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Mutant calreticulin (CALR) proteins resulting from a −1/+2 frameshifting mutation of the CALR exon 9 carry a novel C-terminal amino acid sequence and drive the development of myeloproliferative neoplasms (MPNs). Mutant CALRs were shown to interact with and activate the thrombopoietin receptor (TpoR/MPL) in the same cell. We report that mutant CALR proteins are secreted and can be found in patient plasma at levels up to 160 ng/mL, with a mean of 25.64 ng/mL. Plasma mutant CALR is found in complex with soluble transferrin receptor 1 (sTFR1) that acts as a carrier protein and increases mutant CALR half-life. Recombinant mutant CALR proteins bound and activated the TpoR in cell lines and primary megakaryocytic progenitors from patients with mutated CALR in which they drive thrombopoietin-independent colony formation. Importantly, the CALR-sTFR1 complex remains functional for TpoR activation. By bioluminescence resonance energy transfer assay, we show that mutant CALR proteins produced in 1 cell can specifically interact in trans with the TpoR on a target cell. In comparison with cells that only carry TpoR, cells that carry both TpoR and mutant CALR are hypersensitive to exogenous mutant CALR proteins and respond to levels of mutant CALR proteins similar to those in patient plasma. This is consistent with CALR-mutated cells that expose TpoR carrying immature N-linked sugars at the cell surface. Thus, secreted mutant CALR proteins will act more specifically on the MPN clone. In conclusion, a chaperone, CALR, can turn into a rogue cytokine through somatic mutation of its encoding gene.

Recent evidence shows an association between type 2 diabetes mellitus (T2DM) and liver steatosis, fibrosis, and hepatocellular carcinoma. Studies suggest that improved glycemic control, weight loss, and GLP-1 agonists may halt progression and even improve non-alcoholic fatty liver disease (NAFLD); however, little is known about the extent of improvement these factors have on liver fibrosis. ADA guidelines suggest only patients with T2DM or pre-DM and elevated liver enzymes (or fatty liver on ultrasound) should be evaluated for NAFLD and fibrosis. Three patients ages 65-71 with long-standing T2DM, HLD, obesity, and no history of liver disease or current alcohol use were followed in our diabetic clinic and referred for screening for liver steatosis and fibrosis. The transient elastography method (fibroscan) was used to determine the degree of liver fibrosis. On initial fibroscans, all three patients were found to have F4 scores consistent with liver cirrhosis. The first patient is a 71-year-old male with T2DM (BMI 37.5kg/m 2), treated with semaglutide, empagliflozin, glipizide, and insulin. Over two years, he experienced a 4-lb weight loss and improvement in HbA1c(7.8% to 7.1%). Repeat fibroscan after two years showed improved fibrosis score – F3 (11.6 kPa to 10.2 kPa), consistent with moderate fibrosis. The second patient is a 69-year-old female with T2DM (BMI 37.3kg/m 2) and transient mild LFT abnormalities, found to have cirrhosis on fibroscan – F4 score. She was managed with semaglutide, empagliflozin, metformin, and insulin; weight decreased 9-lbs over two years and HbA1c improved(10.8% to 7.8%). Repeat fibroscan showed improvement in fibrosis score to F2 (15.1 kPa to 8.9 kPa), and her LFT abnormalities resolved. The third patient is a 65-year-old male with T2DM (BMI 35kg/m 2). His initial fibroscan showed cirrhotic liver – F4 score. He was maintained on empagliflozin and dulaglutide- then switched to semaglutide. He experienced a 10-lb weight loss during this time and had a stable HbA1c(6.6%). Repeat fibroscan after one year showed improvement to F0-F1(11.3 kPa to 6.1 kPa) consistent with little to no liver scarring. These cases highlight two important factors: 1) ADA liver screening guidelines should be expanded to include obesity and dyslipidemia; and 2) advanced liver fibrosis in patients with diabetes is, to some degree, reversible. Not all of the patients initially met standard ADA screening criteria, mostly without elevated liver enzymes and no prior abnormal liver imaging. Yet, all were diabetic and obese with dyslipidemia, which are independent risk factors for NAFLD. Additionally, patients with improved glycemic control on GLP-1 treatment as well as some weight loss have promising results on repeat fibroscan. These findings may improve screening and management of diabetes-related liver disease. Further studies are needed to investigate to what extent GLP-1 agonists alone or in combination with glycemic control and weight loss contribute to these improvements. Presentation: No date and time listed

Relevance: A rare yet distinct cause of sudden onset paralysis is severe hypokalemia associated with thyrotoxicosis. This is thought to be associated with mutations in genes encoding cellular potassium channels. We report a case of acute onset paralysis with profound hypokalemia and a new diagnosis of Graves’ thyrotoxicosis in a previously asymptomatic African American, HIV+ man on highly active antiretroviral therapy (HAART) for over 8 years. Clinical case: A 49-year-old man with hypertension and HIV presented with acute paralysis of his bilateral upper and lower extremities. His initial potassium was 1.8 mEq/L (3.5-5.0). Prior to sudden onset loss of motor strength, he denied any preceding palpitations, tremor, anxiety, diaphoresis, hyper-defecation, weight loss, heat or cold intolerance, neck pain, increase in neck girth or difficulty swallowing, proptosis or other ocular symptoms. He has no family history of thyroid disease. He had an enlarged palpable thyroid without nodules and no audible bruit. There was no periorbital edema or proptosis, and no signs of dermopathy. A thyroid ultrasound showed a hyperemic and diffusely enlarged thyroid gland without nodules. Labs included a TSH of 0.007 mCU/mL (0.43-3.8), Free T4 2.1 ng/dL (0.71-1.85), Total T3 229.6 ng/dL (58-194), and thyrotropin receptor antibody 2.6 IU/L (0-1.75). The CD4+ count was 146 in 2010 with a slow gradual rise to 673 in 2019, and HIV viral load was undetectable. There were no offending medications or supplements identified. With aggressive potassium repletion, the serum potassium improved to 4.6 mEq/L and he regained normal strength within several hours. He was started on Methimazole 10mg daily and propranolol 10mg TID. At one month, the thyroid function tests normalized. Methimazole 10mg daily was continued and propranolol was tapered off. He remains euthyroid. Genetic testing is pending. Conclusions: Graves’ disease is the most common thyroid disease triggered by immune reconstitution in HIV + individuals on HAART. The incidence of thyroid disease in HIV patients on HAART is higher in women and Africans with a 1.5-2 fold increase compared with the general population [Muller et. al, Eur Thyroid J 2019;8:173-185]. Despite a higher incidence of hyperthyroidism in women, over 95% of cases of hypokalemic periodic paralysis have been reported in men, with a 10-fold higher incidence among Asians compared to Westerners. There has been one prior case report of Thyrotoxic Periodic Paralysis in an HIV + patient and this was in a Polynesian male (Brown JD et al. Hawaii Med J 2007). To our knowledge, this is the first case report of an African American HIV+ patient with this disorder. An association with mutations in the Kir2.6 gene (encodes a potassium channel, is expressed in skeletal muscle, and is transcriptionally regulated by thyroid hormone) has been proposed [Ryan et. al, Cell 2010 January 8; 140(1):88-98].

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Long non-coding RNAs (lncRNAs), an emerging group of gene expression regulators in normal and cancer cells, are defined as endogenous RNAs consisting of more than 200 nucleotides and lacking an ope...

We studied the impact of driver mutations of JAK2, CALR, (calreticulin gene) or MPL on clinical course, leukemic transformation, and survival of patients with primary myelofibrosis (PMF). Of the 617 subjects studied, 399 (64.7%) carried JAK2 (V617F), 140 (22.7%) had a CALR exon 9 indel, 25 (4.0%) carried an MPL (W515) mutation, and 53 (8.6%) had nonmutated JAK2, CALR, and MPL (so-called triple-negative PMF). Patients with CALR mutation had a lower risk of developing anemia, thrombocytopenia, and marked leukocytosis compared with other subtypes. They also had a lower risk of thrombosis compared with patients carrying JAK2 (V617F). At the opposite, triple-negative patients had higher incidence of leukemic transformation compared with either CALR-mutant or JAK2-mutant patients. Median overall survival was 17.7 years in CALR-mutant, 9.2 years in JAK2-mutant, 9.1 years in MPL-mutant, and 3.2 years in triple-negative patients. In multivariate analysis corrected for age, CALR-mutant patients had better overall survival than either JAK2-mutant or triple-negative patients. The impact of genetic lesions on survival was independent of current prognostic scoring systems. These observations indicate that driver mutations define distinct disease entities within PMF. Accounting for them is not only relevant to clinical decision-making, but should also be considered in designing clinical trials.

We studied the impact of driver mutations of JAK2, CALR, (calreticulin gene) or MPL on clinical course, leukemic transformation, and survival of patients with primary myelofibrosis (PMF). Of the 617 subjects studied, 399 (64.7%) carried JAK2 (V617F), 140 (22.7%) had a CALR exon 9 indel, 25 (4.0%) carried an MPL (W515) mutation, and 53 (8.6%) had nonmutated JAK2, CALR, and MPL (so-called triple-negative PMF). Patients with CALR mutation had a lower risk of developing anemia, thrombocytopenia, and marked leukocytosis compared with other subtypes. They also had a lower risk of thrombosis compared with patients carrying JAK2 (V617F). At the opposite, triple-negative patients had higher incidence of leukemic transformation compared with either CALR-mutant or JAK2-mutant patients. Median overall survival was 17.7 years in CALR-mutant, 9.2 years in JAK2-mutant, 9.1 years in MPL-mutant, and 3.2 years in triple-negative patients. In multivariate analysis corrected for age, CALR-mutant patients had better overall survival than either JAK2-mutant or triple-negative patients. The impact of genetic lesions on survival was independent of current prognostic scoring systems. These observations indicate that driver mutations define distinct disease entities within PMF. Accounting for them is not only relevant to clinical decision-making, but should also be considered in designing clinical trials.

We searched for JAK2 exon 12 mutations in patients with JAK2 (V617F)-negative myeloproliferative disorders. Seventeen patients with polycythemia vera (PV), including 15 sporadic cases and 2 familial cases, carried deletions or duplications of exon 12 in circulating granulocytes but not in T lymphocytes. Two of the 8 mutations detected were novel, and the most frequent ones were N542-E543del and E543-D544del. Most patients with PV carrying an exon 12 mutation had isolated erythrocytosis at clinical onset, unlike patients with JAK2 (V617F)-positive PV, most of whom had also elevations in white blood cell and/or platelet counts. Both patients with familial PV carrying an exon 12 mutation had an affected sibling with JAK2 (V617F)-positive PV. Thus, several somatic mutations of JAK2 exon 12 can be found in a myeloproliferative disorder that is mainly characterized by erythrocytosis. Moreover, a genetic predisposition to acquisition of different JAK2 mutations is inherited in families with myeloproliferative disorders.