Changes in plasma membrane curvature and intracellular ionic strength are two key features of cell volume perturbations. In this hypothesis we present a model of the responsible molecular apparatus which is assembled of two molecular motors [non-muscle myosin II (NMMII) and protrusive actin polymerization], a spring [a complex between the plasma membrane (PM) and the submembrane actin-based cytoskeleton (smACSK) which behaves like a viscoelastic solid] and the associated signaling proteins. We hypothesize that this apparatus senses changes in both the plasma membrane curvature and the ionic strength and in turn activates signaling pathways responsible for regulatory volume increase (RVI) and regulatory volume decrease (RVD). During cell volume changes hydrostatic pressure (HP) changes drive alterations in the cell membrane curvature. HP difference has opposite directions in swelling versus shrinkage, thus allowing distinction between them. By analogy with actomyosin contractility that appears to sense stiffness of the extracellular matrix we propose that NMMII and actin polymerization can actively probe the transmembrane gradient in HP. Furthermore, NMMII and protein-protein interactions in the actin cortex are sensitive to ionic strength. Emerging data on direct binding to and regulating activities of transmembrane mechanosensors by NMMII and actin cortex provide routes for signal transduction from transmembrane mechanosensors to cell volume regulatory mechanisms. Cell swelling leads to regulatory volume decrease (RVD) mediated by activation of K+ and Cl− ion channels, K+-Cl− cotransporters (KCC) and the efflux of organic osmolytes , phosphatidylinositol 3,4-bisphosphate [PtdInsP2], phosphatidylinositol 3,4,5-trisphosphate [PtdInsP3], phosphatidylethanolamine, and is anchored to cell cortex by phosphatidylinositol transfer protein Vibrator [C-terminal fragments of heavy chains of NMMII bind directly to phosphatidylserine (PS) lipid vesicles and liposomes ,104,105 Vibrator . This sq+/K+-ATPase α1 subunit and play a role in its transport and trafficking both in neuronal and non-neuronal cells [2+ channel and is involved in its localization to membrane [2+ homeostasis regulates NMMIIA activity that regulates platelet generation from megakaryocytes [Ion channels are pore-forming membrane proteins located in cellular membranes and allow the passage of ions through the channel pore . NMMII sal cells . Furthermembrane . Alternaryocytes . Direct ryocytes ,99.Integrins are heterodimeric adhesion molecules consisting of an α and a β subunit and link the intracellular signaling components and actin cytoskeleton to the extracellular matrix. They are structural proteins playing an important role in mechanosensory transduction and they mediate both inside-out and outside-in signaling . IntegriSeveral lines of evidence suggest that, at least in hepatocytes, they act as volume sensors and are clustered in response to cell volume changes ,115,116.Interaction between integrins and NMMII is less studied field. The Stouffer lab first reported accumulation of NMMII heavy chains at alpha v beta 3 integrin complexes in response to vascular injury in smooth muscle cells . Later R1A receptor [It is well documented that activation of various GPCRs can influence the activity of NMMII via activation of downstream kinases and phosphatases and thus may play a role in cell-volume regulation . Howeverreceptor .There is the surprising observation that epidermal growth factor receptor (EGFR) can be activated by both swelling and shrinkage . DownstrTyrosine-phosphorylated EGFRs and platelet-derived growth factor receptors (PDGFRs) can directly interact with growth factor receptor bound protein 2 (Grb2) which transmits the signal further onto the small GTPase Ras . In NIH3ACE is a membrane protease expressed in several tissues and an important component of the renin-angiotensin system (RAS). It converts angiotensin (Ang) I to Ang II which regulates blood pressure through its effects on the kidneys, brain, heart and blood vessels. In addition to the cleavage of Ang I to Ang II, ACE has been reported to interact directly with NMMII and participate in outside-in signaling in endothelial cells . Based oExperimental testing of our model requires the real-time live-cell observation of the cascade of the following signaling and executive events.Visualization of activation of cell surface receptors—that is observation of signaling that fine-tune the NMMII and smACSK;Visualization of signaling proteins that transmit signal from cell surface receptors onto NMMII and smACSK;Observation of activation of NMMII and re-arrangements of the smACSK;Recording of pulling force and pushing force exerted by NMMII and actin protrusions, respectively;Visualization of activation of transmembrane signaling proteins by NMMII-driven pulling force and actin protrusion-driven pushing force;Observation of activation of signaling proteins that transduce signal from the transmembrane signaling proteins onto ion transport systems;Observation of activation of ion transport systems responsible for RVI and RVD.In our model cell surface receptors are to be activated twice, when they adjust cell volume point to execution of particular cell program, and when they are activated by mechanical stresses generated by cell swelling, shrinkage and NMMII and protrusive F-actin. Thus, signaling from EGFR is suggested to tune volume sensing machinery see  and EGFRProtein tyrosine kinases of the Src family, presented by nine members , participate in both RVI and RVD . Real-tiin versus HPout) across the PM by two active ATP-consuming mechanisms (NMMII and F-actin polymerization), (3) perfect elasticity of the PM-SMACSK complex, allowing its return to normal shape, and (4) sensing of changes in intracellular ionic strength by NMMII and machinery performing F-actin polymerization. Direct associations of NMMII and F-actin with several transmembrane signaling molecules provide the route for signal transduction onto ion transporting systems.Tight connections between cell volume and cell fate highlight the importance of the search for the molecular apparatus that performs cell volume sensing and regulation. NMMII and F-actin assembly acting together with a PM—smACSK spring seem a promising candidate volume sensor to us. We present an hypothesis that comprises four key features, (1) fine-tuning of cell volume set point  by cell fate, (2) probing of changes in difference in hydrostatic pressures (HPThe search for any particular volume-sensitive—or, as we assume here, active, energy-consuming cell volume sensing—molecular apparatus would be of importance for, e.g., anti-cancer treatment. Uncontrollable cell proliferation, resistance to apoptosis, migration—these features are among the hallmarks of cancer cell biology . These c


The incidence of renal cell carcinoma (RCC) is rising and metastatic RCC carries a very poor prognosis. The treatment paradigm for metastatic RCC has shifted dramatically in the last decade with multi-targeted tyrosine kinase inhibitors (TKI) previously used as first-line treatment but its utility is limited by short-lived efficacy and rapid disease progression. The dysregulation of immune cells in the tumour microenvironment contributes to unregulated growth of RCC. Thus, the use of immune checkpoint inhibitors has become first-line treatment for metastatic RCC and has offered dramatic improvement in clinical benefit and survival. Treatment with immune checkpoint inhibitor in combination with TKI appears to be promising in offering even greater response rates. The treatment for metastatic RCC continues to evolve and ongoing advances with new targeted agents and biomarkers are needed to continue to improve prognosis in the future. Renal cell carcinoma (RCC) has an incidence of approximately 400,000 cases per year globally, which is highest in North America, Europe and Australia . The incThe prognosis of RCC is poor as 30% of patients have metastatic disease at diagnosis with a 5-year survival rate of only 12% . PrognosThe advent of targeted treatment such as tyrosine kinase inhibitors improved survival outcomes for patients with metastatic RCC in the last decade . HoweverThe development of RCC is underpinned by abnormal angiogenesis. The von Hippel-Lindau (VHL) gene is a tumour suppressor gene that regulates activity of hypoxia-induced factor (HIF) and expression of vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) . VHL is RCC is highly immunogenic and contributes to mobilisation of immune cells such as Tumour Infiltrating Lymphocytes (TILs) and natural killer cells into the tumour microenvironment, which promotes tumour growth , 12. FurThe use of immune checkpoint inhibitors nivolumab and ipilimumab is now approved for first-line treatment of intermediate and poor-risk metastatic RCC and has demonstrated improved overall survival across multiple clinical trials . Nivolumvs. 27%, p<0.001), progression-free survival and a significant increase in 12-month overall survival rate , when compared to the control arm of sunitinib in those with intermediate or poor IMDC risk  and a significantly higher progression-free survival  in comparison to the control arm of sunitinib. This benefit was observed regardless of PD-L1 level and IMDC risk. However, avelumab and axitinib did not offer a significant overall survival benefit compared to the control arm in an updated analysis in 2020 , progression-free survival  and overall survival  in comparison to sunitinib. This benefit was observed regardless of PD-L1 expression or IMDC risk. There were no unexpected treatment toxicities but there was a higher incidence of hepatotoxicity and rates of treatment discontinuation in the intervention arm. Hypertension and diarrhoea were common toxicities in both groups. Treatment with pembrolizumab and axitinib appears to offer durable anti-tumour response at long-term follow up with an objective response rate of 85%, progression-free survival and overall survival rates of 94.7% and 74.8% at 36 months respectively  and significant improvement in progression-free survival  compared to sunitinib but this did not translate into an overall survival benefit , longer progression-free survival  and 12-month overall survival  compared to sunitinib. This benefit was observed across all subgroups including the group with ccRCC with sarcomatoid differentiation. Survival benefit was observed independent of PD-L1 expression and IMDC risk. No unexpected treatment-related adverse events were identified although rates of hepatotoxicity were higher in the intervention group. 19% of patients in the intervention arm required high dose corticosteroid treatment due to immune-related toxicities. However, patient-reported quality of life was greater in the intervention arm compared to sunitinib. First-line treatment with nivolumab and cabozantinib for metastatic RCC was most recently FDA-approved in January 2021 based on results from this trial. However, follow-up duration in this trial is reasonably short at 18 months and therefore durability of treatment response will need to be assessed at long-term follow up.The phase III trial, Checkmate-9ER included patients with ccRCC with sarcomatoid differentiation, which constituted 11.5% of the study population and investigated the role of nivolumab and cabozantinib, a second-generation anti-VEGF agent, in treatment-naïve advanced ccRCC . Resultsvs. 53.5% vs. 36.1%) compared to lenvatinib with everolimus and sunitinib respectively. Lenvatinib and pembrolizumab offered significantly higher progression-free survival  and higher overall survival  when compared to sunitinib. This benefit was observed regardless of PD-L1 level or IMDC risk. Lenvatinib and everolimus also offered longer progression-free survival  compared to sunitinib, but this did not translate into an overall survival benefit. However, toxicity appears to be an issue in this treatment and commonly included hypertension, diarrhoea, elevated lipase and hypertriglyceridaemia. 68.8% of patients in the lenvatinib and pembrolizumab group required dose reduction of lenvatinib and 37.2% of patients discontinued treatment as a result of toxicities. The FDA approved the use of lenvatinib and pembrolizumab in 2021 for treatment of metastatic RCC in 2021. Nonetheless, longer follow up data is required to continue to assess the efficacy and durability of response in this treatment.The phase III trial, CLEAR has recently been completed and investigated the efficacy of anti-VEGFR TKI lenvatinib either in combination with everolimus alone or combined with both everolimus and pembrolizumab in treatment-naïve ccRCC . This trThe advent of immune checkpoint inhibitors has led to a plethora of new treatment options for metastatic RCC. The approach of combining immune checkpoint inhibitor with a TKI as first-line treatment of metastatic RCC appears promising, yielding higher response rates and improved survival outcomes, demonstrated across multiple phase III trials , 27–30. There is no head-to-head trial evidence to compare the efficacy of the various treatment options available including immune checkpoint inhibitors, anti-VEGF therapy or a combination of both. There are multiple factors to consider when selecting first-line treatment for metastatic RCC. The IMDC prognostic risk model remains important in guiding selection of treatment. In favourable-risk disease, first-line treatment options include an anti-angiogenic agent alone or in combination with an immune checkpoint inhibitor, the latter option is favoured as illustrated in both NCCN and EAU guidelines in 2021. In favourable-risk disease, treatment with immune checkpoint inhibitor with TKI offers higher response rate and improved survival outcomes, when compared to treatment with sunitinib alone. In intermediate or poor risk disease, treatment options include ipilimumab and nivolumab or combining an immune checkpoint inhibitor with a TKI. Treatment with immune checkpoint inhibitor and TKI may be favoured in patients who are highly symptomatic with high disease burden and a rapid treatment response is desired, which may be offered by the TKI component of this treatment. Durability of treatment response should also be considered as there is now long-term follow up data to demonstrate the durable response and survival benefits offered by treatment with ipilimumab and nivolumab. In contrast, most clinical trials investigating various treatment regimens with immune checkpoint inhibitor and TKI have shorter follow up and immature long-term data, therefore it is unclear whether this treatment also offers similar durable responses when compared to ipilimumab and nivolumab. Toxicity is also an important consideration given higher rates of immune-related toxicities and requirement for high dose corticosteroid treatment associated with ipilimumab and nivolumab treatment compared to treatment with immune checkpoint inhibitor and TKI.There are now multiple FDA-approved immune checkpoint inhibitor and anti-VEGF treatment regimens available, which further complicates the decision-making process in selecting treatment for patients with treatment-naïve RCC  accumulates abnormally in VHL inactivation, which results in tumour growth and progressive clear-cell RCC . BelzutiIndoleamine 2,3-dioxygenase 1 (IDO1) mediates anergy of effector T cells and contributes to the immunosuppressive tumour microenvironment . TherefoThe identification of new predictive biomarkers and treatment targets is important to continue to improve the treatment of metastatic RCC. It has been demonstrated in many pivotal phase III clinical trials that PD-L1 expression is not a predictive biomarker as patients with negative PD-L1 expression also benefit from immune checkpoint inhibitor treatment. This is likely due to variable PD-L1 expression across different metastatic sites . HoweverNeutrophil lymphocyte ratio (NLR) is a measure of inflammation secondary to tumour growth and is the ratio of absolute neutrophil count to absolute lymphocyte count, which has been postulated as a potential biomarker that predicts treatment response to immune checkpoint inhibitor , 42. NLRPBRM1 is a possible biomarker, which is a gene that plays a role in remodelling of chromatin , 44. PBRnccRCC is a diverse group of RCCs with various histological subtypes that are vastly different but treated as one group due to rarity of the individual subtypes . The mosnccRCC often have positive PD-L1 expression, which is associated with more advanced disease and poorer prognosis, similar to in ccRCC , 49. TheThe treatment landscape of metastatic RCC has evolved in the last decade with the rise of immune checkpoint inhibitors in addition to the development of novel TKIs. This has resulted in the improvement of prognosis and survival for patients with metastatic RCC. However, given the rising incidence of metastatic RCC and the lack of evidence to guide treatment of nccRCC, there is a strong need for ongoing research in identification of new biomarkers and development of novel targeted agents to overcome persistent challenges posed by tumour resistance and to guide treatment decisions.IT contributed to the writing and completion of this manuscript. AS contributed to the conception of the research topic and the supervision of this project. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Invasive bacterial infections (IBI) in children present a difficult clinical challenge. They are often life-threatening, however in the early stages they can be hard to differentiate from benign viral infections. This leaves clinicians with the risk of missing a serious IBI diagnosis or inappropriately using antimicrobials in a child with a viral infection- contributing to the ongoing development of increased antimicrobial resistance. Hence, biomarkers which could aid in early detection of IBI and differentiation from viral infections are desirable. Mid-Regional pro-Adrenomedullin (MR-proADM) is a biomarker which has been associated with IBI. The aim of this systematic review was to determine its diagnostic accuracy in identifying children with IBI.A strategy was devised to search online databases MEDLINE, Embase, Web of Science and Scopus for human clinical trials reporting the accuracy of MR-proADM in children. Against predesigned inclusion and exclusion criteria full texts were selected for inclusion and data extraction. True positives, false positives, true negatives and false negatives were extracted from each included study to fill 2 × 2 tables. Using the Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool methodological quality of each study was assessed.A total of 501 articles were initially identified. After the removal of duplicates and abstract screening 11 texts were fully reviewed and four texts  were included in the systematic analysis. Only one study was of a high quality and that study accounted for the vast majority of patients. A single study reported the diagnostic accuracy of MR-proADM for invasive bacterial infection reporting an Area under the Curve of 0.69. The paucity of available studies made meta-analysis and studies of heterogeneity impossible.There is a paucity of research regarding the diagnostic accuracy of MR-proADM in the diagnosis of invasive bacterial infections in children. Initial results would suggest that MR-proADM testing alone is poor at identifying IBI in young children. It remains unclear if MR-proADM performs differently in older children or in children with signs and symptoms of IBI.CRD42018096295.PROSPERO The online version contains supplementary material available at 10.1186/s12887-022-03255-9. Invasive bacterial infections (IBI) are defined as the identification of pathogenic bacteria from a sterile fluid or body tissue by either culture or molecular diagnostic techniques such polymerase chain reaction (PCR). Worldwide IBIs are the leading cause of neonatal and paediatric morbidity and mortality . As manyBiomarkers for severe infections or IBI are often measured from plasma samples taken at i) presentation to hospital to initiate treatment ii) as an inpatient to guide treatment and iii) to prognosticate on expected outcomes including the need for Intensive Care Unit (ICU) admission. In current clinical practice C-Reactive Protein (CRP) and Procalcitonin (PCT) are the most widely used biomarkers to used detect IBI and monitor response to treatment . UnfortuThe index test for this systematic review is Mid-Regional pro-Adrenomedullin (MR-proADM). MR-proADM is a peptide that is related to adrenomedullin (ADM). ADM was first isolated and detected in 1993 and has been found to be involved in a wide range of human physiological processes . ADM actAlthough ADM has near ideal characteristics as a biomarker of IBI its use in clinical practice has been limited. This is due to the difficulty in reliably measuring ADM levels in-vivo . ADM is MR-proADM, a 48 amino acid fragment that is produced in conjunction with ADM at a ratio 1:1 is a significantly more stable molecule making it easier to measure in-vivo . There iResearch assessing MR-proADM as a predictive diagnostic biomarker has been increasing in recent years with researchers investigating its role in paediatrics. A recent systematic narrative review suggested that MR-proADM may have a diagnostic role in the diagnosis of serious bacterial infections (non-IBI) in children as well as in non-infective inflammatory conditions such as juvenile arthritis . HoweverThe aim of this review was to establish the diagnostic accuracy of MR-proADM in the detection of IBI in children under the age of 18. The secondary aim was to determine the diagnostic accuracy of MR-proADM in various age-related subgroups such as newborns, neonates, children and adolescents and whether any differences in accuracy or optimal cutoff values existed between these groups.Prior to conducting this systematic review a protocol was produced in adherence to the standards of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) and registered prospectively on the 30/05/2018 with the International Prospective Register of Systematic Reviews (PROSPERO) - registration number CRD42018096295. The protocol underwent external peer review and was published in 2020 . These characteristics are summarised in Table Four studies including 1404 patients aged between day one of life and 12 years were included in the final systematic review –23. One tandards –22. Thesn = 1077) provided data directly assessing the diagnostic test accuracy of MR-proADM for the identification of invasive bacterial infection in children [Only the study by Benito et al. (children . They prchildren .n = 128) and Hagag et al. (n = 60) were both performed in newborn populations [r = 0.67 p < 0.05) [The studies by Oncel et al. . They also reported that the MR-proADM level was positively correlated with severity of sepsis (r = 0.62 p < 0.05) [The study by Lan et al. reported that MR-proADM levels were significantly higher in children with paediatric sepsis in the intensive care unit compared to healthy controls ( < 0.05) .The paucity of available studies made meta-analysis and studies of heterogeneity impossible.n = 1035) [The systematic review was primarily designed to determine the accuracy of MR-proADM at identifying invasive bacterial infection in children less than 18 years of age. The review included four studies of 1404 patients –22. Only = 1035) . The rem = 1035) . This wo = 1035) .Many potential novel biomarkers have been studied in children at risk of IBI, however most do not demonstrate better diagnostic accuracy than the traditionally used biomarkers of CRP and PCT . Some noIncreasingly there is an argument that individual biomarkers may have limited diagnostic utility in detecting IBI. Some novel biomarkers have been included in clinical scoring systems in combination with physical findings to improve diagnostic accuracy . Whilst n = 16). The available studies suggest that MR-proADM may have a role in identifying and stratifying sepsis in children, but further studies are required to understand the clinical utility of the test.The numbers of studies reporting on the test accuracy of MR-proADM for the diagnosis of invasive bacterial infections in children are small and there is only one high quality paediatric study with a small number of cases of IBI (There is a paucity of research regarding the diagnostic accuracy of MR-proADM in the diagnosis of invasive bacterial infections in children. Initial findings indicate that MR-proADM is not a good biomarker for the diagnosis of IBI although further research is required. Specifically, research is required to investigate if MR-proADM has a role in predicting the need for ICU admission in children admitted to hospital with severe infection.Additional file 1.


Immunotherapy in the metastatic setting has drastically altered the landscape of treatment for various types of malignancy, including colorectal cancer. The category of immune checkpoint inhibitors has especially emerged as a class of therapy predicated on a more comprehensive understanding of immune cell-cancer cell regulation and evolution of the tumor microenvironment over time. Strategies including adoptive cellular therapies, tumor vaccines, and antibodies have also demonstrated the ability to enhance antitumor immunity. In this article, we provide a comprehensive review of the current landscape of immunotherapeutic strategies in colorectal cancer and provide insight into how these strategies may evolve in the next decade and be adapted to more localized forms of cancers of the colon and rectum. We provide particular focus on various combination approaches under investigation for reversing cancer-induced immunosuppression, especially in mismatch repair-proficient/microsatellite-stable colorectal tumors. Finally, we summarize current understanding on a recently identified integral factor in local immune regulation, the colonic microbiome. The aim of this article is to identify current challenges and barriers to improvement and to specify opportunities for applying knowledge in the immunotherapy sphere to rational design of clinical trials intended to improve survival and related outcomes in patients treated in the neoadjuvant setting.  Evolving insight in tumor immunology and the colonic microbiome will allow expansion of current immunotherapeutic strategies to early stages or neoadjuvant settings and to overcome intrinsic biologic resistance, which can improve survival and related outcomes in patients with colorectal cancer. Implementation of immunotherapy has long been a desired goal in the field of cancer research due to its promise for tailoring treatments to individual patients while also promising potential for inducing a longer-term form of immune surveillance that theoretically could decrease risks of future recurrence of disease. The reality of biologic discovery and implementation of immunotherapy has been a longer process than anticipated but has certainly entered more mainstream acceptance over the past decade with randomized controlled trials (RCTs) demonstrating success in the form of longer survival for patients with metastatic melanoma and non-small cell lung cancer, most prominently. However, the extent of that success has not immediately transferred as well to many other forms of systemic malignancies, including cancers of the gastrointestinal tract, and CRC quite specifically. Thus far, evidence supporting the use of immune checkpoint inhibitors (ICIs) is most abundant in cases of metastatic treatment-refractory cases of gastric cancer and hepatocellular carcinoma;Development of CRC involves complex processes accumulating genetic mutations, resulting in the heterogeneity of response to treatment . The use,Even further, as investigations in this field to date have focused on patients with metastatic drug-refractory forms of CRC, findings thus far demonstrating exceptional response rates in some subsets of tumors and patients point to a fertile opportunity to also reexamine regimens currently being used for the neoadjuvant setting for resectable and locally advanced forms of rectal carcinomas, as these patients reside within a window of opportunity to succeed in curative intent treatment due to a relatively limited extent of measurable disease. Furthermore, as a majority of cases of recurrent disease will manifest with distant metastatic disease, consideration of upfront immunotherapy offers an additional theoretical benefit by adding an immune-surveillance component that would further reduce recurrence risk in the long term and also potentially lowering risk of short- and long-term adverse side effects compared to cytotoxic chemotherapies in certain patient populations. To get to that point, there are major challenges and unanswered questions that will need to be addressed in well-designed human trials. The many variables to consider include and are not limited to variability of the host immune system between individual patients and patient populations, co-morbid conditions, and varying immunological milieu within tumor microenvironments (TMEs) that will result in variation of responses and require further investigation. In recent years, a spotlight has been placed on the role of the gut microbiome on an individual host’s response to immunotherapy, with ongoing research underscoring this previously unknown wild card.Understanding the current context of treatment of patients with mCRC, which has been in place for nearly two decades, helps to provide a foundation of understanding for how translation can occur to the neoadjuvant setting. There are multiple combination regimens available as part of current standards of care for mCRC. These generally include a 5-fluorouracil (5-FU) backbone-based regimen in combination with either the platinum compound oxaliplatin (FOLFOX) or topoisomerase I inhibitor irinotecan (FOLFIRI) or all three (FOLFOXIRI); either of these chemotherapy doublet or triplet regimens may be combined in some patients with biologic agents targeting vascular endothelial growth factor (VEGF) or epidermal growth factor receptor (EGFR), the latter of which has biologic activity only in tumors harboring wild type but not mutant isoforms of RAS.Various strategies in activating immunity against cancer have been explored in CRC . The mec,The emergence of TME resistance to immune surveillance is mainly induced through several immune checkpoint signaling cell surface receptors, most prominently programmed death-1 (PD-1), PD-L1, and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4).,Initial studies of ICIs in CRC demonstrated very limited activity in unselected CRC populations.NCT02997228) or to FOLFOXIRI plus bevacizumab  in mCRC. In the second-line treatment setting, pembrolizumab showed a 33% ORR; a trial comparing the effect of anti PD-L1 antibody, avelumab, to standard chemotherapy is underway .There are also trials investigating the effects of the addition of atezolizumab to FOLFOX plus bevacizumab  with another phase II study planned to study the efficacy of pembrolizumab in early-stage colon cancer . The combination of ICIs to neoadjuvant therapy in CRC is also under investigation based on the hypothesis that chemoradiotherapy may increase the neoantigen formation and increase the efficacy of ICIs .Studies in early-stage melanoma showed that neoadjuvant ipilimumab + nivolumab expands more tumor-resident T cell clones than adjuvant application with a pathological response; this finding prompts consideration of approach and implications in the setting of locally advanced CRC as well.NCT03552718, NCT03639714, NCT03953235, and NCT04147078). Various methods of antigen delivery have been investigated including molecular-based vaccines , cell-based vaccines (tumor cells and dendritic cells [DCs]), and vector-based vaccines . These have shown mixed results with limited efficacy in improving clinical outcomes.Therapeutic cancer vaccines have been applied in various cancer types to enhance immune response and immunological memory to prevent recurrence.in vivo following infusion into patients. Currently, there are two main classes of ACT in the process of development for clinical use, as follows.ACT is a novel therapeutic strategy that has made great strides over the past several decades. The strategy allows selection, activation, and expansion of effector immune cells (EIC) NCT01420874, NCT02487992, NCT03904537, and NCT04282044). The strategy that combines ACT with conventional chemotherapy treatment has also shown promising results .TILs are lymphocytes that infiltrate within tumors or at the tumor margin. The presence of TILs is associated with better prognosis due to selective antitumor activity.NCT04426669). This combined phase I/II trial will determine both safety as well as efficacy of this new approach to ACT; if successful, it is conceivable that this approach in the neoadjuvant setting could be utilized to induce significant tumor regression for an overall intent-to-cure approach combined with surgical resection and possibly lead to higher rates of clinical CR using current modalities. Furthermore, the concept of immune surveillance using this ACT method has intriguing opportunities for also inducing a longer-term form of specific anti-tumor immunity in any given individual patient, introducing an interesting possibility of simultaneously decreasing the risk of short- and long-term recurrence beyond the current capabilities of cytotoxic chemotherapy given in the adjuvant setting.Advances in deriving TILs from solid tumors, including from CRC, in combination with improvements in gene-editing techniques including the CRISPR-Cas9 system, have collectively opened the door toward enhancing potential for this therapeutic strategy in treatment of patients with CRC. Novel checkpoint targets have emerged beyond PD-L1 for other gastrointestinal cancers  and MSI for CRC and others. Evolution and improvement of gene-editing techniques that have higher efficiency as well as decreased off-target editing rates, in combination with such novel immune checkpoints, represent a new frontier for therapeutics that, if successful in the metastatic setting, may provide a foundation for use in the neoadjuvant setting in the not-too-distant future. One such example is already in clinical trial at our institution and comprises CRISPR-Cas9 inhibition of the intracellular immune checkpoint CISH. Inhibition of CISH has been achieved at >90% success rates, and the preclinical research at the University of Minnesota in collaboration with researchers at the National Cancer Institute led to a current clinical trial, opened in May 2020, enrolling patients with metastatic gastrointestinal cancers refractory to at least one line of treatment .Carcinoembryonic antigen (CEA) is an attractive target for immunotherapy in CRC as the molecule is frequently overexpressed in over one-half of patients with CRC. One study recruited three patients and demonstrated a significant decrease in serum CEA level, with one patient having an objective response in metastatic lesions following T cell treatment.NCT03310008, NCT03370198, NCT03692429, NCT04107142, and NCT04550663). Other potential targets undergoing clinical studies include EGFR (HER-2) , mesothelin , guanylyl cyclase C (GUCY2C), and mucin 1 (MUC1), with most of the studies still in an animal model or early phase of clinical trials.NCT03970382). Major challenges in the application of CAR cells in solid tumors include physical barriers surrounding cancer cells leading to insufficient infiltration, immunosuppressive TME, and barriers on logistics and cost of the treatment.Natural killer group 2D ligands (NKG2DLs) are gaining interest, as they are commonly overexpressed in cancer and can be a target for NKG2D receptor-based CAR-T cellsNCT02324257) or combination therapy .BiTEs are artificially produced antibodies containing two distinct single-chain variable regions. The mechanism is through binding of the target antigen by one antibody fragment and engaging the T cell by another fragment specific to CD3. This action increases T cell infiltration and generates a highly inflamed TME leading to cancer cell apoptosis.Virotherapy for cancer treatment is another evolving field that harnesses unique biologic characteristics of the viruses. Oncolytic viruses are designed to preferentially infect and replicate in the cancer cells, causing cell lysis, and induce host anti-tumor immune responses.NCT01274624) and recombinant vaccinia virus in combination with irinotecan  are also under evaluation. Pre-clinical studies have also provided promising results for oncolytic virus combination with ICIs and were recently translated into clinical trials in CRC,NCT03206073 and NCT04348916). A combination therapy of binary oncolytic adenovirus in combination with HER2-specific autologous CAR-T cells is currently under investigation as well .Methods have been employed to increase efficacy and safety for this therapy option. Combination of pelareorep (oncolytic reovirus) with FOLFOX/bevacizumab was tolerable with an increased ORR, but PFS was inferior.Effectiveness of ICI treatment is limited to the dMMR-MSI-H subset, which represents around 15% of localized CRC and about 4% of mCRC.,The differences in TME composition and immune cell distributions between dMMR-MSI-H and pMMR-MSI-L tumors contribute to response rate to therapy and prognosis.,NCT02291289) and BACCI trial  revealed minimal effects on PFS and OS.Immunogenic cell death induced by chemotherapy leads to antigen release and immune activation, which could sensitize tumor cells to ICI therapy.RT has long been postulated to activate antitumor immunity at distant sites through what has been termed “the abscopal effect.” A study in melanoma models also showed that this combination can reverse treatment resistance due to upregulation of PD-L1 on cancer cells.in vitro study revealed a subgroup of patients who carry certain Fc gamma receptor (FcγR) polymorphisms with higher antibody-dependent cell cytotoxicity (ADCC) activity has significantly longer PFS.Monoclonal antibodies targeting EGFR (cetuximab or panitumumab) are a standard of care for treatment of wild-type RAS mCRC. A previous + T cells from baseline was significantly associated with a better outcome.Regorafenib, a multi-kinase inhibitor, possesses pleiotropic effects in inhibiting angiogenesis and immunomodulation, mainly through TAM inhibition.,Inhibition of mitogen/extracellular signal regulated kinase (MEK), a downstream effector of the RAS-MAPK pathway, was found to result in upregulation of PD-L1 and have synergistic effects with the PD-1 inhibitor in preclinical models.,Currently, there are extensive ongoing trials exploring knowledge in the field to improve the efficacy of immunotherapy.Fusobacterium nucleatum correlated with CD3+ TIL depletion,,,The microbiome has been well described in CRC carcinogenesis,Bifidobacterium abundance.Bifidobacterium-containing pro-biotics were all found to be able to transfer this effect. Evaluation of the immune environment revealed that enhanced tumor-specific CD8+ T cell activity and DC maturation in correlation with Bifidobacterium abundance were the keys to this response, reflecting earlier findings on the microbiome and carcinogenesis.Given the limitations in CRC-directed immunotherapy, early studies focused on the interaction between the microbiome and anti-PD-1/CTLA-4 therapies in metastatic melanoma.Bacteroides taxa and abrogated by antibiotic induced dysbiosis.Bacteroides thetaiotaomicron, Bacteroides fragilis, and Burkholderia capacia was able to restore anti-tumor activity of anti-CTLA-4 antibodies in germ-free mice. Further, these taxa were noted to induce Th1 immune response and promote DC maturation. This particular study went further, showing that human-mouse fecal transfer of feces enriched for B. fragilis and B. thetaiotaomicron induced a marked response to anti-CTLA-4 therapy.Similarly, anti-CTLA-4 antibody efficacy against mouse-model sarcoma and melanoma was found to be promoted by the presence of specific Ruminocaccaceae faecalibacterium, and Akkermansia muciniphila.Faecalibacterium,Faecalibacterium was associated with significantly longer PFS/OS.A. muciniphila was positively associated with a treatment response in patients with multiple advanced cancers.Subsequent human studies identified additional taxa involved in immunotherapy treatment response, including + lymphocyte infiltration in clinical CRC samples. Specifically, the abundance of the families Lachnospiraceae and Ruminococcaceae was correlated with expression of CCR5 and CXCR3 chemokines, the latter of which is associated with tissue T cell trafficking.Ruminococcaceae family members, suggest that microbiome-based therapies may be useful in priming tumors for response to immunotherapies. Another landmark study demonstrated that a group of 11 low-abundance bacteria was necessary and sufficient to enhance anti-PD-1 antibody activity against a mouse xenograft colon cancer model.+ CD8 T cells, as depletion of this population abrogated this effect. Mice treated with the 11-strain mix also demonstrated lower frequency of colitis in response to anti-PD-1 therapy, suggesting that these bacteria could both enhance treatment efficacy and limit colon-specific side effects.Given these promising results, more recent studies have focused on identifying a subset of bacteria that may modify the tumor immune environment,NCT04264975 and NCT04130763), and their outcomes could significantly alter the prognosis and management of mCRC. Preliminary results are promising, with renewed response to anti-PD-1 therapy seen in previous immunotherapy-refractory metastatic melanoma.Several clinical trials are currently underway to evaluate these hypothesesThe advent of immunotherapy—in the active sense with drug intervention that stimulates the innate immune system to better recognize and counteract tumors through recognition of their antigens or cell surface markers—has changed the paradigm of treatment of solid tumors such as melanomas and non-small cell lung cancers and in the process of current investigation, challenging traditional notions that chemotherapy for CRC in general, and radiation for rectal cancers, needs to be the mainstay of treatment. Current progress most prominent with ICIs, and also CTLA-4 inhibitors and various forms of cellular therapy that are actively being investigated, is beginning to make an impact and will provide better knowledge for how these modalities may improve upon current strategies for intent-to-cure neoadjuvant therapies for these diseases. The current frontier includes many investigators aspiring to unlock and validate new markers beyond dMMR-MSI-H that will predict susceptibility to ICI and other immunotherapies. Understanding and differentiating biologic differences of these and similar populations in the context of molecular drivers, and their overall impact in different stages of carcinogenesis as well as in cases of CRC at different anatomic stages of progression, will be essential. Emerging knowledge of the effects of the microbiome, not just on CRC biology at all stages but also on modulating the efficacy of immune-based therapies, is another angle that will need attention in correlative studies to clinical trials as well as in design of the therapeutic approaches to care in all settings, including neoadjuvant approaches. Growing understanding in complex interactions between tumor cells and the immune system, analyzed in the context of the dynamic microbiome, all assessed using more sophisticated molecular and genetic techniques will merge our understanding and be a major basis for developing a better understanding of this process and will need to be taken into account for rational design of better and more strategically savvy therapeutic clinical trials.


Laura Spring, MD, a clinical investigator and breast medical oncologist at Massachusetts General Hospital Cancer Center and Harvard Medical School, shared insights on breast cancer research presented at the 2021 San Antonio Breast Cancer Symposium (SABCS). Laura Spring, MDThe Oncologist: Starting with human epidermal growth factor receptor 2 (HER2)-positive breast cancer, we saw updates from the DESTINY-Breast03 trial, which compared trastuzumab deruxtecan and trastuzumab emtansine in patients with HER2-positive metastatic breast cancer. What are your thoughts on this trial, particularly the findings in patients with brain metastases?Dr. Spring: The DESTINY-Breast03 subgroup results were great to see. In particular, the development of brain metastases is a major issue in HER2-positive metastatic breast cancer, and we need more therapeutic strategies. We had previously seen some data suggesting trastuzumab deruxtecan may have some intracranial activity. With this update from DESTINY-Breast03, now we see further results showing many patients with brain metastases did benefit from trastuzumab deruxtecan, further suggesting that this can be an option for patients with brain metastases as well.The Oncologist: Another trial in the HER2-positive metastatic setting was the phase III PHOEBE trial comparing pyrotinib plus capecitabine with lapatinib plus capecitabine after prior treatment with trastuzumab and taxanes. What are the key take-home messages from this trial?Dr. Spring: Pyrotinib outperformed lapatinib, but the PHOEBE trial occurred during a time of active drug development in the HER2-positive space, making it difficult to know how the agent will fit into the landscape. However, being an oral agent, it may be a good option in areas where access to certain anti-HER2 therapies is limited.The Oncologist: We also saw several trials in hormone receptor (HR)-positive, HER2-negative breast cancer. The phase III EMERALD trial evaluated elacestrant in patients with previously treated estrogen receptor (ER)-positive metastatic breast cancer. What are your thoughts on this trial?Dr. Spring: The EMERALD trial was the first phase III study of a novel oral selective estrogen receptor degrader (SERD) to demonstrate a statistically significant improvement in progression-free survival (PFS) versus standard of care. The results suggest elacestrant may become a new treatment option for patients with metastatic breast cancer. The results were more impressive in patients with ESR1 mutations. Both arms of the study had a relatively short PFS, suggesting more work is needed to address the issue of endocrine-resistant disease.Being first in class is always important and has potential to lead to an approval. Although elacestrant outperformed the treatment of physician’s choice (fulvestrant or an aromatase inhibitor), the outcomes overall were somewhat disappointing in either arm. This highlights that we still have a lot of work to do for this disease. Many of these patients were likely resistant to estrogen-blocking therapy—this is subset that needs a lot of research focus. Ultimately the novel oral SERDS may be even more helpful in the adjuvant breast cancer setting, and trials are ongoing and planned.The Oncologist: The phase III PADA-1 trial examined the strategy of switching from palbociclib plus an aromatase inhibitor to palbociclib plus fulvestrant upon detection of the ESR1 resistance mutation in patients with ER-positive metastatic breast cancer. What can we learn from this trial?Dr. Spring: PADA-1 was an interesting study that looked at the impact of ESR1 mutations detected via circulating tumor DNA (ctDNA). These are mutations that typically breast cancers don’t have right off the bat; they develop later as a resistance mechanism to certain types of endocrine therapy. This trial asked, if you identify ESR1 mutations earlier and change the endocrine therapy backbone from an aromatase inhibitor to fulvestrant, is that helpful? This was an interesting proof-of-principle study. I don’t think it will change the standard of care right away, but it will influence other studies and it will be important to explore this paradigm more.The Oncologist: Updates from the phase III MONALEESA trials, which evaluated ribociclib plus endocrine therapy in patients with HR-positive, HER2-negative advanced breast cancer, focused on treatment outcomes across subgroups. What are the take-home messages from these updates?Dr. Spring: The updates from MONALEESA continued to show a treatment benefit with ribociclib regardless of tumor subtype or site of metastatic disease. Overall survival benefit was consistent across the intrinsic subtypes , which is important because the intrinsic subtypes speak to disease biology. These different subtypes can behave quite differently, so it’s nice to see that ribociclib was still helpful regardless. In terms of the number and sites of metastases, which speaks to burden of disease, treatment with ribociclib was again helpful regardless of the situation.The Oncologist: Several updates from the phase II BYLieve trial also examined outcomes across patient subgroups—in this case, patients with HR-positive, HER2-negative, PIK3CA-mutated advanced breast cancer who progressed on or after prior treatment with a cyclin dependent kinase (CDK) 4/6 inhibitor. What does this research tell us about the ability to identify predictive biomarkers?Dr. Spring: The BYLieve trial is important because the study that led to the approval of alpelisib, SOLAR-1, included relatively few patients treated with a CDK 4/6 inhibitor, as the standard of care was evolving at that time. The recent data presented at SABCS 2021 helps confirm the efficacy of alpelisib, even in patients with a short duration of prior treatment with CDK 4/6 inhibitors or with ESR1 mutations. A short duration of response to CDK 4/6 inhibitors and the presence of an ESR1 mutation can both suggest endocrine resistance, and so it was nice to see that alpelisib plus fulvestrant showed efficacy in these settings. Like many targeted therapies, alpelisib has side effects; ultimately it would be great for patients if we could better predict who may benefit more, and who may benefit less, from a drug. There is also interest in biomarkers that may help predict side effects, to help with personalization. As we heard with the EMERALD study, we’re going to continue to have more agents available, and therefore personalization of therapy selection becomes even more important.The Oncologist: Yes, speaking of personalized treatment, we also saw an update from the phase III Southwest Oncology Group (SWOG) S1007 RxPonder trial, which evaluated the benefit of adjuvant chemotherapy based on clinical and genetic risk factors in patients with HR-positive, HER2-negative, early breast cancer. What have we learned about personalizing therapy in this setting?Dr. Spring: This update confirmed previous overall findings from the RxPonder trial, which has helped many patients avoid unnecessary treatment with chemotherapy. One challenge involves the premenopausal subset, which continues to show benefit from adjuvant therapy regardless of Oncotype DX recurrence score according to the study results. The most recent RxPonder update reviewed how many premenopausal patients received ovarian function suppression (OFS), and that number was relatively low (6.3% in the chemoendocrine arm and 19.0% in the endocrine-only arm). Therefore, it is still an unanswered question whether OFS could replace chemotherapy and provide the same benefit. The RxPonder trial just wasn’t the right trial to answer the question, but it was informative to see data showing that not many premenopausal patients received OFS.The Oncologist: Lastly, updates from the KEYNOTE-355 and KEYNOTE-522 trials examined the role of checkpoint inhibition with pembrolizumab in patients with triple-negative breast cancer (TNBC). What are your thoughts on these trials?Dr. Spring: Updated results from KEYNOTE-522 and KEYNOTE-355 continue to establish the role of pembrolizumab both in programmed death-ligand 1 (PDL-1)-positive metastatic TNBC and in stage II and III TNBC regardless of PDL-1 subtype. KEYNOTE-355 more specifically examined the PDL-1 cutoff that identifies patients who are most likely to benefit from pembrolizumab, whereas KEYNOTE-522 focused on the updated event-free survival data. The further follow-up data from both trials helped solidify the role of this agent becoming a standard of care in TNBC.


The consequences of strict COVID-19 mobility restrictions on motor/non-motor features in Parkinson’s disease (PD) have not been systematically studied but worse mobility and quality of life have been reported. To elucidate this question, 12 mild to moderate PD patients were assessed in March 2020 before and after two months of isolation as part of a clinical study that had to be interrupted due to the pandemic and the implementation of COVID19 mobility restrictions.Twelve patients were systematically evaluated before and after the lockdown period as part of a larger cohort that previously underwent thermal water rehabilitation. Clinical outcomes were the Body Mass index, the Mini-Balance Evaluation Systems Test, the MDS-Unified Parkinson’s Disease Rating Scale part III, the 6 Minute Walking Test and the New Freezing of Gait Questionnaire. Global cognition was evaluated with the Montreal Cognitive Assessment scale. The impact of COVID-19 restrictions on quality of life and functional independence was evaluated with The Parkinson’s disease Quality of life (PDQ-39), the Activities of Daily Living (ADL) and Instrumental Activities of Daily Living questionnaires (IADL) and the Parkinson’s disease cognitive functional rating scales (PD-CFRS).p=0.005), and four patients reported one or more falls during the lockdown. BMI increased (p=0.031) while the remaining clinical variables including quality of life did not change.After two months of isolation the Mini-BESTest score worsened (We observed moderate worsening at Mini-BESTest, greater risk of falls and increased body weight as consequence of prolonged immobility. We believe negative effects were partially softened since patients were in contact with our multidisciplinary team during the lockdown and had previously received training to respond to the needs of this emergency isolation. These findings highligh the importnace of patient-centered interventions in PD management. The COVID-19 pandemic has been declared as a global emergency by the World Health Organization. Unprecedented restrictions have been implemented to control the viral spread, including a complete lockdown of quarantine isolation in certain countries including Italy –5. FallsSince, physical exercise programs are essential components in the management of motor and non-motor symptoms in PD patients –13, the Twelve PD participants from our Movement Disorders and Parkinson Unit of the University Hospital of Padua  were tested before and after the 2-months lockdown period in Italy.All participants had at screening (max 1 week before the baseline visit): (i) diagnosis of idiopathic PD ; (ii) HoIn order to be sure that participants had no close contact with potential COVID-positive people or became COVID-positive themselves between the two evaluation sessions, a triage questionnaire was administered both by telephone and in person at post-isolation visit. Namely it was checked whether i) their body temperature was lower than 37.5 °C, ii) they experienced COVID-19 symptoms; iii) their family members or close contacts (less than 1 meter in the same room for at least 15 min without protective equipment) confirmed or suspected to have contracted COVID-19 in the 20 days prior the evaluation; iv) they have ever been tested for COVID-19, especially in the 20 days prior the evaluation; and eventually if v) they have ever been positive to COVID-19 tests. None of the patients included in this study had history of positive COVID-19 or had contact with positive people.Participants were evaluated in the context of a project that aimed to study the effect of rehabilitation in thermal water on people with PD. This rehabilitation program was interrupted due to the COVID-19 lockdown, but all clinical assessments at baseline were successfully performed the 2-weeks before lockdown. Participants were evaluated by the multidisciplinary team in our clinic, which includes a nurse case manager, neurologists, psychologists, physiotherapists and bioengineers working in collaboration with the rehabilitation unit and a study coordinator. Our approach to patients’ management includes education and training according to current recommendations , 18. AssThe study was carried out according to the Declaration of Helsinki and ethical approval to data collection was granted by the local review board. Participants read and signed an informed written consent form before participating to the study.Clinical description , and motor aspects   and Hoeh2) and carried out the motor evaluation. The Mini-Balance Evaluation Systems Test (Mini-BESTest), a shortened version of the Balance Evaluation Systems Test (BESTest), was employed as clinical tool to measure balance and predict the risk of falls [A physical therapist specialized in movement disorders collected the participants’ body mass and height to calculate the participants’ body mass index  was used to assess participants’ aerobic capacity and endurance. The distance covered over a time of six minutes is recorded as the outcome on which to contrast changes in performance and capacity. The test was carried out in an obstacle-free corridor with a length of 30 meters .The New Freezing of Gait Questionnaire (NFOG-Q) is a self-reportable questionnaire consisting of nine items that measure presence, severity and relevant impact of freezing of gait (FOG) in individuals’ daily life . The higA clinical semi-structured interview was carried out by two experienced neuropsychologists performing the cognitive-behavioral evaluation. The Montreal Cognitive Assessment scale (MoCA) was administered to evaluate global cognitive status as it has been shown that MoCA is a sensitive and clinically useful cognitive screening instrument in parkinsonisms –30; The Statistics were calculated using IBM SPSS Statistics .2 [For each variable, Minimal Detectable Change (MDC) derived from literature was considered to evaluate the noticeable change in ability : Mini-BE2 ), MoCA (2 ).dZ measure correcting for small sample size and between-repetitions Pearson correlation [The normality of scores distributions was assessed using the Shapiro-Wilk test. Pre-post lockdown effects on continuous variable were assessed with paired Two-tailed Student’s t-tests and on categorical variables with McNemar test. For those variables that do not satisfy normality distribution, Median, 25-75-percentile values and the paired Wilcoxon signed-rank test were considered. Effect size power analysis was calculated using Cohen’s relation . Achieverelation . An achiParticipants’ sociodemographic and clinical characteristics at the screening visit (pre-isolation period) are shown in Table Ten participants asked for telephone assistance during the lockdown period as they reported pain associated with poor daily mobility. The other two participants reported a good autonomy in self delivering stretching exercises that were performed on a daily basis.p = 0.031, dz = 0.703). Differences in MDS-UPDRS part III total score were not significant (p = 0.092). Six subjects showed an increasing of the MDS-UPDRS part III equal or superior to +4.63 points. Four of them remained stable and the other two showed a variation of -3.25 points. The Mini-BESTest total score was significantly reduced . Namely, an increased number of participants (six at the post-isolation vs. one at pre-isolation visit) obtained less than 17.5 points, the PD cutoff for potential fallers at the Mini-BESTest total score. In particular, four participants displayed a decline in the APAs sub-score; four participants worsened in the RPC, whereas similar values were observed in the SO and DG sub-scores in pre- and post-isolation visits. Post-hoc power analysis indicated that the power to detect obtained effects at the 0.2 level was 0.861 for the effect of lockdown on Mini-BESTest performance.Table p=0.799) and the NFOG-Q questionnaire (p=0.313) between pre- and post-isolation evaluations.Additionally, statistical analysis revealed no significant differences in the 6MWT (p=0.392), nor in the PD-CFRS total score (p=0.123). Qualitative analysis of the PD-CFRS sub-items showed that four patients reported language (anomia/comprehension) problems and inability to remember drug intakes. Overall, PDQ-39 score did not change (p=0.472) but qualitative analysis shows a worsening in the "emotional well-being" domain in two subjects while 10 was in a similar condition.No differences were found in the ADL and IADL questionnaires. No significant effect was found on global cognitive performance as assessed with MoCA scale (To the best of our knowledge, this is the first study assessing the consequences of the COVID-19 related restrictions on both motor and non-motor symptoms of people with PD who had never contracted COVID-19. Considering that our subjects had been assessed just before the implementation of strict lockdown, we had the unique opportunity to test the impact of isolation without any treatment change and through a systematic and complete evaluation.Our findings showed that no significant worsening in clinical measures and quality of life were detected for this cohort of PD patients in the mild and moderate stages of the disease. These findings should be interpreted considering that PD participants were followed regularly at our clinic and attended several educational activities at our local Parkinson lay association. We suggest that these results further point to the importance of multidisciplinary care interventions for PD patients to hand and learn self-management strategies , 18.The only worsening item we detected was the Mini-BESTest total score. This finding could indicate that, if the lockdown period had continued, we could have possibly observed a clinically significant deterioration in balance performance of these individuals. The Mini-BESTest has proven to be a significant predictor of falls and detector of balance problems in PD , 23, andMDS-UPDRS motor examination total score highlighted only a modest median worsening. However, considering the minimal threshold of +4.63 points for detecting clinically meaningful worsening changes in the score of the MDS-UPDRS part III, this variation was clinically relevant for six of these participants . The facFinally, it should be noticed that baseline clinical, cognitive and functional status was relatively mild for this cohort and PDQ-39 score confirmed that patients were stable, and symptoms were well controlled with pharmacological treatment. Emotional wellbeing was the only domain showing a trend for worsening most likely due to prolonged social isolation . OverallThe present study has two main drawbacks. The unprecedented conditions of this pandemic gave us the unique opportunity to explore the impact of a full lockdown on motor and non-motor symptoms in PD patients, but also limited the possibility to both perform an a-priori sample size calculation for our population, and to recruit an aged-matched control group. It is also worth noting that seven of the 12 participants completed an intensive motor rehabilitation program before the lockdown period, thus the overall outcome of the results might be affected. All the participants were yet at similar stage of the disease at the baseline. Another limitation worth mentioning is the unavoidable lack of control on how each participant managed to achieve a good quality of either the suggested or self-administered low-intensity physical exercises during the lockdown period.Our study provides the unique opportunity to test, with the same clinical measurements, in real life and in unprecedented mobility restrictions, the effects of isolation in PD. Even if the overall changes were modest, we observed an increased risk of falls in selected individuals which is nonetheless potentially relevant. The potentially negative effects of lockdown may have been softened by previous participation of these patients to educational activities and continued availability of our multidisciplinary team.


The adrenal gland is a common site of metastasis due to its rich blood supply. Adrenalectomy is typical treatment in the management of oligometastatic disease. We present an unexpected finding of cytomegalovirus (CMV)-related adrenalitis mimicking adrenal metastasis. A 54-year-old female was reviewed with a history of BRCA2-mutated, hormone receptor-positive invasive ductal cancer of the right breast diagnosed 12 years prior. Surveillance fluorodeoxyglucose positron emission tomography (FDG-PET) demonstrated a new focus of FDG avidity in the left adrenal gland, for which she underwent adrenalectomy. Histopathology revealed CMV-related adrenalitis in an otherwise immunocompetent patient without history of human immunodeficiency virus (HIV) or other immunocompromise. We describe the first case of CMV adrenalitis in a patient without acquired immunodeficiency syndrome. This case was initially presumed to be adrenal metastasis in the context of disseminated metastatic breast cancer and a PET-avid left adrenal lesion. The adrenal gland is a common site of metastasis due to its rich sinusoidal blood supply , with 13A 54-year-old female was referred to a tertiary referral surgeon for consideration of left adrenalectomy for presumed adrenal metastasis. The patient had a background of BRCA2-mutated, hormone receptor-positive invasive ductal cancer of the right breast diagnosed 12 years prior, for which she had undergone right mastectomy and axillary lymph node dissection, followed by completion of adjuvant endocrine, radiation and chemotherapy. She had also undergone subsequent radiation therapy for metastases to the brain and lumbar spine, with good response. There was no history of human immunodeficiency virus (HIV), AIDS or lymphopenia. There were no symptoms of CMV infection, and serology was not performed.Surveillance fluorodeoxyglucose positron emission tomography (FDG-PET) demonstrated a new focus of FDG avidity in the left adrenal gland . The adrThe patient underwent elective laparoscopic transabdominal left adrenalectomy. There were no intraoperative or postoperative complications, and she was discharged the day after surgery. On subsequent phone follow-up 4 weeks post-operation, the patient was well and reported return to normal function.No mass lesions could be seen on macroscopic examination of the left adrenal specimen . On micrPubMed and MEDLINE searches confirmed that there are no cases in the literature that describe CMV-related adrenalitis in an immunocompetent patient. This pathology is described in patients with AIDS, with a series of 74 autopsy cases of AIDS demonstrating CMV adrenalitis in 30 (41%), and remarking that the adrenal gland is the most frequently affected organ in AIDS .Outside of concurrent AIDS, CMV-associated adrenalitis has been described in two case reports of renal transplant recipients , 7. In bAnother case report of CMV-related adrenal insufficiency not associated with AIDS is described in a patient with Stage IV diffuse large B-cell lymphoma who initially demonstrated clinical remission following completion of eight cycles of rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP), but then developed progressive endocrine failure preceding death . AutopsyWe describe the first case of CMV adrenalitis in a patient without AIDS. This case was initially presumed to be adrenal metastasis in the context of disseminated metastatic breast cancer and a PET-avid left adrenal lesion. This unexpected finding reinforces that the surgeon must entertain a broad list of differentials when resecting lesions of indeterminate nature.None of the authors have any financial or professional disclosures.None.


ABSTRACT IMPACT: This work should provide further insights to mechanisms of the negative consequences of chemotherapy drugs, specifically in the cardiovascular system. OBJECTIVES/GOALS: Cardiotoxicity remains a safety concern in the development or utilization of chemotherapeutics largely due to the gap in knowledge of the mechanisms of toxicity. The pathophysiology of this cardiotoxicity has not been fully elucidated but data from our lab as well as other recent studies hint toward implications of mitochondrial (mito) biogenesis. METHODS/STUDY POPULATION: Prophylactic use of the beta-blocker carvedilol as well as the ACE inhibitor enalapril have been shown to inhibit the development of anthracycline-induced toxicity, but the mechanism of this cardio-protection remains elusive. To explore this, human stem cell-derived cardiomyocytes and endothelial cells will be either treated with the anthracycline doxorubicin or pretreated with carvedilol or enalapril followed by doxorubicin treatment before cellular lysates are harvested. Western blotting and qPCR will be performed to determine the expression of mito biogenesis markers including Nrf1, TFAM and the master regulator of mito biogenesis, PGC-1α. RESULTS/ANTICIPATED RESULTS: We anticipate that doxorubicin treatment alone will result in decreased expression of the mito biogenesis markers Nrf1, TFAM and PGC-1α and that pretreatment with either carvedilol and/or enalapril prior to doxorubicin treatment will either prevent or reverse this. DISCUSSION/SIGNIFICANCE OF FINDINGS: Doxorubicin’s role in causing mitochondrial dysfunction as well as suppression of biogenesis has already been established. Ideally, generation of new mitochondria would offset the occurrence of dysfunctional mitochondria. Confirming carvedilol/enalapril’s involvement with mito biogenesis would provide a mechanism of cardio-protection.


F1 hybrid varieties with higher performance, a phenomenon known as heterosis. The inability to fully exploit heterosis in outcrossing grasses is partially responsible for lower levels of improvement in breeding programmes compared with inbred crops. However, SI can be overcome in forage grasses to create self-compatible populations. This is generating interest in understanding the genetical basis of self-compatibility (SC), its significance for reproductive strategies and its exploitation for crop improvement, especially in the context of F1 hybrid breeding.Self-incompatibility (SI) systems prevent self-fertilization in several species of Poaceae, many of which are economically important forage, bioenergy and turf grasses. Self-incompatibility ensures cross-pollination and genetic diversity but restricts the ability to fix useful genetic variation. In most inbred crops it is possible to develop high-performing homozygous parental lines by self-pollination, which then enables the creation of Lolium–Festuca complex and the use of SC to develop immortalized mapping populations for the dissection of a wide range of agronomically important traits. The germplasm available is a valuable practical resource and will aid understanding the basis of inbreeding depression and hybrid vigour in key temperate forage grass species.We review the literature on SI and SC in outcrossing grass species. We review the currently available genomic tools and approaches used to discover and characterize novel SC sources. We discuss opportunities barely explored for outcrossing grasses that SC facilitates. Specifically, we discuss strategies for wide SC introgression in the context of the A better understanding of the genetic control of additional SC loci offers new insight into SI systems, their evolutionary origins and their reproductive significance. Heterozygous outcrossing grass species that can be readily selfed facilitate studies of heterosis. Moreover, SC introduction into a range of grass species will enable heterosis to be exploited in innovative ways in genetic improvement programmes. S locus containing at least two tightly linked genes, encoding the male and female-specific SI determinants that mediate the self/non-self-pollen recognition. If the S alleles carried by the pollen and pistil match, the pollen is recognized as ‘self’, thus preventing self-fertilization and the resulting inbreeding.Self-incompatibility (SI) is the most widespread mechanism for promoting outcrossing in hermaphrodite flowering plants. Described by Charles Darwin as ‘the ability of some plants to reject their own pollen’, SI is considered the mating system of ancestral angiosperms . The firThe breakdown of SI caused a change from outcrossing to self-fertilization in the evolutionary history of many flowering plants , genetic mapping studies employ highly heterozygous pseudo-F1 or pseudo-F2 populations that need to be maintained clonally. This is labour-intensive and potentially compromises the genetic integrity of the population over time.Thus, genetic mapping and dissection of agronomic traits have been useful strategies for the development of targeted approaches to DNA marker trait selection in many inbred crops by developing durable immortalized inbred line-based genetic resources. These include recombinant inbred lines (RILs) produced through recurrent rounds of inbreeding . These pSeveral self-incompatible species of the Poaceae are economically valuable forage crops and represent a fundamental component in many ruminant production systems. Self-incompatibility has acted as a barrier to their improvement and forage yield advances in breeding have been small . CurrentThe need to accelerate breeding progress in forage grasses for traits such as biomass and seed yield, nutritional quality and disease resistance is stimulating grass breeders to develop hybrid breeding technologies as an alternative to recurrent population selection strategies. For this reason, overcoming SI and exploiting SC to breed forage grasses as an inbred line-based crop is gaining interest  or in tapetal cells . In a self-pollination event, the products of male and female determinants interact with each other and trigger the pollen inhibition response that leads to SI  (Beta vulgaris) (Lilium martagon)   and fourartagon) . Of thesS and Z, which are located on chromosomes 1 and 2, respectively    containing the DUF247 domain of unknown function appeared to be the most likely candidate for the pollen SI determinant as it possessed the high level of allelic variability expected of an SI determinant gene affected by frequency-dependent selection   and rye cereale) , later bL. perenne Z locus. Two genes were identified as plausible candidates for either the pollen or stigma determinants. Interestingly, one of these was a gene encoding a protein containing a DUF247 domain (LpDUF247) similar to the S locus-linked LpSDUF247 gene, with the other candidate gene encoding either a putative tetratricopeptide repeat-like domain-containing protein or a ubiquitin-specific protease (LpTC116908). These findings lend support to a hypothesis put forward by LpSDUF247 have also been identified in other grass species, such as wild rice (Oryza barthii) and sorghum (Sorghum bicolor), suggesting that this gene could function as an SI determinant throughout the Poaceae (2+) signalling and protein phosphorylation are involved in the recognition and/or inhibition of incompatible pollen. Two independent expression studies found that transcripts predicted to code for proteins containing calcium-binding domains were enriched during the SI response in L. perenne and in sheepgrass (Leymus chinensis) , a naturally self-compatible species. By analysing SC segregation in backcrossing generations of L. temulentum–L. perenne and L. temulentum–annual ryegrass (Lolium multiflorum) interspecific hybrids, Z locus. This was inferred from distorted segregation of the GOT/3 isozyme locus in selfed progeny of first-generation backcross plants. Information available at the time showed that the GOT/3 locus was not linked to the S locus and therefore distorted segregation was inferred to have been associated with the Z locus. Later, linkage mapping experiments located the GOT/3 locus on linkage group (LG) 3  (Arabidopsis lyrata (Identifying sources of SC is more effective with the advent of next-generation sequencing (NGS)-based technologies and reference genome assemblies of grass species . Performin vitro self-pollination to phenotype SC with high resolution and accuracy  (Zea mays) (Pisum sativum) (Solanum lycopersicum) (Hordeum vulgare)  offers a high-throughput screening of point mutations at genes of interest in populations mutagenized by chemical treatment  proceed to the next generation. Rounds of recurrent phenotypic selection are made to gradually increase performance and at the same time populations are tested for uniformity. When the desired levels of performance and uniformity are reached the population is then intermated in isolation from possible contaminant pollen from outside sources for several generations to produce commercial quantities of seed. Levels of genetic improvement in forage grass yield have been noticeably modest in comparison with other crops ((Z. mays . RegardlL. perenne (Panicum virgatum)   and otheirgatum) . These aF1 hybrid breeding strategies by producing elite homozygous parental lines capturing favourable genetic diversity. These lines can be crossed for testing heterotic combinations to maximize combining ability in F1 hybrids. Although the challenges are considerable, the transition from populations to inbred line-based F1 hybrid breeding is gaining traction , overcoming inbreeding depression by incorporating SC in selected germplasm is feasible and provides the possibility of creating novel, valuable inbred germplasm for F1 hybrid breeding . For effective exploitation of SC in breeding programmes it is crucial that a broad base of self-compatible germplasm is available. Introgressing SC within the Lolium genus would present little difficulty because all species have the same ploidy level (2n = 2x = 14) and show high levels of homoeologous chromosome pairing in interspecific hybrid offspring  autopolyploids, exhibit an effective SI system and transfer of SC into a range of inter-fertile polyploid Lolium and Festuca species and hybrids could be proposed. For SC to be expressed in grass polyploids, introgressing SC into polyploid germplasm is a necessity and offers potential advantages over self-compatible diploid counterparts. For instance, theory predicts that polyploidy reduces the effect of the inbreeding depression that inevitably arises after repeated selfing of highly heterozygous plants (n = 2x = 28) L. perenne the source of SC on LG 5 is functional  complex produce viable inter-generic hybrids (× Festulolium) that are of increasing interest to farmers because of their inherent superior abiotic stress tolerance and adaptability to a range of climate change scenarios  is straightforward and natural hybrids are commonly found where the species are sympatric. Though previous reports have shown difficulties in producing seeds in sufficient quantity from the interspecific hybrids between L. perenne and F. pratensis, embryo rescue can increase the numbers of individuals recovered . Thus, the generic Lolium × Festuca hybrids, × Festulolium, provide a valuable target for transferring SC, especially as they display heterosis and reduced genetic load brought about by the combination of contrasting genomes  (Dactylis glomerata)  (Z. mays (Heterosis is a complex trait expressed through the action of multiple loci . In riceF1 hybrids exploiting genetic variation for hybrid vigour. Fundamental studies of the genetic basis of the heterosis that SC facilitates will be critical for targeted practical application. The systematic investigation of the agronomic performance of hybrids and the design of efficient seed production systems will be pivotal for efficient production of first-generation hybrid or pure-line cultivars. Afforded by SC mutations, research exploring heterosis in normally outcrossing species, with high levels of accumulated heterozygosity and genetic diversity, offers the prospect of contributing significantly to our understanding of heterosis currently gained from model species and self-fertile crops.Identifying and characterizing the basis of SC in grasses serves to provide a better understanding of the genes involved in the induction of SC, and knowledge of how SC arises in normally self-incompatible species can help biologists determine the pathways involved in the SI process in grasses. This offers insights into the evolutionary relationship of the grass SI system with other SI systems and, more widely, with other cell–cell recognition processes in living organisms. The subsequent exploitation of SC has the potential to promote more effective breeding methodologies based on the development of population or https://academic.oup.com/aob and consist of the following. Table S1: summary of published literature on SI breakdown to SC in flowering plants. References S1: references cited in Table S1.Supplementary data are available online at mcab043_suppl_Supplementary-Material-S01Click here for additional data file.mcab043_suppl_Supplementary-Material-S02Click here for additional data file.


CYP21A2 genotyping.We thank Lao Q and Merke DP for their interest in the EMQN-CAH-practice guidelines for molecular genetic testing of CAH  and theiTNXB gene. In the majority of those patients CAH-X is currently probably missed.We completely agree with the authors of this corresponding letter that CAH-X (CAH with a hypermobility type Ehlers–Danlos syndrome (EDS) due to pathogenic tenascin-X variants) is present in a considerable number of CAH patients , particuTNXB and its pseudogene TNXA, resulting in a contiguous CYP21A2 and TNXB deletion associated with EDS (CAH-X).As addressed in previous work, the use of the CAH-MLPA assay also covering exon 35 of the TNXB-gene could help in identification of a chimeric recombination between TNXB chimeras and pathogenic variants [So far, CAH-X diagnostic evaluation was not included in the CAH genotyping guideline, as from the EMQN-CAH scheme there is only very limited knowledge and experience concerning CAH-X genotyping. Comprehensive and reliable CAH-X genotyping would also include analysis of additional variants  not currTNXB-variants in the course of routine CYP21A2 gentoyping (in case MLPA is performed) is seen as incidental finding and to which extent such findings are communicated to the individual the blood sample of whom was analyzed . This has been controversially discussed during a previous EMQN-CAH-scheme-meeting, when a sample showed a reduced signal for TNXB.Moreover, the respective institutions/laboratories have to define whether findings of CYP21A2 genotyping. So, it can be foreseen that a considerable number of individuals who undergo CAH/CAH-X genotyping have signs and symptoms of hypermobility not related to pathogenic variants of TNXB. Thus, adequate strategies have to be developed and defined in phenotypically hypermobile individuals without CAH-X in cooperation and accordance with the respective professional societies.Another important aspect in that context is that hypermobility is frequently seen and only a minority of cases is due to CAH-X. This has to be taken into account when family members or partners of CAH-patients undergo We fully agree that CAH-X newborn screening would enable early diagnosis for young children before hypermobility evaluation is possible and would allow early screening for cardiac defects. In conclusion, we greatly appreciate Lao’s and Merke’s comment emphasising the importance of detecting CAH-X in patients suffering from 21-OH-defiency, as early detection and adequate therapeutic management of CAH-X will help to improve the patients’ quality of life and could prevent serious cardiac and vascular complications.In that context it would be of major importance that further studies are performed in order to broaden the knowledge and expertise on CAH-X before including respective methods in routine diagnostic procedures.


Enzymatic cleavage of the new substrate by MAGL was demonstrated, and kinetic constants Km and Vmax were determined. 6-O-arachidonoylluciferin has proved to be a highly sensitive substrate for MAGL. The bioluminescence assay  is much more sensitive and should suffer fewer biological interferences in cells lysate applications than typical fluorometric methods. The assay was validated for the identification and characterization of MAGL modulators using the well-known MAGL inhibitor JZL184. The use of PLG2 displaying distinct bioluminescence color and kinetics may offer a highly desirable opportunity to extend the range of applications to cell-based assays.A novel bioluminescent Monoacylglycerol lipase (MAGL) substrate 6-O-arachidonoylluciferin, a D-luciferin derivative, was synthesized, physico-chemically characterized, and used as highly sensitive substrate for MAGL in an assay developed for this purpose. We present here a new method based on the enzymatic cleavage of arachidonic acid with luciferin release using human Monoacylglycerol lipase ( In the last decade, the endocannabinoid system (ECS) has stimulated renewed interest, thanks to medical research results based on modern technologies highlighting its large distribution in human tissues ,2 and itMAGL inhibitors are potential antiproliferative and cytostatic drugs for cancer chemotherapy not only for enhancing endocannabinoid signaling but also controlling fatty acid release for synthesis of protumorigenic signaling lipids. Moreover, these inhibitors are also antineurodegenerative compounds that induce an anti-inflammatory activity by reducing the production of eicosanoids. The role of MAGL as a therapeutic target has increased interest not only in the development of selective inhibitors but also in the set-up of efficient protocols amenable to high-throughput screening (HTS) formats. In this context, different analytical methods to determine MAGL activity, generally based on its enzymatic action on fatty acid compounds, have been developed using various techniques. The procedures most used to screen MAGL inhibitors quantify the arachidonic acid released from 2-AG hydrolysis using high-performance liquid chromatography (HPLC) coupled with UV  or mass 2), a benzothiazole-containing compound, in a two-step reaction pyrrolidin-2-one) and ArLuc-2 produced. The values of negative controls were subtracted from the enzymatic curve. The initial velocity was determined from the linear portion of the resulting curve. All data were elaborated using GraphPad Prism 6.0c , and kinetic parameters Km and Vmax were calculated applying a nonlinear regression analysis (Michaelis–Menten). The quantitative data were calculated as means ± standard errors. The IC50 was calculated with Prism 6 (GraphPad software) using a log (inhibitor) vs. response–Variable slope (four parameters) function.All experiments were performed in triplicate and independently replicated at least once. A calibration curve was constructed to convert luminescence data in to LH1, has been synthesized and was found to enable highly sensitive  bioluminescent detection of MAGL activity in vitro for potential use in the study of MAGL inhibitors. Recombinant PLG2 was also produced and purified from bacterial cells in good yield (28.7 mg/L). This luciferase variant functioned well in the presence of 2.5% DMSO, confirming its stability and resistance to potentially denaturing conditions. The compatibility of the BL-based MAGL assay with DMSO, which is essential for solubilizing ArLuc-1, suggests that additional applications of PLG2 can be developed with reagents that are poorly soluble in aqueous buffers. The BL-assay described here will very likely expand the scope of MAGL inhibitor screening, which often involves solubilization of inhibitors in DMSO. The major advantages of this new bioassay for MAGL activity are high sensitivity and rapidity making it very suitable in applications when the amount of biological sample is limited. ArLuc-1 represents a valuable addition to the available toolkit of substrates useful for determining the activity of MAGL and can allow the development of efficient HTS protocols for the screening of potential selective inhibitors of this endocannabinoid metabolic enzyme. Furthermore, the very high sensitivity of this method together with the lower biological interference associated with bioluminescence compared to fluorescence-based methods suggests likely applicability to ex vivo or in vivo research.Fervent research on the ECS in the last decade has focused on the application of new techniques to deeply investigate the biochemical mechanisms of action of this ubiquitous system. The modulation of the activity of the ECS enzymes remains an important challenge for mechanistic and pharmacological purpose. For this reason, having a range of useful tools for varied research interests such as HTS methods for enzymes inhibitors or activators and development of methods for ex vivo and in vivo assessments is of fundamental importance. Our work, focused on the enzyme MAGL, aimed to add a new useful and very sensitive tool for ECS research. The efficacy of the newly developed luminescence-based assay for measuring the activity of MAGL has been demonstrated. D-luciferin esterified with arachidonic acid, ArLuc-


The rising rates of women in prison is a serious public health issue. Unlike men, women in prison are characterised by significant histories of trauma, poor mental health, and high rates of substance use disorders (SUDs). Recidivism rates of women have also increased exponentially in the last decade, with substance related offences being the most imprisoned offence worldwide. There is a lack of evidence of the effectiveness of post-release programs for women. The aim of this systematic review is to synthesise and evaluate the evidence on post-release programs for women exiting prison with SUDs.We searched eight scientific databases for empirical original research published in English with no date limitation. Studies with an objective to reduce recidivism for adult women (⩾18 years) with a SUD were included. Study quality was assessed using the revised Cochrane Risk of Bias tool for randomized trials (RoB2) and the Risk of Bias in Non-randomized Studies - of Interventions (ROBINS-I) tools.n = 3799 women) met the inclusion criteria. Recidivism was significantly reduced in five (42%) programs and substance-use was significantly reduced in one (8.3%) program. Common attributes among programs that reduced recidivism were: transitional, gender-responsive programs; provision of individualised support; providing substance-related therapy, mental health and trauma treatment services. Methodological and reporting biases were common, which impacted our ability to synthesize results further. Recidivism was inconsistently measured across studies further impacting the ability to compare results across studies.Of the 1493 articles, twelve (Recidivism is a problematic measure of program efficacy because it is inconsistently measured and deficit-focused, unrecognising of women’s gains in the post-release period despite lack of tailored programs and significant health and social disadvantages. The current evidence suggests that women benefit from continuity of care from prison to the community, which incorporated gender-responsive programming and individualised case management that targeted co-morbid mental health and SUDs. Future program design should incorporate these attributes of successful programs identified in this review to better address the unique challenges that women with SUDs face when they transition back into the community.The online version contains supplementary material available at 10.1186/s40352-021-00162-6. There was a low overall prevalence for preference of prescription drugs (8.0%), however twice as many women strongly preferred this category compared to men     programs are interventions that are delivered in the community. Transitional programs are interventions that start pre-release (in custody) and support people during the transition from prison to community Baldry, . Post-reDespite the growth of the women’s prison population, and their profoundly different criminogenic profile compared to incarcerated men, the majority of prison programs available have been designed for men and extended to women with little alteration  programs offered to women with SUDs to inform program development to decrease reoffending. Further, as the link between criminal offending and substance-use is well established, we also aim to review the effectiveness of interventions to reduce substance-use outcomes post-release and whether this impacts recidivism. This review addresses the following research questions (RQ):Post-release”, “Prison”, “Women”, and “Interventions”. The electronic database searches were supplemented with manual searches of the reference lists from relative articles. Due to the limited number of publications found an updated search was conducted in February 2020 following the method by Bramer and Bain (2017), adding search terms related to “Substance use” and “Recidivism”  guidelines  with an objective to reduce recidivism for adult women (⩾18 years) with a known SUD. The program had to be either a post-release or transitional intervention, published in English in a peer-reviewed journal. In this review substance-use included individuals using occasional drugs or alcohol, those who were dependent, or those who had other drug and alcohol related problems prior to their current offence. Studies that included both men and women were included if the results relating to women could be isolated. Due to the limited published studies of women in prison Baldry,  there weSearch results were imported into Endnote X7 software, duplicates removed, and results exported into Covidence online software. Two investigators independently applied eligibility criteria to titles and abstracts and discrepancies identified through the platform were discussed in a face-to-face meeting. Studies that were included were progressed to full-text review where the investigators systematically went through individual articles thoroughly to check eligibility and documented reasons for exclusion. Discrepancies were resolved through face-to-face discussion and a third reviewer was approached when needed. The lead reviewer extracted data according to the template for intervention description and replication (TIDieR) checklist and guide       (n = 6)      of the studies        were compared with usual care (standard probation/parole) in 67% of studies      . A major benefit for transitional programs is the continuity of care from prison to the community. Transitional support has been previously shown to assist participants in retaining rehabilitative health gains and reducing the risk of injury and death which is high for women with SUDs post-release . This is problematic considering all studies included women with SUDs and the direct correlation between substance-use and criminal offending for women is well understood . The attributes that contributed to the success of these programs were transitional, gender-responsive interventions which provided individualised support through community case management, with the use of cognitive behavioural therapies, as well as having substance-use, MH and trauma services available  (RQ3). Six studies reviewed substance-use post-release, and of those, three reported reduced substance use among program participants (RQ1). Reductions in substance-use was associated with programs that offered gender-responsive support, used cognitive behavioural therapies, and provided substance-use treatment, MH and trauma services (imbedded or referred) (RQ3).These findings reinforce the existing evidence that the design of transitional programs need to address criminogenic risk factors of women in prison  TEMPLATE FOR COMPLETION.Additional file 4. The Risk Of Bias In Non-randomized Studies – of Interventions (ROBINS-I) assessment tool.