative concepts brought by academic research and their integration with market needs, as well as the inaccessibility of biological samples, known as real-world samples, and limited experience on marketable devices. Collaboration between academies and companies could yield a viable solution to the need for more durable, stable, and low-cost sensors.5The prospects presented by MOFs-based wearable sensors are limited in certain ways by several obstacles that have yet to be fully resolved. Ionic strength, temperature, and liquid-junction potentials can all be adjusted for or taken into consideration . LeachinThe use of ordinary addition to overcome the matrix effect is incompatible with the wearable concept and will almost certainly increase the cost of the sensing platform for environmental analysis . For weaIt appears that there aren't any convincing designs for inexpensive, mass-producible ion-selective microelectrodes that can be used for wearable electronics and environmental monitoring. Out-sensor computing consumes a lot of energy and adds a lot of latency. It is critical to create physical intelligent devices that allow near-sensor or in-sensor computing in order to solve this problem. Such a problem might be resolved by machine learning, which might also enhance the utilization of artificial e-skins with multiple wearable sensing nodes for gathering external information. On the other hand, a significant issue for everyday monitoring is the service life of wearable electromechanical sensors. Based on their self-power supply, chemical stability, wearing comfort, and mechanical durability, wearable electromechanical sensors will have an effect on long-term use. Self-repairing hydrogels and elastomers may open new avenues to overcome wearable sensors stability in the future. A unique wearable sensor needs to be built through material design and structural optimization in order to enable long-term monitoring and high-fidelity electrophysiological recording.In conclusion, there is still a lot of work to be done in order to create wearable sensors based on MOFs that are affordable and dependable for research on untreated real samples.66.11Stability of MOFs-based wearable sensors is a major issue since they are relied on a biorecognition system that requires a stable environment for the biorecognition element such as enzymes especially for long-term storage.2The sensitivity, selectivity and LOD of wearable sensors are the key features of any wearable sensor and ideal biosensors must have a very low value of LOD and outstanding selectivity toward the studied analyte so that they can provide accurate data with negligible hysteresis at lowest interference.3Up-scaling wearable technology always confront reproducibility related challenges. To ensure accurate fabrication and marketing of MOFs-based wearable sensors they must have excellent reproducibility and repeatability, so similar sensors with similar features could be produced.4To be consider for human health diagnosis, MOF-based wearable sensor must be able to operate properly in a real sample such as saliva, blood, urine, sweat, body fluid, tears, etc. The real sample collection is itself a challenge; some factors need to be considered for collecting a real sample for detection.5Limitations due to MOFs morphology, safety and biodegradability for constructing highly performed wearable sensors are one of the most critical issues in developing MOFs-based wearable sensors for health diagnosis. MOFs-based sensors must be safe and have mechanical features so they can be compatible with the studied area (e.g. skin) without interfering the results causing no damage to the patient.6MOFs-based wearable sensors must be removed from the body once their mission is accomplished to avoid any side effects which require additional sophisticated procedures. Some health complications may appear if one of the component of the wearable sensor failed to be removed and/or in case of an in-vivo damage which may lead to serious in-vivo injuries during the diagnosis of a disease. Wearable sensors must be resistant to any mechanical and physicochemical changes to avoid any in-vivo issues.7In case of commercialization and scaling-up, production costs and customer behavior toward MOFs-based wearable sensor could be considered as a serious challenge. To get a rapid and efficient market diffusion, the customer needs to be convinced about the advantages of the product taking into account its cost, ease of operation with minimal efforts and product safety.8Several sensors require calibration before every use and incubation in conditioning solution. Such conditions are incompatible with wearable technology. Furthermore, many forms of sensors have the issue of drift that causes significant error in calculating analyte concentration. Therefore, these sensors need to be re-calibrated at regular intervals. Researchers are addressing this issue by developing calibration-free sensorsMOFs-based Wearable sensors are characterized by various challenges including usual sensing challenges such as pretreatment, stability and reproducibility, sensitivity challenges and real sample sensitivity along with MOFs wearability related challenges such as materials related challenges, Self-destroyed sensor-based challenges and consumers’ behavior toward such devices. These are also the major challenges in the fabrication of electrochemical biosensors.6.21The issue related to conformal contact between wearable electrochemical sensors and the human body can be resolved by matching the mechanical properties of the device with that of the human tissues. The human tissue is characterized as soft, stretchable and curvilinear with unique mechanical properties. In this regard, researcher must conduct more efforts in developing MOFs with outstanding mechanical properties including but not limited to curvilinearity, softness, flexibility and outstanding stretchability.2A combination of MOFs and silicon based materials could help in improving MOFs-based wearable sensor toward extreme mechanical and physichochemical changes such as pressure and temperature. Such innovative strategies must be explored by researchers to realize highly biocompatible and biodegradable chemical sensors that are eventually consumed by the body.3MOFs functionalization and their combinations with other materials such as graphene, reduced graphene oxide (rGO), carbon nanotubes (CNTs), metals and metal oxide nanoparticles may improve the stability of the biorecognition matrix and then, ensure long-term storage.4Developing cost-effective and eco-friendly synthesis procedures to prepare MOFs is highly recommended to ensure safe, biodegradable and cheap wearable sensors.5Fast response and Micro-nano MOFs-based wearable sensors can be disposed and removed easily and may improve their performance would involve fewer efforts.6The gap between research laboratory and hospitals should be bridged so more trails will conducted and MOFs-based wearable sensors can be tested in real-life circumstances, which will help in evaluating their performance and convincing people to use such tools to diagnosis their health.7To ensure up-scaling and excellent market diffusion of such sensors, more efforts should be conducted focusing on the key element for a successful up-scaling strategy including but not limited to production costs, stability and advertising.8MOFs-based wearable sensors should benefit from machine learning and artificial intelligence to overcome the aforementioned challenges. Such technologies may offer excellent opportunities to improve sensors analytical performance and improving their stability, controlling and miniaturizing their size.7•Designing eco-friendly and more conductive MOF-based composites to create more stable and sensitive MOF-based sensing platforms that ensure the accuracy and linearity of the produced MOF-based wearable sensors.•To expand their applications in wearable sensing, MOFs-based sensing platform should be stretchable and flexible which require more efforts on improving their mechanical properties and their biocompatibility taking into account their costs of production.•To increase wearing comfort and skin affinity, MOF wearable sensors should be created using bionic structures rather than artificial glue.•Integration of 3D technologies in the fabrication of MF-based wearable sensors since such technologies helps in improving the long term stability and the reproducibility of the sensors, and may also help in the up-calling of these sensors.•The operational conditions of wearable sensors should be optimized to produce more adaptable MOF-based wearable sensors for each application and to meet a variety of end-user biological needs.•To produce more accurate and smart MOF-based wearable sensors, machine learning and artificial intelligence should be integrated during the process of production.MOF based wearable sensors showed promising performance for future applications in human health care applications and disease diagnosis. However, these sensors must be sensitive, selective and stable to maintain long term performance. The design and architecture of the MOF-based sensing platform while being doped with other materials such as carbon based materials and metals nanoparticules which have high flexibility, stability, conductivity and trustworthy durability can be useful in the fabrication of high performance MOF-based wearable sensors. The development of high performance MOF-based sensors depends ongoing research and technological advances. Therefore, future researchers will likely focus on the following areas, as stated below.All data generated or analyzed during this study are included in this published article.The publication of this article was funded by the Qatar National Library (QNL). The authors would like to acknowledge the library for supporting the publication of this article.Hicham Meskher: Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Project administration, Resources, Writing – original draft, Writing – review & editing. Samir Brahim Belhaouari: Funding acquisition, Project administration, Resources, Supervision, Validation, Writing – review & editing. Fariborz Sharifianjazi: Methodology, Project administration, Resources, Supervision, Visualization, Writing – review & editing.The authors declare that they have no known competing financialinterestsor personal relationships that could have appeared to influence the work reported in this paper.


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