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Supplemental Figure 1. CDK4/6i + MEKi treatment does not modulate proteins within the apoptotic signaling pathway. Supplemental Figure 2. Growth of UM001 xenografts treated with MEKi and/or CDK4/6i. Supplemental Figure 3. RNA-seq samples cluster based treatment. Supplemental Figure 4. MEKi, CDK4/6i, and OxPhosi have reversible effects. Supplemental Figure 5. Cleaved caspase 3 staining in UM001 xenograft tumors.

Targeting cyclin-dependent kinases 4/6 (CDK4/6) represents a therapeutic option in combination with BRAF inhibitor and/or MEK inhibitor (MEKi) in melanoma; however, continuous dosing elicits toxicities in patients. Using quantitative and temporal in vivo reporting, we show that continuous MEKi with intermittent CDK4/6 inhibitor (CDK4/6i) led to more complete tumor responses versus other combination schedules. Nevertheless, some tumors acquired resistance that was associated with enhanced phosphorylation of ribosomal S6 protein. These data were supported by phospho-S6 staining of melanoma biopsies from patients treated with CDK4/6i plus targeted inhibitors. Enhanced phospho-S6 in resistant tumors provided a therapeutic window for the mTORC1/2 inhibitor AZD2014. Mechanistically, upregulation or mutation of NRAS was associated with resistance in in vivo models and patient samples, respectively, and mutant NRAS was sufficient to enhance resistance. This study utilizes an in vivo reporter model to optimize schedules and supports targeting mTORC1/2 to overcome MEKi plus CDK4/6i resistance.Significance: Mutant BRAF and NRAS melanomas acquire resistance to combined MEK and CDK4/6 inhibition via upregulation of mTOR pathway signaling. This resistance mechanism provides the preclinical basis to utilize mTORC1/2 inhibitors to improve MEKi plus CDK4/6i drug regimens. Cancer Discov; 8(5); 568–81. ©2018 AACR.See related commentary by Sullivan, p. 532.See related article by Romano et al., p. 556.This article is highlighted in the In This Issue feature, p. 517

Supplemental Table 1. RNA Seq z-score values for genes significantly up- or downregulated in MEKi-R or CDK4/6i-T groups compared to controls, corresponding to figure 3A. Data are indexed to show which genes are significant for each comparison (BHFDR < 0.05 and absolute log2(FC) > 1).

Frequent GNAQ and GNA11 mutations in uveal melanoma hyperactivate the MEK–ERK signaling pathway, leading to aberrant regulation of cyclin-dependent kinases (CDK) and cell-cycle progression. MEK inhibitors (MEKi) alone show poor efficacy in uveal melanoma, raising the question of whether downstream targets can be vertically inhibited to provide long-term benefit. CDK4/6 selective inhibitors are FDA-approved in patients with estrogen receptor (ER)–positive breast cancer in combination with ER antagonists/aromatase inhibitors. We determined the effects of MEKi plus CDK4/6 inhibitors (CDK4/6i) in uveal melanoma. In vitro, palbociclib, a CDK4/6i, enhanced the effects of MEKi via downregulation of cell-cycle proteins. In contrast, in vivo CDK4/6 inhibition alone led to cytostasis and was as effective as MEKi plus CDK4/6i treatment at delaying tumor growth. RNA sequencing revealed upregulation of the oxidative phosphorylation (OxPhos) pathway in both MEKi-resistant tumors and CDK4/6i-tolerant tumors. Furthermore, oxygen consumption rate was increased following MEKi + CDK4/6i treatment. IACS-010759, an OxPhos inhibitor, decreased uveal melanoma cell survival in combination with MEKi + CDK4/6i. These data highlight adaptive upregulation of OxPhos in response to MEKi + CDK4/6i treatment in uveal melanoma and suggest that suppression of this metabolic state may improve the efficacy of MEKi plus CDK4/6i combinations.

Supplementary Figures

Materials and Methods

The SARS-CoV-2 infection has been predominately associated with lung disease. However, emerging evidence has associated the COVID-19 infection with a hypercoagulable state. This hypercoagulable state can occur despite the use of anticoagulants and antiplatelets. In fact, it may even be the presenting symptom of COVID-19 in some patients. Thromboembolism associated with COVID-19 carries a worse prognosis and should be identified as early as possible. Therefore, we report 2 patients with arterial thrombosis in the form of limb ischemia in the setting of COVID-19.

Frequent GNAQ and GNA11 mutations in uveal melanoma hyperactivate the MEK–ERK signaling pathway, leading to aberrant regulation of cyclin-dependent kinases (CDK) and cell-cycle progression. MEK inhibitors (MEKi) alone show poor efficacy in uveal melanoma, raising the question of whether downstream targets can be vertically inhibited to provide long-term benefit. CDK4/6 selective inhibitors are FDA-approved in patients with estrogen receptor (ER)–positive breast cancer in combination with ER antagonists/aromatase inhibitors. We determined the effects of MEKi plus CDK4/6 inhibitors (CDK4/6i) in uveal melanoma. In vitro, palbociclib, a CDK4/6i, enhanced the effects of MEKi via downregulation of cell-cycle proteins. In contrast, in vivo CDK4/6 inhibition alone led to cytostasis and was as effective as MEKi plus CDK4/6i treatment at delaying tumor growth. RNA sequencing revealed upregulation of the oxidative phosphorylation (OxPhos) pathway in both MEKi-resistant tumors and CDK4/6i-tolerant tumors. Furthermore, oxygen consumption rate was increased following MEKi + CDK4/6i treatment. IACS-010759, an OxPhos inhibitor, decreased uveal melanoma cell survival in combination with MEKi + CDK4/6i. These data highlight adaptive upregulation of OxPhos in response to MEKi + CDK4/6i treatment in uveal melanoma and suggest that suppression of this metabolic state may improve the efficacy of MEKi plus CDK4/6i combinations.

Aging is accompanied by a progressive decline in immune function termed “immunosenescence”. Deficient surveillance coupled with the impaired function of immune cells compromises host defense in older animals. The dynamic activity of regulatory modules that control immunity appears to underlie age-dependent modifications to the immune system. In the roundworm Caenorhabditis elegans levels of PMK-1 p38 MAP kinase diminish over time, reducing the expression of immune effectors that clear bacterial pathogens. Along with the PMK-1 pathway, innate immunity in C. elegans is regulated by the insulin signaling pathway. Here we asked whether DAF-16, a Forkhead box (FOXO) transcription factor whose activity is inhibited by insulin signaling, plays a role in host defense later in life. While in younger C. elegansDAF-16 is inactive unless stimulated by environmental insults, we found that even in the absence of acute stress the transcriptional activity of DAF-16 increases in an age-dependent manner. Beginning in the reproductive phase of adulthood, DAF-16 upregulates a subset of its transcriptional targets, including genes required to kill ingested microbes. Accordingly, DAF-16 has little to no role in larval immunity, but functions specifically during adulthood to confer resistance to bacterial pathogens. We found that DAF-16-mediated immunity in adults requires SMK-1, a regulatory subunit of the PP4 protein phosphatase complex. Our data suggest that as the function of one branch of the innate immune system of C. elegans (PMK-1) declines over time, DAF-16-mediated immunity ramps up to become the predominant means of protecting adults from infection, thus reconfiguring immunity later in life.

Focal segmental glomerulosclerosis (FSGS) is a common cause of nephrotic syndrome, accounting for 40% of nephrotic syndrome in adults. FSGS has diverse clinical and morphological features and underlying pathogenesis. We present a case of a 33-year-old male presenting with acute systolic heart failure complicated with left ventricular thrombus with embolism to coronary circulation and bilateral deep vein thrombosis. He was found to have nephrotic range proteinuria with kidney biopsy showing FSGS. Association of FSGS with cardiomyopathy has been reported in children. However, in adults, according to our best knowledge, there have not been any report of FSGS and non-ischemic cardiomyopathy or it is at least underreported. Abbreviations FSGS: Focal segmental glomerulosclerosis; ESRD: End-stage renal disease; NOS: Not otherwise specified; LV: Left ventricle

AimTo describe and correlate electroretinographic responses with clinical and angiographic findings in retinal vasculitis (RV).MethodsMedical records of patients with diagnosis of RV at a tertiary eye centre from December 2017 to May 2021 were reviewed. Cases in which fluorescein angiography (FFA) and full field electroretinography (ffERG) were done within 1 month were included. FFAs were graded according to the Angiography Scoring for Uveitis Working Group from 0 to 40, where 0 is normal. A novel ffERG grading system was implemented where individual waves were graded for timing and amplitude and general ffERG score was determined with 6 being a perfect score.Results20 patients (34 eyes) were included. Mean age was 43.9±19.8 years; 70% were female. Median best-corrected visual acuity was 0.8 (0.08–1). Mean FFA score was 12.6±6.5. Median general ffERG score was 5 (0–6). 68% and 91% of eyes had responses with general ffERG scores ≥5 and 4, respectively. Flicker timing was most commonly affected.FFA scores weakly correlated with delayed photopic cone b-wave and flicker timing (p=0.03 and 0.016, respectively). Vitreous haze moderately correlated with delayed cone b-wave timing (pConclusionIn this study, RV was not frequently associated with severe global retinal dysfunction Higher FFA scores, and vitreous haze grading were weakly, but significantly, correlated with cone-generated ffERG responses.

Background: Liponeurocytomas are rare neurocytic neoplasms that most often arise in the posterior fossa and affect individuals in the third and fifth decades of life. Most reported cases of this unique tumor in the literature have described a favorable clinical prognosis without recurrence. However, increasing reports of recurrent cases prompted the World Health Organization, in 2016, to recategorize the tumor from Grade I to the less favorable Grade II classification. We conducted a systematic review to identify recurrent cases of this unique tumor and to summarize differences between the primary and recurrent cases of liponeurocytoma. Methods: A systematic review exploring recurrent liponeurocytoma cases was conducted by searching the PubMed, Google Scholar, and Scopus databases for articles in English. Abstracts from articles were read and selected for full-text review according to a priori criteria. Relevant full-text articles were analyzed for symptoms, imaging, location, histological, pathological, treatment, and recurrence-free time between the primary and recurrent cases. Results: Of 4392 articles, 15 articles accounting for 18 patients were included (level of evidence: IV) in the study. Recurrence-free time decreased from an average of 82 months between the primary tumor resection to first recurrence to 31.3 months between the first and second recurrence. Recurrent tumors demonstrated increased pleomorphic neural cells, necrosis, vascular proliferation, and MIB-1 index when compared to the primary tumor. Several cases also demonstrated decreased lipidizing components when compared to the primary tumor, further indicating increased dedifferentiation. The primary treatment for this tumor was surgical resection with occasional adjunctive radiotherapy. Conclusion: Recurrent cases of liponeurocytoma have features of increased malignant proliferation compared to the primary cases. The standard treatment for these primary and recurrent tumors is gross total resection. The role of adjunctive radiotherapy remains a matter of debate.

Aim To describe the clinical characteristics and outcome of hospitalized COVID-19 patients with diabetic ketoacidosis (DKA). Methods We report eight cases of diabetic ketoacidosis in COVID-19 who presented to our institution in New Jersey, USA. COVID-19 was diagnosed by nasopharyngeal swab reverse transcription polymerase chain reaction (RT-PCR). The patients' electronic medical records were reviewed. Data on patients' age, sex, ethnicity, laboratory values, glycosylated hemoglobin level, oral antihyperglycemic agents (OHAs), insulin, and clinical outcomes were collected. Results The median age of the patient was 42.5 years, and seven were males and one was female. Out of eight patients, five had type 2 diabetes mellitus (DM), two had undiagnosed DM, and one had type 1 DM. Median value of initial glucose on presentation was 454 mg/dL. Median value of HbA1c on presentation was 11.4% and of anion gap was 26.5 mEq/L. Four patients had large ketonemia, one patient had moderate ketonemia, and three patients had small ketonemia. All the patients were started on standard treatment protocol for DKA with intravenous fluids and IV insulin infusion. Acute kidney injury (AKI) was seen in four patients, and one patient required renal replacement therapy. Out of eight patients, three required mechanical ventilation, and the same three patients died. Conclusion Our case series shows that COVID-19 infection can precipitate DKA in patients with known diabetes mellitus patients or as a first manifestation in undiagnosed DM patients; COVID-19 with DKA is associated with substantial mortality. Further studies are needed to characterize poor risk factors associated with mortality in these patients.

Coronavirus disease 2019 (COVID-19), caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is characterized by an overwhelming inflammatory response in a subset of patients, resulting in respiratory compromise, multiorgan failure, and death. A common complication seen in patients hospitalized with COVID-19 infections is the development of venous and arterial thromboemboli. This occurs especially in patients who suffer from severe respiratory or systemic manifestations on the COVID-19 spectrum of disease. We present a case of acute limb ischemia as the initial presentation in a patient who tested positive for COVID-19.

BACKGROUND AND OBJECTIVES: To describe the clinical characteristics and outcomes of hospitalized coronavirus disease 2019 (COVID-19) patients with diabetic ketoacidosis (DKA) -- a single center tertiary hospital experience MATERIALS AND METHODS: A retrospective study was conducted among patients admitted to our hospital in the United States between March 1st and June 15th, 2020 with DKA and severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection known as COVID-19 We compared the baseline characteristics, laboratory data, and clinical course between survivors and nonsurvivors to identify the risk factors associated with mortality in the patients with DKA RESULTS: A total number of 43 patients were included in this study The median age was 52 years Thirty-three (76 7%) patients were male Median value of initial glucose on presentation was 553 mg/dL (300 0-1927 0 mg/dL) On admission, 33 (76 7%) patients had glycated hemoglobin (HbA1c)>=8% (64 mmol/mol) and HbA1cwas not obtained in 10 (23 3%) patients Acute kidney injury (AKI) was seen in 37 (86 0%) patients, 6 (14%) patients required renal replacement therapy and 22 (51 2%) required mechanical ventilation Among the 43 patients, 25 (58 1%) died Out of 25 patients who died 15 (60 0%) were Hispanics, 6 (24 0%) were White, 3 (12 0%) were African American, 1 (4%) was Arabic, and 1 (4%) was Asian The patients who died were older in age than who survived (mean age 58 +/- 6 13 vs 46 +/- 9 39;p = 0 023) Some 95% of the patients requiring mechanical ventilation died (odds ratio [OR]: 89 25;95% confidence interval [CI]: 9 10-874 96);p=0 001) Compared to survivors, nonsurvivors had significantly higher d-dimer (13 00 +/- 3 20 mcg/mL vs 6 15 +/- 3 66 mcg/mL;p< 0 006) and peak ferritin values (2763 66 +/- 1105 32 ng/mL vs 835 16 +/- 257 07 ng/mL;p= 0 016) Conclusion: Our retrospective study shows COVID-19 infection may present as DKA in patients with diabetes mellitus (DM) Older age, mechanical ventilation, elevated d-dimer, and ferritin are associated with poor prognosis in these patients Our study shows that COVID-19 is associated with substantial mortality in DKA patients and adds to the limited literature available regarding poor risk factors associated with mortality in these patients

Methamphetamine-induced keratitis is a recognized but not widely studied cause of corneal stromalysis and infections. These 2 cases describe the presentation and clinical course of keratitis in patients with preceding methamphetamine use.

Extraintestinal Clostridium difficile infection (CDI) is extremely uncommon. High mortality and poor outcomes have been observed among individuals with this rare medical condition. Empyema is one of the extraintestinal manifestations of CDI. Possible mechanisms to develop this parapneumonic effusion are aspiration and contamination of the chest tube. We present a 42-year-old Hispanic male with C. difficile empyema without any prior history of CDI.

Targeting cyclin-dependent kinases 4/6 (CDK4/6) represents a therapeutic option in combination with BRAF inhibitor and/or MEK inhibitor (MEKi) in melanoma; however, continuous dosing elicits toxicities in patients. Using quantitative and temporal in vivo reporting, we show that continuous MEKi with intermittent CDK4/6 inhibitor (CDK4/6i) led to more complete tumor responses versus other combination schedules. Nevertheless, some tumors acquired resistance that was associated with enhanced phosphorylation of ribosomal S6 protein. These data were supported by phospho-S6 staining of melanoma biopsies from patients treated with CDK4/6i plus targeted inhibitors. Enhanced phospho-S6 in resistant tumors provided a therapeutic window for the mTORC1/2 inhibitor AZD2014. Mechanistically, upregulation or mutation of NRAS was associated with resistance in in vivo models and patient samples, respectively, and mutant NRAS was sufficient to enhance resistance. This study utilizes an in vivo reporter model to optimize schedules and supports targeting mTORC1/2 to overcome MEKi plus CDK4/6i resistance. Significance: Mutant BRAF and NRAS melanomas acquire resistance to combined MEK and CDK4/6 inhibition via upregulation of mTOR pathway signaling. This resistance mechanism provides the preclinical basis to utilize mTORC1/2 inhibitors to improve MEKi plus CDK4/6i drug regimens. Cancer Discov; 8(5); 568–81. ©2018 AACR. See related commentary by Sullivan, p. 532. See related article by Romano et al., p. 556. This article is highlighted in the In This Issue feature, p. 517

We present an interesting case of a 73-year-old female who presented to the emergency department (ED) with complaints of abdominal pain for the past few days. Patient admitted history of “stomach problems” for the past few years with similar attacks every 3 - 4 months and attributed this to her “gallbladder”. After her dinner the night before, she started experiencing intense abdominal pain, which prompted her to come to the ED for evaluation. A portable chest X-ray was acquired which showed herniation of the gas-distended stomach overlying the left lung base. CT scan with IV contrast was then performed which confirmed a left-sided diaphragmatic hernia containing the antrum of the dilated stomach but the antrum was twisted to the left of the esophagogastric (EG) junction. There was pneumatosis in the wall of the caudal aspect of the gastric body, suggestive of ischemia. The patient underwent emergent exploratory laparotomy, derotation, partial gastrectomy and gastrostomy where she was found to have an adult Bockdalek hernia on the left side. We discuss this rare case and review relevant literature. J Curr Surg. 2017;7(1-2):15-19 doi: https://doi.org/10.14740/jcs315w

Peliosis hepatis (PH) is a rare vascular condition of the liver characterized by the presence of cystic blood filled cavities distributed randomly throughout the liver parenchyma. We describe a case of a 42-year-old previously healthy male patient, airlifted to us in a state of hemorrhagic shock after undergoing percutaneous diagnostic liver biopsy for lesions seen on CT scan. Repeat CT scan with IV contrast on presentation in our hospital showed intraperitoneal bleeding. Hepatic angiography failed to identify any specific bleeding source. A diagnostic laparoscopy was performed and approximate 9 L of hemoperitoneum was evacuated. The postoperative course was complicated with rapid hemodynamic deterioration, associated with acute hepatic failure progressively leading to multiorgan failure and death in spite of aggressive intensive care support. We suggest that PH should be considered as differential diagnosis of hypervascular hepatic lesions. It potentially can cause fatal acute non-traumatic liver hemorrhage.

Aging is accompanied by a progressive decline in immune function termed “immunosenescence”. Deficient surveillance coupled with the impaired function of immune cells compromises host defense in older animals. The dynamic activity of regulatory modules that control immunity appears to underlie age-dependent modifications to the immune system. In the roundwormCaenorhabditis eleganslevels of the PMK-1 p38 MAP kinase diminish over time, reducing the expression of immune effectors that clear bacterial pathogens. Along with the PMK-1 pathway, innate immunity inC. elegansis regulated by the insulin signaling pathway. Here we asked whether DAF-16, a Forkhead box (FOXO) transcription factor whose activity is inhibited by insulin signaling, plays a role in host defense later in life. While in younger worms DAF-16 remains in an inactive state unless stimulated by environmental insults, we found that even in the absence of acute stress the transcriptional activity of DAF-16 increases in an age-dependent manner. Beginning in the reproductive phase of adulthood, DAF-16 upregulates a subset of its transcriptional targets, including genes required to kill ingested microbes. Accordingly, DAF-16 has little to no role in larval immunity, but functions specifically during adulthood to confer resistance to bacterial pathogens. We found that DAF-16-mediated immunity in adults requires SMK-1, a regulatory subunit of the PP4 protein phosphatase complex. Our data suggest that as the function of one branch of the innate immune system ofC. elegans(PMK-1) declines over time, DAF-16-mediated immunity ramps up to become the predominant means of protecting adults from infection, thus reconfiguring immunity later in life.

Targeting cyclin-dependent kinases 4/6 (CDK4/6) represents a therapeutic option in combination with BRAF inhibitor and/or MEK inhibitor (MEKi) in melanoma; however, continuous dosing elicits toxicities in patients. Using quantitative and temporal in vivo reporting, we show that continuous MEKi with intermittent CDK4/6 inhibitor (CDK4/6i) led to more complete tumor responses versus other combination schedules. Nevertheless, some tumors acquired resistance that was associated with enhanced phosphorylation of ribosomal S6 protein. These data were supported by phospho S6 staining of melanoma biopsies from patients treated with CDK4/6i plus targeted inhibitors. Enhanced phospho S6 in resistant tumors provided a therapeutic window for the mTORC1/2 inhibitor, AZD2014. Mechanistically, upregulation or mutation of NRAS was associated with resistance in in vivo models and patient samples, respectively, and mutant NRAS was sufficient to enhance resistance. This study utilizes an in vivo reporter model to optimize schedules and supports targeting mTORC1/2 to overcome MEKi plus CDK4/6i resistance.

We report 11 cases of combined diabetic ketoacidosis (DKA) and hyperglycemic hyperosmolar nonketotic coma (HHNK) in coronavirus 2019 patients who presented to our institution in New Jersey, USA. The median age was 47 years (range 12-88 years). Out of the 11 patients, 7 were male and 4 were female. Out of 11 patients, 8 had type 2 diabetes mellitus (DM), 2 had undiagnosed DM, and 1 had type 1 DM. Presenting complaints included altered mental status, weakness, shortness of breath, cough, fever, vomiting, abdominal pain, chest pain, and foot pain. Out of 11 patients, pneumonia was diagnosed at presentation in 8 patients, while in 3 patients, chest X-ray was clear. Median value of initial glucose on presentation was 974 mg/dL (range 549-1556 mg/dL), and hemoglobin A1c on presentation was 13.8%. The median value of anion gap was 34 mEq/L. Out of the 11 patients, ketonemia was moderate in 6 patients, large in 3, and small in 2 patients. Acute kidney injury (AKI) occurred in 9 patients and 2 patients required renal replacement therapy. Out of the 11 patients, 6 required mechanical ventilation and 7 patients died. All the 6 patients requiring mechanical ventilation died. Our case series shows COVID-19 infection can precipitate acute metabolic complications in known DM patients or as first manifestation in undiagnosed DM patients. Patients can present with DKA/HHNK symptoms and/or respiratory symptoms. Mechanical ventilation is a poor prognostic factor. Further studies are needed to characterize prognostic factors associated with mortality in this vulnerable patient population.

Type IV paraesophageal hernia (PEH) is very rare and is characterized by the intrathoracic herniation of the abdominal viscera other than the stomach into the chest. We describe a case of a 90-year-old male patient who presented at our emergency department complaining of epigastric pain that he had experienced over the past few hours and getting progressively worse. On the day after admission, his pain became severe. Chest radiography revealed an intrathoracic intestinal gas bubble; emergency exploratory laparotomy identified a type IV PEH with herniation of only the jejunum with perforated diverticula on mesenteric side through a hiatal defect into mediastinum. There are a few published cases of small bowel herniation into the thoracic cavity in the literature. Our patient represents a rare case of an individual diagnosed with type IV PEH with herniation of jejunum with perforated diverticula.