The molecular understanding of carcinogenesis and tumor progression rests in intra and inter-tumoral heterogeneity. Solid tumors confined with vast diversity of genetic abnormalities, epigenetic modifications, and environmental cues that differ at each stage from tumor initiation, progression, and metastasis. Complexity within tumors studied by conventional molecular techniques fails to identify different subclasses in stromal and immune cells in individuals and that affects immunotherapies. Here we focus on diversity of stromal cell population and immune inhabitants, whose subtypes create the complexity of tumor microenvironment (TME), leading primary tumors towards advanced-stage cancers. Recent advances in single-cell sequencing (epitope profiling) approach circumscribes phenotypic markers, molecular pathways, and evolutionary trajectories of an individual cell. We discussed the current knowledge of stromal and immune cell subclasses at different stages of cancer development with the regulatory role of non-coding RNAs. Finally, we reported the current therapeutic options in immunotherapies, advances in therapies targeting heterogeneity, and possible outcomes. The ansThere are several mechanisms of cellular plasticity. Differentiation is the most common route by which a pluripotent stem cell evolves into a mature cell type . ContrarNon-coding RNAs function as significant players in post-transcriptional gene regulation within diverse cell types. Cancers are primarily regulated by a different set of non-coding RNAs, classified according to their sizes . MicroRNTME is remarkably a complex ecosystem with a heterogeneous population of cancer cells and the associated stroma, which drives tumor initiation and growth. Stromal cells, extracellular matrix, paracrine signaling molecules , and immune landscape formulates TME . InteracCAFs are the major stromal component of many solid tumors and are well known phenotypic remodelers of the stromal environment . They covia TGF-β pathway  which are known to drive terminal differentiation and exhaustion  activity that blocks the transcription factors  T-cell therapy, an oncolytic virus therapies . Advancevia extensive studies of gene expression and metabolomic analysis. Targeting only the αSMA+ tumor population would not be helpful as it may result in the loss of immuno-supportive residents and may hamper significant homeostatic functions  mainly expressed by macrophages in aggressive human breast cancer subtypes . High CSMZB1, JCHAIN, and IGLL5) in ICBs responders relative to non-responders indicating the driving force of B cells in immunotherapy (T cell therapeutics is advancing since the discovery of a breakthrough of chimeric antigen receptor (CAR) -modified T cells and immune modulation using antibodies that block immune regulatory checkpoints in 2013. Both CD8+ and CD4+ cells have multifaceted roles in antitumor immunity. CD4+ cells secrete interleukin (IL)-2 activating CD8+ T cells, cytokine secretion , activation of CD40 ligands that activates dendritic cells and leads to differentiation and maturation of B cells . Immune otherapy .NK cells can induce direct killing of tumor cells, and recruit or activate of T cells and DCs. Three forms of NK therapy have emerged: Bi-specific antibodies, induction of antibody-dependent cell-mediated cytotoxicity (ADCC), and gene therapy with NK cells. Current possibilities to use NK cells are (a) targeting inhibitory NK cell surface receptors or (b) targeting activating NK cell receptors . NKG2A iResearch in the field of the tumor microenvironment is an impressive and significant stage. Despite progress in cancer immunotherapies, the incidence of tumor relapse and recurrence had necessitated a profound study of tumor heterogeneity. Clinically we are struggling with therapies that would target a wide range of cells, being unaware of subtypes that escape and show resistance. Many drugs target tumor biome by using monoclonal antibodies, angiogenic, and protein kinase inhibitors, but the precise targets are rare. With advances in single-cell technology, intra-tumoral heterogeneity is not much explored. Characterization of stromal and immune cell subtypes inhabiting at different tumor stages, early or late-stage, and the corresponding normal tissues revealed a broad range of cell subsets, differentiation patterns, and immunological regulations within TME. Targeting these well-explored immuno-modulatory cells directs the development of next-generation therapeutics. These next-generation therapies are at an early stage of development and would involve trials on small molecule inhibitors, non-coding RNAs, and cell-specific receptors that may lead to better cancer cure.There are still several questions in the TME that needs careful investigation. One aspect is the lineage tracing. A challenge in tumor biology is associating molecular differences among progenitor cells and their capacity to generate differentiated cell types. Studies are needed to associate changes in progenitor and mature cell types and answer questions such as a) Does a differentiated cell have the capability to transdifferentiate in other differentiated cell types in TME? If yes, what are these cells in the tumor microenvironment that shows cellular plasticity? b), and how the cellular plasticity is regulated? To answer these questions, we need to understand the origin of stromal or immune cell types in TME.The second aspect is understanding the heterogeneity of stromal and immune cells that make up most of the TME. Fibroblasts and macrophages are among the most diversified populations whose characteristics change with their location in tumors- perivascular, hypoxic, necrotic areas, and regions of interaction with blood vessels and lymph nodes. Rare information is available on the interaction of these cells with other cell types and how different tumor zones influence the development of subtypes. Technologies such as spatial transcriptomics  and multiplex immunofluorescence would be pivotal to identify prevailing subtypes.The third aspect requiring attention is tumor secretome. Investigations on exosomal secretions, non-coding RNA transcriptomics, and epigenetic remodeling would boost our understanding of tumor biome. Single-cell genomic technologies are an efficient approach to understand every subtype residing within TME. It can also enhance our ability to discover novel target molecules, specific pathways targeted by these drugs, that would facilitate effective strategies. Identification of precise sub-typical population by scRNAseq analysis can serve as biomarkers to develop accessory treatments. Assessment at a single cell level can also provide us cell variations before and after immunotherapies with which medications can be optimized. In summary, we anticipate that single-cell analysis would be a great approach to understand tumor biology and designing therapies that would revolutionize the tumor treatments as it may reduce the gap between responders and non-responders.LA and DP contributed to the study conception and design. LA collected the relevant paper and drafted the manuscript. DP revised the manuscript and supervised the study. LA and DP prepared the review figures and tables of the manuscript. All authors contributed to the article and approved the submitted version.This work was supported by SERB women excellence award (SB/WEA-02/2017) to DP Also, DP gratefully acknowledges financial support of IIT Ropar to the TERM laboratory.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


In vitro studies confirm some neuronal interactions between the HPA and HPO; and immuno-histochemistry clarifies the location and sequence of inter-neurone activity within the brain. In both species, exogenous corticoids, ACTH and/or CRH act at the pituitary (reduce LH release by GnRH), and hypothalamus (lower GnRH pulse frequency and delay surge release). This requires inter-neurones as GnRH cells do not have receptors for HPA compounds. There are two  pathways for CRH suppression of GnRH release via CRH-Receptors: one being the regulation of kisspeptin/dynorphin and other cell types in the hypothalamus, and the other being the direct contact between CRH and GnRH cell terminals in the median eminence. When we domesticate animals, we must provide the best possible environment otherwise animals trade-off with lower production, less intense oestrus behaviour, and impaired fertility. Avoiding life-time peri-parturient problems by managing persistent lactations in cows may be a worthy trade-off on both welfare and economic terms – better than the camouflage use of drugs/hormones/feed additives/intricate technologies? In the long term, getting animals and environment in a more harmonious balance is the ultimate strategy.The environment contributes to production diseases that in turn badly affect cow performance, fertility and culling. Oestrus intensity is lower in lame cows, and in all cows 26% potential oestrus events are not expressed (to avoid getting pregnant). To understand these trade-offs, we need to know how animals react to their environment and how the environment influences hypothalamus-pituitary-adrenal axis (HPA) interactions with the hypothalamus-pituitary-ovarian axis (HPO). Neurotransmitters control secretion of GnRH into hypophyseal portal blood. GnRH/LH pulse amplitude and frequency drive oestradiol production, culminating in oestrus behaviour and a precisely-timed GnRH/LH surge, all of which are disrupted by poor environments. Responses to peripheral neuronal agents give clues about mechanisms, but do these drugs alter perception of stimuli, or suppress consequent responses?  First: over a 2–3 week period, more than 80% non-domesticated wildebeest roaming the plains of the Serengeti in east Africa mate and calve with no interference by man . The tratreated cows take longer to get pregnant, 64, 100 and 58 days, respectively,  reduce fertility, and this is even more dramatic for high yielding cows that also generate more heat than they can dissipate  improve oestrus detection? When progesterone profiles identify possible oestrus events , motion sensors and observations by farm staff detect only 74% potential oestrus events . So, whyTo understand these trade-offs, it is necessary to know how animals react to their domesticated environment  and how cows/ewes translate their responses in terms of controlling fertility. In large part, this involves hormonal control of the hypothalamus-pituitary-adrenal axis (HPA) and its interaction with the hypothalamus-pituitary-ovarian axis (HPO). To examine these relationships, all our studies  are carried out under UK Home Office licenses for work on living animals and with the approval of the University of Liverpool Ethical Review process.In many of the above ‘trade-off’ situations, either luteinising hormone (LH) pulse or surge patterns (or both) in dairy cows are disrupted leading to reduced fertility feeding: . GreaterBriefly, in cows and ewes neurotransmitters in the brain  control secretion of gonadotrophin releasing hormone (GnRH) into hypophyseal portal blood, and thus LH release from the pituitary into the peripheral circulation. During the luteal phase, feedback from progesterone and oestradiol restrain small discrete GnRH/LH pulses to approximately one per four hours. As peripheral progesterone concentrations decline, LH pulse amplitude and frequency increase to approximately one per hour further driving ovarian follicular growth and oestradiol production. Towards the end of the follicular phase, when progesterone concentrations are low and oestradiol concentrations are at a maximum , there iIf the environment is not ideal, in cows and ewes GnRH/LH pulsatility is disrupted, as well as the timing and amplitude of the GnRH/LH surge, resulting in failure to initiate a pregnancy. This key disruption is usually temporary so that when prevailing conditions improve, normal hormonal profiles will resume.A major difficulty encountered by high-yielding dairy cows is achieving sufficient food dry matter intake (DMI) around calving. Feeding in the dry period has very important consequences for events in the following postpartum period. For example, in the week before calving, feeding duration and DMI are both more than 20% lower in cows that subsequently have mild-or-severe metritis and these same cows produce 6-8 kg milk/day less than healthy herd-mates during the first three weeks after calving . Low DMImonthly milk yield and maximum values are not different from normal in those cows that develop ovarian cysts, but weekly analysis reveals shorter durations of peak yield  and low urea:BHB ratios are more useful after calving as predictors of an increased likelihood of cystic ovaries and delayed commencement of luteal activity.A further example regarding the consequences of the genetic drive for higher yields: patterns of ak yield . It is nak yield . High miak yield . Howeverpulsatility . Evidence for this suggestion follows:Prolonged anoestrus after calving is characterised by low oestradiol and progesterone values due to suppression of GnRH/LH  review: . WhereasDairy cows with naturally-occurring follicular cysts have lower plasma progesterone concentrations than those with luteal cysts ; but cowAs it is impossible to closely monitor cows at the precise moment ovarian cysts are spontaneously formed, experimental models have been developed. Starting on Day 15 of a ‘normal’ oestrus cycle, in heifers receiving either:high doses of oestradiol and progesterone to mimic high values at the end of pregnancy; 3 out of 8 formed persistent follicles ;repeated doses of ACTH to stimulate adrenal secretion; 6 out of 13 formed persistent follicles ;or prolonged low doses of progesterone to replicate low progesterone values seen after adrenal stimulation; all 13 formed persistent follicles .Combined observations from these models reveal that during treatmentfollicle stimulating hormone (FSH) concentrations are normal;LH pulse and surge concentrations are lower;some prolonged dominant follicles ovulate after 10 days with higher oestradiol concentrations and greater internal diameters than those follicles that persist for > 20 days;occasionally persistent follicles luteinise;for the second half of the life-span of persistent follicles, oestradiol values are basal, thus the structure can remain for up to 50 days after endocrine function declines, and other smaller follicles appear ;Figure 3subsequent treatment with very low dose progesterone or GnRH decreases plasma oestradiol and a new follicular wave emerges.In short, heifers with experimentally-produced persistent follicles resemble cows with clinically-diagnosed spontaneous cysts. But questions remain – why/how are pulsatile and surge LH secretion disrupted?Ewe models have also been used to examine in greater detail the functionality, and consequences, of persistent follicles. In ewes, endogenous LH is suppressed by a GnRH antagonist but when replaced with frequent injections of low dose LH for 60 h, normal follicular growth occurs with ovulation after a large LH injection at 60 h, followed by a luteal pattern of plasma progesterone . HoweverRegarding the consequences of prolonged oestradiol exposure, ovariectomised ewes treated with oestradiol implants for 2 to 12 days do not have a normal LH surge after a final challenge with oestradiol . HoweverClearly, in cows and ewes continuation of LH pulses are required for persistent (cystic) follicular growth and oestradiol production, but why does an LH surge not occur at the end of what appears to be an otherwise normal follicular phase? Cystic follicles in cows are associated with clinical production diseases, including uterine infection . A betteE coli (LPS). However, there are several important points to note:-The hypothalamus-pituitary-adrenal axis (HPA) protects life by monitoring the environment and activating immediate responses to threatening stimuli. Responses are similar in all mammals; during transport, both corticotrophin releasing hormone (CRH) and vasopressin (AVP) are released from the hypothalamus, which in turn cause the anterior pituitary to secrete adreno-corticotrophin hormone (ACTH); that then triggers the adrenal glands to secrete corticoids  of the activated HPA axis are responsible for disrupting normal function at specific levels of the HPO. While some studies investigate endocrine systems in cattle, many more use ewes that are more easily handled, less expensive to maintain, but closely mimic responses in cows. Different components of the HPO (as well as agonists or antagonists) have been administered to assess changes in responses during exposure to a variety of acute or chronic stimuli; and different HPA hormones have been assessed for their impact on HPO function. Moreover, a variety of reproductive states have been involved; for example, intact ewes in the breeding or non-breeding season, in luteal or follicular phases with or without (synthetic) hormonal manipulation, and short- or long-term ovariectomised animals with or without (synthetic) hormone replacement. There are difficulties in interpreting the responses in ovariectomised animals: the HPO itself adapts after ovariectomy, irrespective of any changes in HPA activity; selecting the correct replacement hormone in appropriate doses/patterns/duration affects responses; and the role of the animal’s own balancing systems are compromised (or often ignored during interpretation).amplitude in ovariectomised ewes  is lower . This ac review: . During  review: . Furtheral blood , 2000.In vitro perifusions of ewe pituitary slices show unequivocally that ACTH reduces the amount of LH released by repeated low dose GnRH; hence, a direct effect occurs at the pituitary and is enhanced with additional oestradiol  and decreases both GnRH biosynthesis in the hypothalamus, and the number of GnRH-Receptors in the pituitary  just before an expected LH surge, the surge is delayed or totally blocked . The delIf oestradiol is given to ewes in the follicular phase followed by 3 low doses of GnRH, an LH surge occurs within 40 h; if additional ACTH is given along with oestradiol, LH surges are blocked. Negative feedback of oestradiol is necessary to accumulate a readily-releasable LH pool, and it is possible that ACTH acts at pituitary level to prevent the replenishment of enough LH for a surge .During the ewe pre-ovulatory period, i.c.v. infusion of CRH suppresses the LH surge causing prolonged oestrus cycles, via effects at the hypothalamus and pituitary .in situ hybridisation studies on the hypothalamus of long-term ovariectomised ewes fail to show an increase in GnIH activity after psychosocial stress , suppresses GnRH/LH secretion in ewes but immunohistochemistry and  review: .infusion (An early series of cow studies using exogenous opioid agonists and antagonists hint that there is opioid suppression of LH release until hour(s) before the LH surge, that is then lifted and the LH surge occurs; however, the suppression is extended by transport (blocking/delaying the LH surge), although the latter is not reversed by one injection of opioid antagonist , 1992a. infusion .From the scarce literature available, mid-cycle FSH is increased by HPA stimulation but this is probably due to a reciprocal reduction in follicular oestradiol after suppression of GnRH/LH pulses. Late follicular phase surges of FSH are delayed in parallel with LH surges .With the back-ground of all our above studies, we have recently focused on two main paradigms:a chronic spontaneously-occurring model comparing clinically compromised cows  with their ‘normal’ herd-mates might reveal ways in which cows utilise coping trade-off strategies, andan acute ewe model with or without insulin or LPS treatment that has focused on how these trade-offs may be controlled by neurotransmitters within the brain.and lameness fail to ovulate  reveals a precise sequence of events . Sniffinintensity is reduced; Lameness does not affect the above sequence or overall duration of oestrus  is reduced by lameness, low BCS or high milk yield no doubt due to the above effects on oestrus intensity (versus 0.029 ng/ml), contrary to luteal values that are markedly lower in lame cows  and the locus coeruleus (LC) with on-going synaptic contacts between these cells and CRH/AVP cell bodies in the hypothalamic paraventricular nucleus (PVN), possibly also with links via the medial pre-optic area mPOA; . As a reSedation induced by sodium pentobarbitone reduces the magnitude or totally blocks the LH surge in cyclic and oestradiol-treated ewes or cows . The expPentobarbitone, diazepam (gamma-amino-butyric acid (GABA) agonist), or xylazine  all immediately decrease basal, as well as transport-induced increases in plasma cortisol, glucose, respiration rate and heart rate , c, 1992afferent sensory input to the brain (perception) or interfere with efferent responses within the brain. Concerning the latter, the paraventricular nucleus (PVN) has alpha-1 adrenergic receptors, is well innervated by adrenergic fibres, and administration of an alpha-1 adrenergic agonist directly into the third ventricle of the brain increases peripheral ACTH and cortisol concentrations  - this response is further enhanced in the presence of oestradiol  neurones is reduced by insulin treatment , complex interactions occur particularly between/within the brain stem and hypothalamus. The hypothalamic mPOA is the main location of GnRH cells and  the transfer of GnRH proceeds along axons to the median eminence for secretion into portal blood, and onward to the pituitary to release LH .There are no adrenergic cell bodies in the hypothalamus but axons extending from noradrenergic cells in the brainstem are in close contact with GnRH cell bodies in the ewe mPOA . There aFurther cell types within the ARC are also activated at varying times before the GnRH/LH surge in ovary-intact ewes . SpecifiKNDy’ cells contain ER and neuropeptides, the latter being both stimulatory ) and inhibitory (Dynorphin) to keep GnRH secretion tightly regulated. However, the overall balance of different neuropeptides within KNDy cells varies throughout the ewe follicular phase. Initially, there is a shift of the net balance towards inhibitory dynorphin before behaviour starts, followed by a swing towards excitatory kisspeptin and NKB after the influence of increasing oestradiol concentrations during the surge  activity in the lower part of the ARC and median eminence . At the Glucocorticoid (cortisol) receptors do not exist on GnRH neurones but are co-localised with progesterone and oestradiol receptors in the mPOA and ARC, possibly in KNDy cells, and it is by this indirect pathway that cortisol signals are transmitted to ewe GnRH neurones . There ipulses in ovariectomised ewes : Activation of ER neurones in the VMN increases 10-fold during ewe sexual behaviour  remain an option  otherwise animals trade-off with lower production, less intense oestrus behaviour, and impaired fertility. The incidence of (sub) clinical problems around calving should be minimalised, but why make cows calve so frequently? Lactation persistency is enhanced by increasing milking frequency, feeding more concentrates during declining lactation, and by genetic selection. Pregnancy rates in later lactation are similar to those soon after calving, and later re-breeding of high-yielding cows improves profitability  until conditions improve. Ultimately, those animals that do develop coping strategies to overcome adverse stimuli, are the ones with the genes that are valuable to achieve trade-offs. From the fore-going discussion, it is clear that survival of the species/individual is so important that the strategy does not depend on a single factor, but is made fail-safe by being multi-factorial, achieved through a complex interplay among excitatory and inhibitory neuronal and hormonal signals that converge on hypothalamic neurones responsible for secretion of GnRH. Understanding the mechanisms of this interplay is of paramount importance for the efficiency of both sexual behaviour and subsequent fertility.Trying to understanding We know that cows and ewes do exert fertility trade-offs, but we need to be sure we are using the right criteria to monitor their impact, e.g., meaningful welfare indices are required, and assessing output per animal/rumen rather than per kg milk/meat may be more appropriate in view of the reverse impact exerted by ruminants upon the environment . Domestication needs to be as efficient as possible both economically and environmentally. However, while optimising the inter-relationship between animals and the environment, caution is urged against the camouflage use of drugs/hormones/feed additives/intricate technologies . In the Finally, there is one phenomenon that remains totally inexplicable. How/why do some animals display oestrus during pregnancy? In one study, ~ 5% dairy cows displayed oestrus (sniffed and mounted by herd-mates) at least once at varying times throughout pregnancy – some even stood willingly to be mounted and mated by a bull. However, dried samples of vaginal mucus did not form characteristic fern patterns; and plasma oestradiol concentrations were similar to non-pregnant oestrus herd-mates, whereas plasma progesterone values were all > 1.5 ng/ml when expressing sexual behaviour . How/why


In this review we have showcased the preclinical development of original amphiphilic nanomaterials designed for ruthenium-based anticancer treatments, to be placed within the current metallodrugs approach leading over the past decade to advanced multitarget agents endowed with limited toxicity and resistance. This strategy could allow for new options for breast cancer (BC) interventions, including the triple-negative subtype (TNBC) with poor therapeutic alternatives. BC is currently the second most widespread cancer and the primary cause of cancer death in women. Hence, the availability of novel chemotherapeutic weapons is a basic requirement to fight BC subtypes. Anticancer drugs based on ruthenium are among the most explored and advanced next-generation metallotherapeutics, with NAMI-A and KP1019 as two iconic ruthenium complexes having undergone clinical trials. In addition, many nanomaterial Ru complexes have been recently conceived and developed into anticancer drugs demonstrating attractive properties. In this field, we focused on the evaluation of a Ru(III) complex—named AziRu—incorporated into a suite of both zwitterionic and cationic nucleolipid nanosystems, which proved to be very effective for the in vivo targeting of breast cancer cells (BBC). Mechanisms of action have been widely explored in the context of preclinical evaluations in vitro, highlighting a multitarget action on cell death pathways which are typically deregulated in neoplasms onset and progression. Moreover, being AziRu inspired by the well-known NAMI-A complex, information on non-nanostructured Ru-based anticancer agents have been included in a precise manner. The prevention, diagnosis, and treatment of neoplastic diseases are a major problem in developed countries . AccordiBC is an extremely heterogeneous malignancy, completely diverging among distinct patients (intertumor heterogeneity) and even within each individual tumor (intratumor heterogeneity) . The femNowadays, as the field of genomics has developed considerably, researchers can perform fast screening to examine gene sequences alterations with their protein products that might be correlated with precise BC subtypes. Interestingly, many of these gene clusters also mapped onto the four major biological markers described above . Due to Currently, different types of treatments are available for patients with BC . In receIn addition, a vast repertoire of chemotherapeutic agents is available, e.g., 5-fluorouracil (5-FU), cyclophosphamide, carboplatin, taxanes such as paclitaxel and docetaxel, anthracyclines such as doxorubicin ,32,33. CCancer can be briefly described as an uncontrolled proliferation of mutant cell clones due to loss of normal controls, resulting in deregulated cell cycle, lack of differentiation, angiogenesis, and local tissue invasion. DNA damage and consequent mutations can also produce devastating effects on the regulation of cell death pathways, thus leading to uninhibited cell proliferation ,38,39. IApoptosis unfolds through two different pathways: the intrinsic pathway—or mitochondria—activated by intracellular signals such as organelles or biomolecular damages , and the extrinsic pathway—or death receptor—activated by extracellular signals, both propagated by caspase cascades ultimately leading to cell death . NormallAutophagy is fundamentally an evolutionary conserved process ensuring cell survival under stress conditions . Originally termed programmed cell death type II, autophagic cell death differs from apoptosis in being caspase independent. It is accomplished by the coordinated activity of a family of autophagy-related proteins (Atg) and can be initiated via canonical Beclin 1 dependent and non-canonical pathways . Basal lA growing number of data proves that dysregulated autophagy, as well as autophagic pathways failure, play a key role in different malignancies onset and progress, including BC ,62. As acDDP) and cytoprotective autophagy in response to docetaxel .,78.cDDP)>250 μM) . IC50 vaHence, preclinical evaluations provided convincing and definitive evidence validating our nucleolipid Ru(III) nanosystems as promising options among advanced therapeutics with superior anticancer activity.cDDP - having nuclear DNA as final target with resultant adducts formation and cell cycle arrest - it has been proved that also AziRu can interact with DNA models, wherein Ru(III) ions can bind stably nucleotide structures . We. We57]. In addition to apoptosis, DOTAP nanoformulations enclosing the nucleolipid AziRu complex trigger extensive and massive autophagy activation in BCC models, including ER and TN subtypes, providing a crucial drive in causing cell death in vitro . Indeed,Breast cancer is certainly among the most studied disease afflicting humankind. This is not surprising given the overall number of women diagnosed with the malignancy, making it a main concern worldwide. In recent years, researchers have made great progress in the management of BC. Nevertheless, mainly because of molecular subtypes heterogeneity and chemoresistance to treatments, the battle is still far from being won and must be fought on several fronts. Although the most exciting area of research are perchance immunotherapy and targeted therapy, chemotherapy is still a necessary weapon to cope with invasive and widespread tumors such as TNBC. Among chemotherapeutics, metal-based drugs have dominated the scene so far, with the research that has increasingly moved on other precious metal alternative to platinum. Many scientists have focused their interest on the design of original ruthenium complexes due to their prospective biomedical and pharmaceutical appliance. Indeed, ruthenium exhibits many ideal drug properties, making it a highly favorable therapeutic agent, including its redox abilities, minimal toxicity profile for healthy cells, high selectivity for cancer cells, ligand exchange properties, and favorable adsorption properties. In this framework, nanostructured materials functionalized with Ru complexes have proved to be a useful and safe way to administer Ru-based drugs for cancer treatment. Herein, starting from ‘traditional’ Ru-based compounds, we have highlighted the recent developments in a specific type of nucleolipid nanosystems enclosing the anticancer ruthenium(III) complex AziRu, including extensive information about the possibility of their decoration and functionalization for a selective and effective drug delivery to tumors. With the aim of expanding the beneficial properties of available amphiphilic Ru(III) complexes, we are investigating novel biocompatible platforms alternative to nucleosides to develop new prospective anticancer agents. Very recently, a trifunctional α-amino acid, in place of the nucleosidic core of amphiphilic Ru(III) complexes, has been selected. The amino acidic scaffold, coupled to the Ru(III) complex, is decorated with both hydrophilic and lipophilic moieties, conferring high tendency to form stable aggregates which are showing promising anticancer activity . MoreoveBeyond chemotherapeutic interventions by nanostructured material, as part of a potential combined targeted therapies planned to improve the anticancer efficacy of Ru-based strategy, and, in the meantime, to overcome the possible occurrence of chemoresistance, a further approach we are pursuing encompasses interference with multiple intracellular oncogenic pathways: in concert with the uncontrolled growth mechanisms counteracted by ruthenotherapy, the simultaneous intercepting of aberrant miRNA expression can be experimented in TNBC patients through the employ of ad hoc designed cationic nucleolipid nanoformulations loaded with both the ruthenium(III) complex AziRu and a selected miRNA with proven antitumor activity towards human BC ,174.Overall, in this review we have attempted to showcase the path toward the preclinical validation of novel amphiphilic nanomaterials designed for effective Ru-based anticancer treatments, to be placed within the current metallodrug approach leading over the past decade to advanced multitarget agents endowed with limited toxicity and resistance. On this point, modern chemotherapeutics must in fact address resistance issues that are afflicting many therapeutic regimens. Besides potent drug combinations, chemoresistance can be overcome by using a multitarget approach to improve anticancer efficacy. Indeed, simultaneous action on multiple molecular targets can increase the chances of reactivating death pathways that are commonly suppressed in cancer cells by the action of oncogenes. To our knowledge, ruthenium complexes show a higher activity in several resistant cell lines, including BCC ,175. In


Dieting regimens such as calorie restriction (CR) are among the most commonly practiced interventions for weight management and metabolic abnormalities. Due to its independence from pharmacological agents and considerable flexibility in regimens, many individuals turn to dieting as a form of mitigation and maintenance of metabolic health. While metabolic benefits of CR have been widely studied, weight loss maintenance and metabolic benefits are reported to be lost overtime when the diet regimen has been terminated—referred to as post-dietary effects. Specifically, due to the challenges of long-term adherence and compliance to dieting, post-dietary repercussions such as body weight regain and loss of metabolic benefits pose as major factors in the efficacy of CR. Intermittent fasting (IF) regimens, which are defined by periodic energy restriction, have been deemed as more flexible, compliant, and easily adapted diet interventions that result in many metabolic benefits which resemble conventional CR diets. Many individuals find that IF regimens are easier to adhere to, resulting in fewer post-dietary effects; therefore, IF may be a more effective intervention. Unfortunately, there is a severe gap in current research regarding IF post-dietary effects. We recognize the importance of understanding the sustainability of dieting; as such, we will review the known physiological responses of CR post-dietary effects and its potential mechanisms through synthesizing lessons from both pre-clinical and clinical studies. This review aims to provide insight from a translational medicine perspective to allow for the development of more practical and effective diet interventions. We suggest more flexible and easily practiced dieting regimens such as IF due to its more adaptable and practical nature. As the rates of obesity and its associated metabolic diseases continue to rise globally, there is a pressing need to develop disease management strategies that are both effective and comprehensive. To combat obesity and its downstream metabolic consequences, many attempt to mitigate these effects through dieting interventions such as calorie restriction (CR) . CalorieDue to CR’s compliance challenges, there has been a shift in interest towards intermittent fasting (IF) regimens, such as time-restricted feeding (TRF) and alternate-day fasting (ADF) ,14. IF iPrior to discussing post-dietary effects of CR, it is important to understand the known benefits and mechanisms of CR. CR involves a reduction in caloric intake, which is often achieved through lowering total caloric intake to an amount below total energy expenditure with a goal of body weight reduction and metabolic improvement . CR studad libitum (free access to food) HFD.Compounding what is already known about CR, there has been overwhelming attention and curiosity surrounding post-dietary effects. The underlying mechanisms of post-dietary effects remain unclear, despite postulations that suggest weight cycling is associated with increased disease risk and may pose a major metabolic health concern. Therefore, understanding this phenomenon and its possible drivers and mechanisms are important to provide insight concerning dieting and disease risk. Much of the current pre-clinical post-dietary studies subject mice to CR with a 30% reduction in normal caloric intake or diet-switch regimens that alternate between a high-fat diet (HFD) and a low-fat diet (LFD), commonly referred to as the “yo-yo diet” ,27. ThisVariability in genetic backgrounds are also investigated in CR studies. Although they are not the focus of this paper, they play an undeniable role in determining the effectiveness of CR-mediated responses in mouse models. CR is shown to decrease IGF-1 (insulin-like growth factor 1), which increases apoptosis and inhibits cell proliferation, thereby delaying tumour progression and expanding longevity. Interestingly, sexual dimorphism is reported in IGF-1 expression . When feConsidering the CR-mediated alterations of the gut microbiome, hormone levels, and adipose tissue remodeling in contributing to metabolic benefits, it would be reasonable to speculate that post-dietary effects of CR may also be attributed to a reversal of these alterations to conditions prior to CR dieting. The notion of post-dietary effects is deemed as the consequences following termination of dieting regimens. In this review, we explore possible drivers such as behavioural and organ system alterations, which contribute to the overall metabolic effects of post-dietary restriction. Taken together, these observations point to the importance of understanding the pathways involved in post-dietary effects in order to develop effective and sustainable diet intervention programs.Although diet interventions often result in successful short-term weight loss and metabolic benefits, these benefits are rarely maintained over time. As such, long-term maintenance of weight loss and the sustainability of metabolic benefits remain important challenges for the treatment of obesity and metabolic disorders. Evidence from mouse models indicates that diet-induced weight loss is associated with enduring behavioural changes such as decreases in activity and intrinsic motivation towards food, thereby contributing to chronic obesity, metabolic dysfunction and relentless post-dietary weight regain . In lighob/ob mice could harvest more energy compared to their lean counterparts [The gut microbiome is one of the key organs involved in energy harvesting and regulation of host metabolism . In a stterparts . These fBacteroidetes and Actinobacteria were increased, while harmful Firmicutes and Verrucomicrobia were slightly decreased. Widely accepted probiotics associated with health-promoting and immunomodulatory properties such as Lactobacillus and Bifidobacterium [Helicobacter, a pathogenic bacteria which predominantly resides in the upper gastrointestinal tract [In a study to investigate the functional role of the gut microbiota in a CR model, antibiotic-induced microbiota-depleted mice were placed on either a normal ad libitum diet or a 30%-reduced CR diet . Depletiacterium ,39 had iacterium . Helicobal tract , was sigal tract . Fecal mal tract . These ral tract . TogetheConsidering the highly sensitive nature and persistent effects of alterations in the gut microbiome, its contribution to the long-term outcomes of dieting is compelling . In a stThe gut microbiome has an active and integral role in regulating metabolic hormones secreted by organs such as the gastrointestinal tract and adipose tissue. It has been shown that depletion of the gut microbiome greatly affects hormone secretion, thereby leading to detrimental effects to metabolic health ,44,45. ABacteroidetes and a significantly greater proportion of Firmicutes in obese mice compared to their lean counterparts [Excess caloric intake is commonly regarded as one of the root causes of obesity, although a distinction in gut microbial populations has been deemed a prominent factor affecting energy, metabolic and endocrine homeostasis. In particular, significant differences in the microbial composition between obese and lean mice have been reported and well-studied . It has terparts . AltogetMechanistic changes in adipocytes are proposed to be a prominent factor driving post-dietary weight regain . White aCol1+ preadipocytes suggest that they are taking on a myofibroblast phenotype with impaired adipogenic differentiation capacity, consequently leading to limited capacity for eWAT expansion and dysfunction [Pref1, Col1a1, Col3a1, Acta2, and Fnl exhibited significantly reduced expression in formerly obese eWAT, which remained at low levels following the HFD rechallenge. This suggests that mice subjected to HFD rechallenge had significantly impaired lipid storage capacity. Overall, the results demonstrate that adipose tissue of obese and formerly obese mice contribute to impaired adipogenesis and the failure to induce new properly functioning adipocyte formation when subjected to HFD rechallenge. This is consistent with previous studies, which demonstrated that prolonged recovery from HFD did not recover the adipose tissue environment, insulin sensitivity, or lipid storing functionality that was disrupted during previous periods of HFD or obesity [In a weight cycling study, mice were subjected to a diet-switch model where mice were fed HFDs, followed by normal chow, then rechallenged again with HFDs to induce weight regain . Resultsfunction . Mature  obesity ,52. TogeThe hypothalamus is the central site where many neuropeptides and hormones act to elicit appetite regulation . HoweverEvidently, body weight maintenance and metabolic homeostasis are intricately regulated by processes such as multi-organ cross-talk, and environmental and behavioural factors. The hypothalamus has a central role in maintaining homeostasis by integrating signals pertaining to energy balance, food intake, and body weight in both pre-clinical and clinical models ,62,63. TDietary interventions are commonly being practiced as non-pharmaceutical treatments for tackling obesity and metabolic diseases; therefore, the need for a more thorough understanding of the benefits and drawbacks of dieting is more imperative than ever. The conventional interventions containing energy-deficit diets such as CR emphasize the critical role of caloric intake in the management of body weight, fat mass, and metabolic abnormalities. Evidence has indicated that CR diets modify cardiometabolic conditions, thereby greatly reducing various risk factors of age-related and cardiovascular diseases, cancers, and metabolic syndromes ,64. HoweDespite this dire need, clinical studies examining the prevalence of weight cycling are limited and the outcomes are relatively inconsistent. While some studies have suggested that the regaining of body weight following diet-induced weight loss is associated with an increased risk for stroke, diabetes, coronary heart disease, and mortality ,67,68, oAla55Val genetic polymorphism in uncoupling protein-2 (UCP-2) was associated with abdominal subcutaneous fat, where ValVal types showed significantly less reduction under CR compared to AlaAla or AlaVal types [Humans exhibit complex and distinctive genomes, and as such, CR may have diverse effects on individuals due to genomic variations. It was demonstrated that single nucleotide polymorphism of the peroxisome proliferator-activated receptor γ (PPARG) gene was responsible for approximately 7% of total reduced body weight variance during a short-term CR regimen (14 weeks) . Furtheral types . These fAs such, the outcomes of current weight cycling studies are largely impacted by the multiple confounding variables. It is rather difficult to isolate the impact of the post-dietary effects. Therefore, a consensual methodology and a more thorough understanding of post-dietary effects are needed to provide insightful, practical, and effective treatments to address obesity worldwide.The post-dietary effects of weight cycling in clinical settings remain inconclusive and unclear. The underlying mechanisms need to be further investigated to develop effective, comprehensive, and evidence-based treatment strategies for obesity and metabolic diseases. Due to the resemblance in genomics between mice and humans, the gut microbiome is heavily studied in clinical studies. Similar to rodent studies , the micFirmicutes and Bacteroidetes are the two dominant beneficial bacterial strains residing in the gut microbiome [Firmicutes compared to Bacteroidetes. After dieting, a dramatic shift in ratio was reported; reduced levels of Firmicutes and elevated levels of Bacteroidetes. The ratio of Firmicutes:Bacteroidetes in obese individuals shifted towards becoming more similar to lean controls following a CR diet, irrespective of macronutrient content. On the other hand, other clinical studies have demonstrated the dynamic and rapidly adaptive nature of the gut microenvironment [Lachnospiraceae and Ruminococcaceae, and increases in Bacteroidetes and Proteobacteria were reported, which highlights that certain residing bacterial strains are more sensitive to dietary interventions, and more specifically that the alterations in microbial populations are quickly adaptive. Thus, post-dietary weight regain loss of metabolic benefits could be the result of the active remodeling of the microbiome, further favoring fat storage. However, although microbiota-mediated post-dietary weight regain have been investigated in rodent studies [Gut microbiota in humans have been reported to be dynamically altered by both CR and nutritional components ,73. Thuscrobiome . Interesironment ,77. In aironment . Decreas studies , whetherBody weight and metabolic homeostasis are mediated by the complex neuro-hormonal system, which coordinates peripheral anorexigenic or orexigenic neuropeptides and hormones . The arcLeptin secreted from adipose tissue predominantly act on POMC neurons in the hypothalamus to induce satiety. In a study where the obese participants underwent a weight-loss program, overall weight loss resulted in a reduction in leptin signaling, which led to reduced or delayed feeding inhibition in the study subjects . FollowiHumans exhibit a much more complex system, whereby extrinsic factors such as the external environment and lifestyle directly affect behaviours, which leads to fluctuations in body weight and metabolic homeostasis. Due to food being conveniently accessible with our modern-day societies, hunger is certainly not the only factor regulating energy intake. Thus, hedonic reward pathways are commonly investigated in clinical settings as it involves cognitive, reward, and emotional factors . PalatabSimilar to reports in rodent models, human adipose tissue alterations are proposed to be involved in the mechanisms driving post-dietary weight regain, because it plays a major role in fat storage ,83. AdipDynamic adipose tissue remodelling has been reported as a result of calorie restriction and overfeeding. A study investigating the initial period of weight gain induced by overfeeding has reported that individuals had an upregulation in genes associated with lipid metabolism and storage, angiogenesis, and extracellular matrix remodelling . SeveralMany other diet-induced mechanisms of adipose tissue remodelling are being investigated in post-dietary studies, such as its metabolic capacity, endocrine functions, and inflammatory responses ,87. For Inconsistencies and gaps in the current literature are rooted in many limitations and challenges associated with the current research on post-dietary weight management and metabolic effects. Some of these barriers include inconsistent definitions and definitive terminology, varying targeted populations, and lack of unified methodologies, thus resulting in major sources of bias between pre-clinical and clinical studies. In the current literature, there are many different key terms used to characterize post-dietary weight regain and metabolic phenotypes ,28,61,90IF regimens and their abundance of variations have revealed many benefits that resemble conventional CR regimens. Early time-restricted feeding (eTRF) has been commonly used as a form of IF in human studies. Sutton et al.’s study  demonstrAlthough IF has been shown to be a promising diet intervention in pre-clinical and clinical models, further research is needed to delineate the post-dietary effects of IF. While IF regimens are flexible, more easily adaptable to suit various lifestyles and often include isocaloric consumption, CR regimens mainly focus on chronic and consistent caloric restriction which likely drives adapted responses in metabolic organs and systems such as adipose tissue, gut microbiome, and hormone levels. Due to the adaptation of chronically reduced caloric intake, an increase in energy absorption and storage often leads to downstream metabolic impairments. In contrast, due to the ability to maintain isocaloric consumption periods in IF regimens, the efficiency of energy utilization can be sustained. Under IF conditions, metabolic organs and systems are able to be well adapted to consistent levels of caloric intake. Therefore, due to the fundamental difference between IF and CR regimens, the outcome of post-dietary effects of IF may also be different from CR. While we can synthesize, extrapolate, and apply the mechanisms known from CR post-dietary research, more work is required to investigate the metabolic effects of IF post-dieting among both pre-clinical and clinical models.


Recent advancements in molecular testing, the availability of cost-effective technology, and novel approaches to clinical trial design have facilitated the implementation of tumor genome sequencing into standard of care oncology practices. Current models of precision oncology practice include specialized clinics or consultation services based on a molecular tumor board (MTB) approach. MTBs are comprised of interprofessional teams of clinicians and scientists who evaluate tumors at the molecular level to guide patient-specific targeted therapy. The practice of precision oncology utilizing MTB-based models is an emerging approach, transforming precision genomics from a novel concept into clinical practice. This rapid shift in practice from cytotoxic therapy to targeted medicine poses challenges, yet brings exciting opportunities to clinical pharmacists practicing in hematology and oncology. Only a few precision genomics programs in the United States have a strong pharmacy presence with oncology pharmacists serving in leadership roles in research, interpreting genomic sequencing, making treatment recommendations, and facilitating off-label drug procurement. This article describes the experience of the precision medicine clinic at the Indiana University Health Simon Cancer Center, with emphasis on the role of the pharmacist in the precision oncology initiative. Translation of genetic testing into standard clinical practice in oncology was pioneered by scientific and clinical efforts in areas related to pharmacogenetics and pharmacodynamics . Recent ® (Olaparib) for first-line maintenance therapy in BRCA-mutated advanced ovarian cancer [Identification of somatic mutations as oncogenic drivers of malignancy has revolutionized the treatment of various malignancies such as chronic myeloid leukemia and non-small lung cancer . We owe n cancer ,15,16. BThe Precision Medicine Initiative, also known as the Cancer Moonshot Initiative, was established by President Barack Obama and led by Vice President Joe Biden in 2016. This initiative was launched with a $215 million investment to pioneer a new model of patient-powered research led by the National Institutes of Health (NIH) and $70 million dedicated to the National Cancer Institute (NCI), to scale up efforts to identify genomic drivers in cancer and to develop more effective approaches to cancer treatment . CurrentThe NCI-MATCH and TAPUR trials are not only expanding the reach of precision genomics, but their results are also anticipated to provide tremendous amounts of new genomic information to the oncology community. Despite the advances in genetic technology, there are challenges to translating the results of genetic sequencing into patient care. This calls for the need of an enterprise consisting of multidisciplinary teams equipped with clinical and scientific tools to help prepare the oncology community to integrate precision medicine into clinical practice.Current models of the precision oncology practice include specialized clinics, consultation services, or molecular tumor boards (MTB) ,24,25,26The pharmacist’s role in precision medicine practice models includes but is not limited to interpreting tumor genome sequencing results, managing anticoagulant use around the timing of tissue biopsy, leading patient education, and obtaining off-label drug therapy . HoweverThe IUSCC precision genomics program (PGP) was established in 2014 by a multidisciplinary team, including a medical oncologist, a medical and molecular genetic scientist, and an oncology pharmacy specialist. The program now includes a diverse team of medical oncologists, scientists, an oncology pharmacy specialist, two nurse coordinators, two genomic counselors, a data manager, two rotating PGY-2 oncology pharmacy residents, and an oncology fellow . The PGPPatients with advanced or rare malignancies are referred to the program by their primary oncologists from both within IU Health and external health systems. A dedicated nurse coordinator schedules patients for an initial visit with the PGP oncologist to introduce patients to the program, obtain past medical history, and complete consent for biopsy. The PGP oncologist specifies the biopsy site, and a laboratory personnel conducts tumor sequencing on the biopsy specimen. The biopsy of the tumor can be performed at the initial visit, or tumor samples from other recent biopsy procedures can be sent to Foundation Medicine for NGS. Before the biopsy is scheduled, the oncology pharmacist reviews patient medications to identify those that may increase the risk of bleeding or impair wound healing. These medications include oral and intravenous chemotherapy agents, such as vascular endothelial growth factor (VEGF) inhibitors, and anticoagulants. Recommendations are made by the pharmacist to hold the high-risk medications for a specified amount of time before proceeding with the biopsy. Treatment options are proposed based on the sequencing results and an extensive literature review during a weekly pre-MTB meeting. The weekly MTB conference consists of medical oncologists, oncology pharmacists, oncology nurses, pathologists, genetic counselors, residents, fellows, and a phase I clinical trials team. The past medical history and tumor sequencing results of each case are presented by either the medical oncologist, genetic scientist, or oncology pharmacist, followed by a discussion of genomic targets and potential therapies. Decisions regarding therapy are given with considerations to patient-specific factors  and genomic factors . To standardize targeted therapy recommendations at IUSCC PGP, identified targets are chosen, with first priority given to DNA mutations, copy number variations, and DNA fusions; second priority to mutations detected by immunohistochemistry or by florescence in situ hybridization; third priority to mRNA overexpression alone. The oncology pharmacist documents the recommendations from the tumor board conference into the EMR. An overview of the precision oncology practice model at IUSCC is shown in After the results are finalized, the patient has a return clinic visit with the medical oncologist, scientist, and pharmacist to discuss the genomic results, molecular biology of genomic aberrations, potential therapeutic options, and if appropriate, clinical trial referral options. A genetic counselor sees all patients with clinically relevant germline mutations that are detected from the tumor biopsy. The oncology clinical pharmacist and PGY-2 oncology pharmacy residents will meet with each of the returning patients for 30 to 45 min to educate them regarding the genomic findings, treatment recommendations, medication details , and clinical trial information. It is crucial to assess patients’ health literacy before each counseling session and use effective communication skills, such as active listening, using patient-friendly language, and assessing patient understanding. Each counseling session will begin by introducing the patient to what will be discussed during the session: a brief overview of genetics, a review of the precision medicine program and the process of how the treatment recommendations are made. Visual demonstrations such as simple drawings of cancer genetics pathways as well as easy-to-understand analogies are utilized when explaining the molecular biology of genetic mutations and the mechanism of action of treatment. Described below is an example of the education session that was conducted for a 61-year-old male with BRAF-mutated metastatic colon cancer who was seen at IUSCC PGP clinic. Pharmacist:“We are going to start with briefly introducing you to what we do here at the Precision Genomics clinic, explain the process of how we came up with the treatment recommendation based on the sequencing results, and go over the recommended treatment with you. At this clinic, we view cancer at a genetic level. We took a biopsy of your tumor, sent a sample to a lab that utilizes technology to identify if there are genetic changes in your tumor that we can target with medications. After we obtain the results from your biopsy, we present the findings to a multidisciplinary tumor board, which consists of 15 to 20 individuals including oncologists, pathologists, nurses, pharmacists, and scientists to discuss and devise treatment specific to you. What questions do you have so far?”Pharmacist:“As you see here in this drawing, every cell has a DNA, which is the blueprint of life as it contains information needed for individuals to grow and survive. Cells must divide in order for us to survive but there must be a balance between cell division and cell death. As we age and with increased exposure to environmental factors and genetic changes, we may acquire certain mutations or “mistakes” in our genes that can lead to the development of cancer. From your tumor biopsy, we found something called a BRAF mutation. BRAF is a gene that tells the DNA to make a protein called B-Raf, which is involved in a pathway called the MAPK pathway that is responsible for controlling how quickly your cells divide, and it acts as a gas pedal that tells your cell to grow and multiply (the pharmacist will use a drawing to show this pathway). Normally, BRAF has an off-switch that controls when the cells should divide. However, when BRAF is mutated, the growth signal will always be on causing cancer cells to grow and divide at a much faster rate than normal.” After answering patient’s questions, the pharmacist proceeds with explaining in detail the recommended treatment including mechanism of action, dose, administration, side effects, drug-interactions, etc.The genomic findings and treatment recommendations are also summarized in writing and given to the patient along with PGP contact information, drug information, and if available, information on the recommended clinical trials. The summary of each visit is documented in the EMR by PGY-2 oncology pharmacy residents and communicated to the referring primary oncologist. The subsequent patient interactions related to clinical trials consultation and drug procurement, including letters of medical necessity, prior authorizations, and peer to peer discussions are completed by the oncology clinical pharmacist or PGY-2 oncology pharmacy residents. p < 0.0001). Furthermore, patients treated with genomically guided therapy had a superior median PFS compared to those treated with non-genomically guided therapy  [In 2016, the precision genomics team including the clinical oncology pharmacist at IUSCC led a formal prospective cohort study evaluating the efficacy of a precision medicine approach, using genome-guided therapy designed by IUSCC multidisciplinary MTB team members. In this study, 101 patients with metastatic solid tumors who progressed on at least one line of standard of care therapy were evaluated from April 2014 to October 2015. The median number of prior regimens was four in both groups. The most common malignancy diagnoses in the genomically vs. non-genomically-directed therapy groups were soft tissue sarcoma (22.7% vs. 19.3%), breast (18.2% vs. 14%), and colorectal cancer (15.9% vs. 8.8%). Of 44 patients treated with genomically guided therapy, 19 (43.2%) patients attained a progression free survival ratio ≥ 1.3 vs. three of 57 (5.3%) patients treated with non-genomically guided therapy (37–0.84) . At IUSCC, the hallmark feature demonstrating the successful experience of transforming precision oncology from a scientific concept to clinical practice was the interprofessional dynamic and strong team-based approach. Particularly, the oncology pharmacy clinical specialist has a leading role throughout the continuum of precision genomics and is heavily relied upon by physicians and nursing colleagues. Key responsibilities of the oncology clinical pharmacist at IUSCC PGP include collaborating with oncologists and scientists to interpret genetic sequencing and provide recommendations, documenting clinical notes in the medical record, educating patients, facilitating off-label drug procurement, and ensuring patient safety when obtaining tissue biopsies.Interpretation of genetic sequencing and provision of treatment recommendations require scientific knowledge of oncogene pathways, tumor suppressor genes, the clinical significance of genetic variance, treatment toxicities, etc. Clinical and scientific proficiency in this area may be achieved via certificate-based training programs and extensive self-directed learning, as well as knowledge of clinical trials, drugs in development, and existing targeted therapies.As experts in pharmacotherapy and oncology treatment, the clinical pharmacists at IUSCC PGP lead the management of anticoagulation and other interacting therapies, to ensure patient safety prior to obtaining tissue biopsy for genomic testing. Timing of anticoagulation and VEGF inhibitors administration are only a few examples of therapy interventions that clinical pharmacists manage at IUSCC.The PGP at IUSCC serves as a one-stop-shop that provides comprehensive services, which greatly impacts patient access to targeted therapies and decreases delay in treatment initiation. This is another area where the oncology clinical specialist plays a leadership role within the genomics program. Being experts in pharmacotherapy, clinical pharmacists when compared to physicians or other allied providers are better equipped to take ownership of the process of obtaining medications, communicating with insurance companies on behalf of physicians, and navigating patient assistance programs. When oral targeted therapy is recommended, a prescription is sent to IUSCC specialty pharmacy. The specialty pharmacy team then submits prescription claims to the patient’s insurance. Outside of clinical trials, the recommended targeted therapy can be off-label, which is often denied by insurance companies. If prescription coverage is denied, the PGP clinical pharmacist sends an appeal letter and performs peer to peer discussions after gathering extensive literature to support the recommendation. If multiple attempts are denied, the clinical pharmacist seeks additional financial resources, such as applying for pharmaceutical manufacturer patient assistance programs. While the process of off-label drug procurement can be time consuming, it is an essential step to ensure patient access to genome-directed therapy.Finally, selection of a particular practice infrastructure may be influenced by the institution’s payment model. Financial sustainability is key when implementing a novel service such as precision oncology. With a healthcare payment model dominated by fee-for-service in the United States, an ambulatory clinic model that bills for individual clinic visits and laboratory tests can be viewed as revenue generators. In contrast, consultation services and MTBs are more likely to be viewed as cost centers whose services might need to be justified. Regardless of what avenue is pursued when implementing precision genomics in oncology practice, oncology pharmacists hold essential clinical and leadership roles in many aspects of precision oncology practice.The number of clinically relevant, druggable genetic changes and approved targeted therapy are anticipated to increase at an exponential rate, given advancement in genomic testing technologies and efficient clinical trial designs. This rapid shift in practice from cytotoxic therapy to targeted medicine poses challenges yet brings exciting opportunities to pharmacists practicing in hematology and oncology.Incorporation of genomic testing and targeted therapy is considered standard of care in many solid tumor malignancies including lung, colorectal, and gastrointestinal . While tThe practice of precision oncology may not be a new concept anymore, but it is still considered a novel approach to patient care in oncology. The IUSCC PGP demonstrated clinical benefit in using precision genomics guided therapy in advanced and refractory solid tumors. These encouraging findings indicate an opportunity for advancing the practice of precision oncology via research. Examples of areas for future research include efficacy of off-label, histology-independent treatment, rare patient cases with successful outcomes associated with targeted therapy, impact of precision genomic programs on improving patient access to targeted therapy, and expansion to multicenter studies.To successfully implement precision genomics services and conduct research, pharmacists need to be equipped with adequate knowledge and proficiency in interpreting genomic testing and targeted therapy selection. This reveals gaps in pharmacy basic and higher-level education . In the Opportunities to bridge current gaps in pharmacy education include offering PGY-2 oncology pharmacy residents practice exposure to facilitate understanding of molecular biology, genomic signaling pathways associated with tumorigenesis, genomic sequencing technology, and targeted therapy selection. Understanding that an on-site experience may not be possible, other avenues for experiential learning include off-site rotations, webinars, case discussions, MTB meetings organized by the American Society of Clinical Oncology, etc. Another approach that can potentially reach a larger population of learners include a virtual learning experience led by experts in the field, consisting of a month-long rotation with various activities such as topic discussions, patient cases, and journal presentations.The practice of precision oncology utilizing an MTB-based model is an emerging approach transforming precision genomics from a novel concept into clinical practice. Only a few precision genomics programs in the United States have a strong pharmacy presence with clinical pharmacy specialists serving leadership roles in research, interpreting genomic testing, and making treatment recommendations. Recent advancements in precision oncology have revealed gaps in pharmacy didactic curriculum and experiential education in precision medicine and genomics. Pharmacists in clinical practice and academia have a professional obligation to transform the education system and to prepare trainees for opportunities in precision medicine and contemporary practice.


Zika virus (ZIKV) is an emerging pathogen associated with neurological complications, such as Guillain-Barré syndrome in adults and microcephaly in fetuses and newborns. This mosquito-borne flavivirus causes important social and sanitary problems owing to its rapid dissemination. However, the development of antivirals against ZIKV is lagging. Although various strategies have been used to study anti-ZIKV agents, approved drugs or vaccines for the treatment (or prevention) of ZIKV infections are currently unavailable. Repurposing clinically approved drugs could be an effective approach to quickly respond to an emergency outbreak of ZIKV infections. The well-established safety profiles and optimal dosage of these clinically approved drugs could provide an economical, safe, and efficacious approach to address ZIKV infections. This review focuses on the recent research and development of agents against ZIKV infection by repurposing clinical drugs. Their characteristics, targets, and potential use in anti-ZIKV therapy are presented. This review provides an update and some successful strategies in the search for anti-ZIKV agents are given.


During the Coronavirus (COVID-19) pandemic, in-person interpreters have been deemed “nonessential,” and thus eliminated to minimize viral exposure and conserve personal protective equipment. Considering alarming patterns of interpreter underuse, we evaluate how substitution for remote modalities (telephone or video) may exacerbate existing inequalities for patients with limited English proficiency. The inherent intimacy, dynamic physicality, and cultural nuances of labor and delivery pose unique communication challenges. Using clinical scenarios, we illustrate the vital role interpreters have in providing accessible obstetric care. We argue that eliminating in-person interpreters in this setting is not justified by COVID-related harms given the potential to exacerbate underlying health disparities. This well-intended infection control measure may reinforce disparities. Limited English proficiency (LEP) can contribute to clinically meaningful and potentially morbid misunderstandings. This is substantiated in the obstetric setting, as illustrated by a confounder-adjusted analysis of parturients whose first language was not English , however, does not justify its use as a substitute in all settings. There are some circumstances in which remote methods are preferred,11–13 and communication breakdown may leave permanent scars. Technical and logistical challenges of remote technology may prompt reduction of clinical communication to the bare minimum.7 This lack of nuance increases confusion among patients, leaves unanswered questions, undermines of trust, and may contribute to lasting traumatic memories of obstetric care.13 Cultural, societal, and family norms of childbirth are unique and important to elicit. Pregnant women who spoke English less than “very well” were more likely to report not having an active role in decision-making with respect to their labor.12 Postpartum women with LEP expressed a strong preference for in-person interpretation compared with remote modalities.9 In a randomized trial among patients seen in a tertiary care emergency department, a setting comparable with labor and delivery triage, those allocated to the in-person arm were significantly more satisfied with their interpreter service, significantly more satisfied with their physicians, and rated their ability to communicate significantly higher than the patients allocated to telephone or ad hoc interpretation (p<0.001).14 Although some patients identify the benefit of anonymity with telephone interpretation in settings where the need for privacy is high and the nature of communication is straightforward, communication quality and effectiveness was overall improved with in-person interpreters.15Language barriers impact health outcomes and patient satisfaction,16 They can act as patient advocates, empower cross-cultural physician–patient relationships, and directly contribute to the therapeutic bond. Although interpreters were equally satisfied with remote and in-person modalities for straightforward information exchange, they found in-person exchange was more satisfactory for establishing rapport and facilitating physician understanding of patients' cultural background.17 Interpreters cite that being in-person to “read the room” and assess body language is key for effective and accurate communication.16Interpreters are uniquely equipped to assess how well information has been communicated with all parties involved. In-person interpreters enable a genuine dialogue by conveying patients' desires, questions, and fears. To develop trust in complex dynamic settings, such as when more than two people are in a room, as is commonly the case during labor and delivery, most interpreters preferred in-person encounters.18 Similar to interpreters, providers rated in-person services significantly higher than video and phone resources, citing frequent technical difficulties while using remote methods.9 Participating in nonverbal communication, serving as a culture broker/patient advocate, and being able to gather more accurate information from patients were cited as benefits of in-person services.7 As providers are principally responsible for orchestrating the involvement of interpretation services, their perspective is paramount.Providers were more likely to indicate improved understanding of patients' cultural beliefs with in-person interpreters compared with video interpretation.19–22 Title VI of the Civil Rights Act mandates health care organizations that receive federal funding must provide patients access to language services,23 but patients report often just “getting by.”22Eliminating in-person interpreters in favor of remote services during the COVID-19 pandemic devalues in-person interpreters and is consistent with underutilization trends. Despite availability of professional interpreters and what we know about the relationship between language barriers and clinical outcomes, certified interpreters are used for <20% of patients who would benefit.ad hoc interpreters such as family members or staff.5 Despite availability of interpreters, providers cite time constraints and organizational-level considerations, including interpreter and telephone availability, among the factors limiting interpreter use.21 Using ad hoc or untrained interpreters increases communication errors such as omission of information, word addition or substitution, and editorialization.24 Universal masking in clinical settings is an additional physical barrier to relationship building25 and clear communication that may be particularly significant across language barriers with elevated importance of facial expression. Eliminating in-person interpreters as nonessential in the care of patients with LEP during COVID-19 compromises communication and sacrifices a critical patient advocate, exacerbating disparities in an at-risk population.Residents frequently rely on their own limited language skills or Wholesale elimination of in-person interpreters during COVID-19 is a mechanism by which health crises hit our vulnerable patients the hardest. The concurrent restriction of most in-person visitors and universal masking requirements exacerbate potential harms and heighten the need for critical patient advocates. Use of remote instead of in-person interpreters is a compromise in quality of care that we should not be willing to accept on our patients' behalf. Although we advocate for broader use of interpreters overall, we acknowledge that not all encounters equally benefit from in-person services. Optimal use of available interpretation modalities, including the decision to invoke an in-person interpreter in specific circumstances, requires judicious clinical judgement.Minimizing health disparities among patients with LEP requires linguistic support. The pandemic has highlighted the need to re-examine and reprioritize in-person interpreter use in clinical settings. Going forward, we encourage institutions and governing bodies to promote this effort accordingly.


The underlying pathophysiology of Parkinson's disease is complex, but mitochondrial dysfunction has an established and prominent role. This is supported by an already large and rapidly growing body of evidence showing that the role of mitochondrial (dys)function is central and multifaceted. However, there are clear gaps in knowledge, including the dilemma of explaining why inherited mitochondriopathies do not usually present with parkinsonian symptoms. Many aspects of mitochondrial function are potential therapeutic targets, including reactive oxygen species production, mitophagy, mitochondrial biogenesis, mitochondrial dynamics and trafficking, mitochondrial metal ion homeostasis, sirtuins, and endoplasmic reticulum links with mitochondria. Potential therapeutic strategies may also incorporate exercise, microRNAs, mitochondrial transplantation, stem cell therapies, and photobiomodulation. Despite multiple studies adopting numerous treatment strategies, clinical trials to date have generally failed to show benefit. To overcome this hurdle, more accurate biomarkers of mitochondrial dysfunction are required to detect subtle beneficial effects. Furthermore, selecting study participants early in the disease course, studying them for suitable durations, and stratifying them according to genetic and neuroimaging findings may increase the likelihood of successful clinical trials. Moreover, treatments involving combined approaches will likely better address the complexity of mitochondrial dysfunction in Parkinson's disease. Therefore, selecting the right patients, at the right time, and using targeted combination treatments, may offer the best chance for development of an effective novel therapy targeting mitochondrial dysfunction in Parkinson's disease. Parkinson's disease (PD) is a common disorder, with over 6 million individuals affected globally . The pathophysiology underlying the vast majority of IPD patients is complex and to date, only partially understood  imbalance and possibly other indirect influences on mitochondrial function from unrelated pathways  clusters, control of cell division, and growth have also been shown to be altered in IPD and mPD , which is then converted to H2O by GSH, as part of the network that removes H2O2  and complex III (C.III), and react with molecular oxygen to form superoxide  oversimplifies their complex role in controlling cellular homeostasis. However, the complexity could also provide an opportunity for drug development, as many cellular alterations in PD (and other neurodegenerative disease for that matter) can be modulated by alleviating mitochondrial dysfunction. Genetic PD may be both primarily mitochondrially involved but also indirectly involved; therefore, targeting only the mitochondria will be insufficient as a stand-alone therapeutic approach. This may particularly be the case if mitochondrial dysfunction is the consequence rather than the cause of disease pathophysiology   fosters this approach. However, the extension of potential findings to the vast majority of IPD patients remains challenging. Polygenic risk scoring of in-silico annotated single nucleotide polymorphisms (SNPs) could translate the concept of mitochondrial impairment found in monogenic PD to IPD  or the in-vivo assessment of key aspects of mitochondrial dysfunction via specific neuroimaging methods is likely to be highly advantageous. Magnetic resonance spectroscopy imaging (MRSI) could provide a non-invasive approach to assess mitochondrial bioenergetics. 31Phosphorus-MRSI (31P-MRSI) bears the potential to measure ATP among other phosphorus-containing metabolites  can be applied to measure lactate levels as a surrogate marker of impaired oxidative phosphorylation  shows potential for discerning in situ mitochondrial impairment  or MRI sequence optimization, offer great potential for MRSI studies  can be employed to study the oxidation status of cytochrome c by multiwavelength absorption and reflection of cortical cytochrome c in the near-infrared spectrum of light , which is one major prerequisite for theranostic-accompanied drug trials  and its derivatives are still the most studied investigational products  and its derivatives are also currently under investigation, where the underlying rationale is to normalise redox levels . NAM, particularly NAD+ and NADH, are also highly relevant for C.I and could be beneficial for ETC disturbances  has also been investigated in a small proof-of-concept study. It showed antioxidative properties measured by increased blood and brain glutathione levels after single-time point administration .The most recent clinical trial candidate showing promise is ursodeoxycholic acid (UDCA), a drug often used in chronic inflammatory liver disease with an extensive safety profile  led to a reduction of mitochondrial factors adenosine monophosphate (AMP)-activated protein kinase, PGC-1α, and tyrosine hydroxylase (TH), and increased expression of silent information regulator T1, TFAM, and Furthermore, endurance exercise restored motor function and reduced apoptosis in a MPTP mouse model -SIRT1-PGC-1α axis may be the most promising approach to enhancing mitochondrial biogenesis, and could involve PGC-1α activating drugs targeting PPAR, AMPK, and SIRT1  or inactivation (missense variants) of the parkin protein   was recently investigated as a protein-based therapy in cellular and animal-based models -antioxidant response element (ARE) signalling cascade plays a key role in several aspects of mitochondrial homeostasis, such as mitochondrial biogenesis, mitophagy, ROS production and scavenging , copper (Cu), and zinc (Zn) levels], their macromolecular organisation (e. g. by forming Fe-Sulphur cluster or heme groups) and their respective distribution in cells  seems to be protective against MPTP-induced toxicity in animal models  are non-coding RNA fragments approximately 22 nucleotides in length that modify gene expression. There is considerable complexity in miRNA mediated gene expression given that one gene can be regulated by many miRNAs and one miRNA can influence many genes  are intimately linked by specialised mitochondria-ER contact sites (MERCs), which act as foci of enzymatic activity  deficiency or inhibition can attenuate parkinsonism in MPTP-treated mice  refers to the use of light in the red to infrared wavelength range to enhance mitochondrial function by displacing nitric oxide, which competitively binds the molecular oxygen site on C.IV of the respiratory chain Hamblin, . Neverthin vitro and in vivo studies of PBM in PD models have explored variations on the theme  is primarily based on the appearance of motor symptoms (Postuma et al., Mitochondrial dysfunction plays a central, multifaceted role in the pathogenesis of PD , whetherThe future of neuroprotective trials will likely rely on combinations of drugs acting directly and indirectly on affected pathways using personalised precision medicine (Titova and Chaudhuri, All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Exercise-induced oxidative stress is linked with the expression level of endogenous antioxidants, but these antioxidants cannot overcome all oxidative stress-related damages in the cells, particularly when cells are under physiological stress. Sometimes, compounds are needed for cellular function, which are produced/activated within the cells, and these compounds can be synthesized by performing exercise, especially high-performance exercise. Taurine is a sulfur-containing amino acid used for various physiological functions. However, its synthesis and accumulation under the oxidative environment may be compromised. Recently, we have shown that taurine level is increased during exercise performance with a decrease in oxidative damage in overused muscles. Other studies have also shown that short-term supplementation with taurine increased physiological performance during severe work intensities, suggesting the role of taurine in improving muscle performance during exercise. However, its precursor cysteine is used in the synthesis of other compounds like GSH and Coenzyme A, which are important for regulating the redox system and energy homeostasis. It is, therefore, important to understand whether taurine synthesis within the cells can blunt the activity of other compounds that are beneficial in preventing oxidative damage during intense exercise. Furthermore, it is important to understand whether taurine supplementation can prevent the conditions observed in the physiological stress of muscles. This review discusses how taurine synthesis could alter exercise-induced ROS generation and the relationship between the physiological stress of muscle and subsequent improvements in exercise performance. Skeletal muscle is one of the most specialized tissues with plasticity in the human body. It accounts for approximately 40–50% of the total body mass, and it can easily withstand different stimuli from exercise-induced mechanical stress and metabolic stress to other environmental stimuli, including heat exposure and nutrient availability . These aReactive oxygen species (ROS) play a crucial role in the exercise-induced pathophysiology of muscle, but it occurs in an exercise-dependent manner . ModeratMuscle-induced oxidative damage is often unavoidable and requires therapeutic interventions, e.g., supplementary foods containing antioxidants, that regulate the redox process . HoweverMuscle overuse affects taurine synthesis by producing ROS, and consequently, taurine-induced benefits are compromised. Taurine biosynthesis is achieved as a result of the following biochemical reactions: oxidation of cysteine by cysteine dioxygenase to form cysteine sulfinic acid; followed by decarboxylation of cysteine sulfinic acid by the cysteine sulfinic acid decarboxylase producing hypotaurine, and finally the oxidation of hypotaurine to taurine. Exercise-induced ROS generation can alter this sequence . As cystExercise affects the taurine turnover , but it Exercise endurance is proportional to the mitochondrial function  and the 2O2 through regulating mitochondrial membrane potential and preventing the formation of semiubiquinone radical  increased concentrations of taurine in the plasma increase the calcium uptake and release to contractile filaments, which further enhances the force production; (II) regulating muscle membrane; and (III) increasing the mitochondrial buffering . Speed aTaurine’s metabolic action in maintaining normal physiological homeostasis may be related to its action on glucose uptake, insulin sensitivity, and further fat oxidation in the skeletal muscle. Studies have shown that taurine improved insulin sensitivity in the fructose-fed rat to stimulate glycolysis and gluconeogenesis . TaurineTaurine affects the availability of antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), but its effect on antioxidant enzymes depends on its concentration. Taurine has increased the antioxidant enzyme activity in a dose-dependent manner in malathion-induced oxidative stress . It alsoMuscle overuse may require a patient to take additional supplements, the function of which is to alter the inflammatory response and reduce oxidative stress without affecting muscle function. However, the duration of supplement administration must be established and monitored. Taurine reverses muscle function under overuse conditions in several ways, including by controlling mitochondrial ROS production, regulating membrane potential, preventing DNA damage, and inducing muscle regulatory proteins. ROS could be the mediator in organizing these conditions. Synthesis of taurine as a means of endogenously improving exercise performance is a major task. Optimal ROS generation due to exercise may facilitate the synthesis of taurine in various ways, such as by primarily attacking sulfhydryl (-SH) terminal of cysteine, the location where several cellular events are organized including the synthesis of taurine. However, how this is achieved through exercise-mediated ROS is yet to be established. Furthermore, the mechanism that drives ROS-induced effects in combination with taurine supplementation must be established in both human and animal models.AT, RP, JB, BI, and YG conceived the idea and wrote the manuscript. All the authors contributed to the final version of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Heart rate variability (HRV) offers insights into humoral, neural and neurovisceral processes in health and disorders of brain, body and behavior but has yet to be fully potentiated in the digital age. Remote measurement technologies (RMTs), such as, smartphones, wearable sensors or home-based devices, can passively capture HRV as a nested parameter of neurovisceral integration and health during everyday life, providing insights across different contexts, such as activities of daily living, therapeutic interventions and behavioral tasks, to compliment ongoing clinical care. Many RMTs measure HRV, even consumer wearables and smartphones, which can be deployed as wearable sensors or digital cameras using photoplethysmography. RMTs that measure HRV provide the opportunity to identify digital biomarkers indicative of changes in health or disease status in disorders where neurovisceral processes are compromised. RMT-based HRV therefore has potential as an adjunct digital biomarker in neurovisceral digital phenotyping that can add continuously updated, objective and relevant data to existing clinical methodologies, aiding the evolution of current “diagnose and treat” care models to a more proactive and holistic approach that pairs established markers with advances in remote digital technology. Remote Measurement Technologies (RMTs) refers to, “any mobile technology that enables monitoring of a person’s health status through a remote interface, with the data then either transmitted to a healthcare provider for review or to be used as a means of education for the user themselves” . Advancein situ using data from personal digital devices  efferent neurons that mediate the function of effector organs. ANS function is generally beyond conscious control and functionally, morphologically and chemically organized into two branches:i.the parasympathetic nervous system (PNS) promotes vegetative activity, such as heart rate (HR) deceleration via the vagus nerve or increasing gut motility predominantly via acetylcholine (Ach) at the neuroeffector junction. The vagus nerve also maintains tonic inhibitory control of proinflammatory cytokines via Ach release into the reticuloendothelial system , mediating the inflammatory reflex through the cholinergic anti-inflammatory pathway .ii.the sympathetic nervous system (SNS) serves to upregulate effector organ function, such as raising blood pressure (BP) or increasing sudomotor activity, via the catecholamines, noradrenaline (NA), and adrenaline .Cardiac tissue has inherent pacemaker properties and the ANS regulates the myocardium’s contractile and electrical output via the vagus nerve and SNS . The ratThe application of spectral analytical techniques to short or long-term neurocardiovascular changes is now widely utilized as a measure of cardiovagal activity. Power spectral analysis can be performed using parametric or non-parametric methodologies. The Fast Fourier transformation (FFT) non-parametric method is typified by discrete peaks of the frequency bands. FFT is a simple and quickly performed equation. The Autoregressive model results in a continuous spectrum of events. It is more complex than the FFT model and must be suitable to the experimental model . High frExercise-induced increases in LF-HRV are linked with metabolic activity in insula, cingulate and somatomotor regions , and HF-“Interoception” is the term given to the transmission of afferent peripheral sensory information, which informs autonomic mediation of homeostasis, allostasis and contributes to psychological and behavioral processes , 2018a. Vagal nerve interoceptive function also has a central role in inflammatory processes via the cholinergic anti-inflammatory pathway. Peripheral proinflammatory mechanisms can be initiated through sympathetic innervation of lymphoid tissue, and anti-inflammatory processes can be promoted through vagal release of Ach or hypothalamic release of corticotrophin-releasing hormone . The vagAs understanding has improved about the role of inflammation in not only neurodegeneration and related neuropsychiatric symptoms , but alsIt could be argued that the current COVID-19 pandemic and related social distancing guidelines have strengthened the case for RMT use in clinical care. Future outbreaks are conceivable and social isolation is likely to be recommended for high-risk groups even after social distancing restrictions are eased, with reduced clinic contact indefinitely suggested for such groups. Social isolation can lead to depression, anxiety, loneliness and hinder clinical care. Loneliness is a modern epidemic that pre-dated COVID-19 and which older adults, who are most at-risk of infection from COVID-19, are particularly susceptible to due to retirement, being widowed, adult children moving away and ill-health causing functional decline and making social activities more difficult . Lonelin1 . DLB has poor prognosis and diagnosis can be complicated by its initial similar presentation to AD, yet early differentiation of DLB from AD is vital due to differing responses to medication and disease courses. Therefore, identifying cheaper yet reliable biomarkers that can differentiate AD and DLB are much-needed. Dysautonomia (autonomic dysfunction) and autonomic failure  are comotension  due to sotension . The impotension  and presotension , genitouotension , gastroiotension , and theotension . Dysautootension , though otension . 123I-MIase  from no (SPECT)  in Lewy Moreover, research has typically not found autonomic symptoms to occur in AD, yet very recent studies have found orthostatic hypotension can present in 42% of AD patients if head-up tilt table testing is used rather than standing tests or subjective self-report measures . In addiFurthermore, capturing acute disease-related episodes, such as fluctuating cognition, seizures or falls, can be challenging in clinic and RMTs, including those that can remotely measure HRV, offer a “real-time” window into the mental and physical health and functional status of the patient. This can also provide relevant insights into treatment responses to therapeutic interventions, whilst negating potential “white coat syndrome,” offering a more realistic and contextualized environment for data-collection and assessment. Moreover, remotely collected data offers the opportunity for greater confidentiality than a physical trip to a hospital, while removing the need for frail patients or carers to commute. RMT-based HRV therefore may have value as an adjunct digital biomarker in health and in neurovisceral digital phenotypes , adding Wearables exist that provide long-term telemonitoring of HRV using low-power biosensors that employ methodologies to acquire ECG signals from on-body sensors . Althougp = 0.005) increase in LF/HF ratio during cognitive loading compared to resting baseline  are key requirements, then wearable sensors are preferable. If HRV is a covariate or secondary measure, then the convenience of camera-based PPG may be more suitable. The RMT selection process is challenging and technical experts should always be consulted, due to the speed with which technology is updated and the wealth of available options . RMTs caData privacy and security remain substantial concerns for users, developers, clinicians, researchers, and regulators . AlthougFor widespread implementation, RMTs must be deployed to measure relevant, and sensitive variables. The wide variety of RMTs in the marketplace, makes selection challenging, particularly as manufacturers continually update their products, offering further challenges for planned deployment in existing healthcare systems. A further potential complication with many consumer devices is that they only provide aggregated rather than high-resolution raw data, complicating cross-device analysis and statistical analysis. Previous studies have indicated HRV could can provide some additional clinically relevant insight into health status (see What role can remotely captured heart rate variability play in telemedicine?), the advent of RMTs that capture indices of HRV offers the prospect of collecting relevant real-time data for clinical purposes. could therefore provide. This will require exploring the feasibility of deployment of RMT-based HRV as a meaningful clinical tool that enhances traditional methods and other digital biomarkers via robust piloting to standardize and define the most relevant temporal and spectral indices of HRV for the particular cohort and how artifacts or missing data can be mitigated.Many RMTs measure HRV, even consumer-grade wearables. HRV offers insights into neurovisceral processes in health and disorders of brain, body and behavior but has yet to be fully potentiated in the digital age. The use of RMTs to capture HRV and other CNS and ANS parameters can provide more detailed data across different contexts, such as activities of daily living or interventions and behavioral tasks. RMT-based HRV therefore has potential value as an adjunct digital biomarker in that has the potential to add continuously updated, objective and relevant data to typical clinical methodologies.The author confirms being the sole contributor of this work and has approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.