Anupam Kotwal

Ca2+-dependent repair of plasma membrane breaches is essential for animal cell viability. An initial passive influx of extracellular Ca2+ triggers the formation of a protein plug that rapidly seals breaches. However, the mechanism of extracellular Ca2+ requirement for subsequent repair remains undefined. EHD2 protein stabilizes the plasma membrane caveolae, which sustain membrane repair, and maintains high surface levels of the caveolae-resident Ca2+ channel Orai1. We establish the requirement of both Orai1 and EHD2 for repair of plasma membrane lesions induced by mechanical injury or by a model bacterial pore-forming toxin. We demonstrate rapid EHD2 recruitment and Orai1-mediated Ca2+ entry at plasma membrane sites of localized mechanical stimulus, the latter requiring EHD2 and CAV1. EHD2 and Orai1 are necessary for mechanosensitive YAP/TAZ-TEAD activation and positive feedback for CAV1 expression that promotes membrane repair. Our studies establish EHD2 and Orai1 as novel components of mammalian plasma membrane repair and mechanoadaptation.

SUMMARY: Ca ²⁺ -dependent repair of plasma membrane breaches is essential for animal cell viability. An initial passive influx of extracellular Ca ²⁺ triggers the formation of a protein plug that rapidly seals breaches. However, the mechanism of extracellular Ca ²⁺ requirement for subsequent repair remains undefined. EHD2 protein stabilizes the plasma membrane caveolae, which sustain membrane repair, and maintains high surface levels of the caveolae-resident Ca ²⁺ channel Orai1. We establish the requirement of both Orai1 and EHD2 for repair of plasma membrane lesions induced by mechanical injury or by a model bacterial pore-forming toxin. We demonstrate rapid EHD2 recruitment and Orai1-mediated Ca ²⁺ entry at plasma membrane sites of localized mechanical stimulus, the latter requiring EHD2 and CAV1. EHD2 and Orai1 are necessary for mechanosensitive YAP/TAZ-TEAD activation and positive feedback for CAV1 expression that promotes membrane repair. Our studies establish EHD2 and Orai1 as novel components of mammalian plasma membrane repair and mechanoadaptation.

Indeterminate thyroid nodules carry a wide range of malignancy prevalence, necessitating incorporation of molecular testing to guide management. Molecular test results help guide prognosis to a certain extent, but additional prognostic factors need to be identified. Limited data have linked PLEKHS1 promoter ( PLEKHS1p ) mutations with higher aggressiveness of thyroid cancer. Hence, we aimed to evaluate the diagnostic and prognostic value of PLEKHS1p mutations in preoperative thyroid nodules undergoing molecular testing. We assessed PLEKHS1p mutations in 9279 patient samples from April 2023 to June 2024 among indeterminate thyroid nodules ((B)ethesda III/IV) with Afirma GSC‐(S)uspicious results as well as among B V/VI cytology thyroid nodules. We also analyzed a subset of 20 consecutive cases positive for PLEKHS1p mutations with surgical resection for histology and for co‐occurring molecular alterations from the Afirma testing. PLEKHS1p mutations were positive in 60/9279 (0.6%) of patient samples with three times higher frequency in B VI compared to B III cytology nodules (1.36% vs 0.47%, p < .01). Among the 60 samples harboring PLEKHS1p mutations, 28 had the C593T hotspot mutation, 38 the G590A mutation, and six samples had both; four samples had concomitant TERTp mutations and 18 samples had concomitant BRAFp.V600E alterations. Our study demonstrated an overall low frequency of PLEKHS1p mutations, but this frequency was highest among malignant (B VI) cytology thyroid nodules. The frequency of PLEKHS1p mutations did not strongly correlate with the severity of thyroid cancer but the surgical sample size was limited. Further research is needed to clarify the role of PLEKHS1p mutations in thyroid nodules and cancer.

Background: Thyroid dysfunction (TD) occurs commonly from immune checkpoint inhibitors (ICI) cancer therapy, but questions remain regarding its predicting factors, appropriate dosing for thyroid hormone replacement, and the strength of association with overall survival (OS). We aim to address these three questions in our study. Methods: We performed a retrospective cohort study of adult patients with cancer who received ICIs from December 1, 2012, to December 31, 2019. After excluding 28 patients with preexisting primary hypothyroidism, 811 patients were evaluated for the development of new-onset ICI-TD. Kaplan-Meier survival and log-rank tests were used to compare OS distributions between ICI-TD status groups, following which Cox regression models addressed immortal time bias (ITB). Results: Of the 811 included patients with a median follow-up of 19.2 months, 122 (15.0%) patients developed ICI-TD. The median age at initiation of ICIs was 64.8 years; women comprised 42.8% of the cohort. There were no significant differences in age, sex, race, malignancy type, or personal history of autoimmunity in patients who developed ICI-TD versus those who did not. ICI-TD occurred most frequently after combination ICI therapy (32%) compared with CTLA-4 ICI and PD-1/PD-L1 ICI monotherapy (p = 0.002). The median levothyroxine dose was the highest, being 1.41 mcg/kg/day in the overt hypothyroidism group. Patients with ICI-TD had a higher median pre-treatment log2(thyrotropin or TSH) level (1.2, corresponding to TSH 2.3 mIU/L) versus those without (0.79, corresponding to TSH 1.7 mIU/L; p = 0.008); however, the area under the curve was Conclusions: The levothyroxine dose needed for overt hypothyroidism from ICI-TD is similar to athyreotic hypothyroidism. While baseline TSH in the upper normal range is associated with an increased risk of ICI-TD, there is no absolute baseline TSH value that accurately predicts ICI-TD in the clinical setting. The link between ICI-TD and OS needs further validation after accounting for ITB.

Introduction: Dosimetry helps calculate the optimal iodine-131 (I-131) dose for treating metastatic differentiated thyroid cancer (DTC). We aimed to evaluate if recombinant human TSH (rhTSH) and thyroid hormone withdrawal (THW) are equivalent methods of preparation for dosimetry-guided I-131 therapy in metastatic DTC. Methods: We performed a retrospective cohort study of 51 adults with metastatic DTC who received I-131 with dosimetry from 2010 through 2022. Gamma camera and blood activity measurements were taken following the pretherapeutic I-131 dose. Statistical analysis compared rhTSH and THW groups; P < .05 was considered significant. Results: Fifty-one adults undergoing 55 I-131 dosimetry-guided treatments were included: 22 by rhTSH and 33 by THW. The median age was lower (P = .0008), and the proportion of stage IV (P = .009) was higher in rhTSH compared to the THW group. The terminal effective half-life at 24 to 48 hours in the whole body was longer in rhTSH compared to THW group (21.9 vs 17.1 hours; P = .014), but this difference was less significant when limited to the n = 37 metastatic cases (P = .046) and not different for red marrow effective half-life. The calculated allowed I-131 dose was lower in rhTSH compared to THW group (187.5 mCi vs 259.9 mCi; P = .0000). Thyroglobulin was higher during treatment in the rhTSH group (P = .031), whereas its reduction at 3 months was not different after adjusting for age and stage. Conclusion: rhTSH is noninferior to THW in preparation for I-131 dosimetry. Compared to THW, rhTSH results in lower calculated allowed I-131 dose after dosimetry, which could translate to fewer side effects or impact on quality of life.