Endocytosis in cancer and cancer therapy.

Endocytosis is a complex process whereby cell surface proteins, lipids and fluid from the extracellular environment are packaged, sorted and internalized into cells. Endocytosis is also a mechanism of drug internalization into cells. There are multiple routes of endocytosis that determine the fate of molecules, from degradation in the lysosomes to recycling back to the plasma membrane. The overall rates of endocytosis and temporal regulation of molecules transiting through endocytic pathways are also intricately linked with signalling outcomes. This process relies on an array of factors, such as intrinsic amino acid motifs and post-translational modifications. Endocytosis is frequently disrupted in cancer. These disruptions lead to inappropriate retention of receptor tyrosine kinases on the tumour cell membrane, changes in the recycling of oncogenic molecules, defective signalling feedback loops and loss of cell polarity. In the past decade, endocytosis has emerged as a pivotal regulator of nutrient scavenging, response to and regulation of immune surveillance and tumour immune evasion, tumour metastasis and therapeutic drug delivery. This Review summarizes and integrates these advances into the understanding of endocytosis in cancer. The potential to regulate these pathways in the clinic to improve cancer therapy is also discussed.

Association of a Statewide Opioid Legislation with Opioid Prescribing Patterns in Facial Plastic and Reconstructive Procedures.

Background: The Michigan Opioid Laws are legislation enacted between 2017 and 2018 as a strategy to combat the growing opioid crisis. Objective: To compare opioid prescription rates and morphine milligram equivalents (MMEs) of opioid prescribed to patients undergoing various facial plastic and reconstructive surgery (FPRS) procedures before, during, and after legislation enactment. Materials and Methods: This is a cross-sectional retrospective review of subjects undergoing any of 10 FPRS procedures between July 2016 and November 2019 at a tertiary care hospital with analysis of demographic factors, opioid prescription rates, and MMEs over time. Results: Of 863 patients included, 107 and 575 patients were prescribed postoperative opioids before and after opiate legislation enactment, respectively, with no difference in baseline demographics between groups. Regression analysis showed no change in MME prescribing in the year before legislation (p = 0.70), followed by a decrease of 0.13 MME per day (p = 0.00), with a subsequent stabilization of MME at a reduced rate for the remainder of the study period (p = 0.74). Conclusion: Enactment of the Michigan Opioid Laws was temporally associated with a decrease in opioid prescriptions for common facial plastic surgery procedures.

Expert Consensus on the Use of Teprotumumab for the Management of Thyroid Eye Disease Using a Modified-Delphi Approach.

BACKGROUND: Teprotumumab is the first treatment for thyroid eye disease (TED), a debilitating autoinflammatory condition, approved by the Food and Drug Administration in the United States, which reduces proptosis and improves quality of life. In the absence of guidelines, clinical recommendations were developed for using teprotumumab in patients with TED in the United States. METHODS: A 3-round modified-Delphi panel was conducted between October 2020 and February 2021 with experts in the management of patients with TED. Key areas regarding the use of teprotumumab were investigated, including eligible patient populations, concomitant treatments, and assessment of response and adverse events. This used 2 survey rounds via an online questionnaire, where statements were scored using 9-point Likert scales. Statements with conflict were included in the third round, involving a consensus meeting via videoconference. RESULTS: Consensus was obtained for all statements (n = 75); of which, 56% were revised to enable agreement of the group. The consensus meeting provided agreement regarding which populations should receive teprotumumab therapy, including all adult patients with TED with a clinical activity score of ≥4. Treatment with teprotumumab can also be considered for TED patients displaying the following characteristics: a CAS of <3, lid retraction of ≥2, and mild or early optic neuropathy with close clinical observation. Further recommendations included suitability of treatment for those beyond 16 months following the initial diagnosis of TED, low CAS concomitant treatment with steroids in some cases, retreatment for those who have relapses, and finally a recommendation to continue therapy for all 8 infusions despite the lack of response by the fourth infusion. CONCLUSIONS: This work constitutes the first consensus on guidelines for the use of teprotumumab. The modified Delphi approach involved physicians with significant experience with the clinical use of teprotumumab, and recommendations were based on current evidence.

Inhibition of the master regulator of Listeria monocytogenes virulence enables bacterial clearance from spacious replication vacuoles in infected macrophages.

A hallmark of Listeria (L.) monocytogenes pathogenesis is bacterial escape from maturing entry vacuoles, which is required for rapid bacterial replication in the host cell cytoplasm and cell-to-cell spread. The bacterial transcriptional activator PrfA controls expression of key virulence factors that enable exploitation of this intracellular niche. The transcriptional activity of PrfA within infected host cells is controlled by allosteric coactivation. Inhibitory occupation of the coactivator site has been shown to impair PrfA functions, but consequences of PrfA inhibition for L. monocytogenes infection and pathogenesis are unknown. Here we report the crystal structure of PrfA with a small molecule inhibitor occupying the coactivator site at 2.0 Å resolution. Using molecular imaging and infection studies in macrophages, we demonstrate that PrfA inhibition prevents the vacuolar escape of L. monocytogenes and enables extensive bacterial replication inside spacious vacuoles. In contrast to previously described spacious Listeria-containing vacuoles, which have been implicated in supporting chronic infection, PrfA inhibition facilitated progressive clearance of intracellular L. monocytogenes from spacious vacuoles through lysosomal degradation. Thus, inhibitory occupation of the PrfA coactivator site facilitates formation of a transient intravacuolar L. monocytogenes replication niche that licenses macrophages to effectively eliminate intracellular bacteria. Our findings encourage further exploration of PrfA as a potential target for antimicrobials and highlight that intra-vacuolar residence of L. monocytogenes in macrophages is not inevitably tied to bacterial persistence.

Efficacy and Safety of Teprotumumab in Thyroid Eye Disease.

Thyroid eye disease (TED; also known as thyroid-associated ophthalmopathy) is an autoimmune condition with disabling and disfiguring consequences. Teprotumumab is the first and only medication approved by the United States Food and Drug Administration for the treatment of TED. We review the efficacy and safety of teprotumumab in TED, highlighting results from the 2 randomized, double-masked, placebo-controlled trials. Post-approval case reports of teprotumumab use in patients with compressive optic neuropathy (CON) and inactive TED were similarly favorable to those from the trials. The preliminarily results of teprotumumab for CON and inactive TED should be investigated in formal clinical trials. Teprotumumab should be avoided in pregnancy. Evidence also suggests that teprotumumab may exacerbate pre-existing inflammatory bowel disease, worsen hyperglycemia, and be associated with hearing impairment. Patients at risk for these adverse events need to be closely monitored with baseline and periodic assessments.

Feasibility of a Risk-Based Approach to Cataract Surgery Preoperative Medical Evaluation.

IMPORTANCE: In 2019, the US Centers for Medicare & Medicaid Services implemented the Patients Over Paperwork initiative, allowing hospitals and ambulatory surgery centers to establish their own policies on preoperative history and physical requirements. A risk-based approach to preoperative medical evaluation may allow surgeons to provide high-value patient care. OBJECTIVE: To assess the feasibility of a risk-based approach to cataract surgery preoperative medical evaluation through a lens of safety and throughput. DESIGN, SETTING, AND PARTICIPANTS: A pilot study was performed to evaluate the implementation of a risk-based approach to preoperative medical evaluation for cataract surgery using a virtual medical history questionnaire. The intervention group, seen from June to September 2020, received the risk assessment and those who were low risk proceeded to surgery without further preoperative evaluation prior to the day of surgery. The preintervention control group included patients who received standard care from January to December 2019. MAIN OUTCOMES AND MEASURES: Primary outcomes included rates of intraoperative complications, noneye-related emergency department visits within 7 days, inpatient admissions within 7 days of surgery, case delays, and rates of case cancellation. The secondary outcome included patient perception regarding preoperative care. RESULTS: A total of 1095 patients undergoing cataract surgery were included in the intervention group (1813 [58.2%] female) and 3114 were in the control group (621/1095 [56.7%] female). The mean (SD) age was 68.6 (11.0) in the control group and 68.4 (10.5) in the intervention group. The intervention group included 126 low-risk individuals (11.5%) and 969 individuals who received standard care (88.5%). There were no differences between the control and intervention groups in terms of rates of intraoperative complications (control group vs intervention group: 21 [0.7%] vs 3 [0.3%]; difference, -0.4% [95% CI, -0.82 to 0.02]), 7-day noneye-related ED visits (5 [0.2%] vs 3 [0.3%]; difference, 0.1% [95% CI, -0.23 to 0.45]), 7-day inpatient admissions (6 [0.2%] vs 2 [0.2%]; difference, -0.01% [95% CI, -0.31 to 0.29]), or same-day cancellations (31 [0.8%] vs 10 [0.6%]; difference, -0.15% [95% CI, -0.63 to 0.34]). The control group had more case delays (59 [1.9%] vs 7 [0.6%]; difference, -1.3% [95% CI, -1.93 to -0.58]). CONCLUSIONS AND RELEVANCE: This study suggests that a virtual, risk-based approach to preoperative medical evaluations for cataract surgery is associated with safe and efficient outcomes. These findings may encourage health care systems and ambulatory surgery centers to tailor preoperative requirements for low-risk surgery patients.

Germline ERBB3 mutation in familial non-small-cell lung carcinoma: expanding ErbB's role in oncogenesis.

Lung cancer is the commonest cause of cancer deaths worldwide. Although strongly associated with smoking, predisposition to lung cancer is also heritable, with multiple common risk variants identified. Rarely, dominantly inherited non-small-cell lung cancer (NSCLC) has been reported due to somatic mutations in EGFR/ErbB1 and ERBB2. Germline exome sequencing was performed in a multi-generation family with autosomal dominant NSCLC, including an affected child. Tumour samples were also sequenced. Full-length wild-type (wtErbB3) and mutant ERBB3 (mutErbB3) constructs were transfected into HeLa cells. Protein expression, stability, and subcellular localization were assessed, and cellular proliferation, pAkt/Akt and pERK levels determined. A novel germline variant in ERBB3 (c.1946 T > G: p.Iso649Arg), coding for receptor tyrosine-protein kinase erbB-3 (ErbB3), was identified, with appropriate segregation. There was no loss-of-heterozygosity in tumour samples. Both wtErbB3 and mutErbB3 were stably expressed. MutErbB3-transfected cells demonstrated an increased ratio of the 80 kDa form (which enhances proliferation) compared with the full-length (180 kDa) form. MutErbB3 and wtErbB3 had similar punctate cytoplasmic localization pre- and post-epidermal growth factor stimulation; however, epidermal growth factor receptor (EGFR) levels decreased faster post-stimulation in mutErbB3-transfected cells, suggesting more rapid processing of the mutErbB3/EGFR heterodimer. Cellular proliferation was increased in mutErbB3-transfected cells compared with wtErbB3 transfection. MutErbB3-transfected cells also showed decreased pAkt/tAkt ratios and increased pERK/tERK 30 min post-stimulation compared with wtErbB3 transfection, demonstrating altered signalling pathway activation. Cumulatively, these results support this mutation as tumorogenic. This is the first reported family with a germline ERBB3 mutation causing heritable NSCLC, furthering understanding of the ErbB family pathway in oncogenesis.

Epidermal Growth Factor Receptor Expression and Resistance Patterns to Targeted Therapy in Non-Small Cell Lung Cancer: A Review.

Globally, lung cancer is the leading cause of cancer-related death. The majority of non-small cell lung cancer (NSCLC) tumours express epidermal growth factor receptor (EGFR), which allows for precise and targeted therapy in these patients. The dysregulation of EGFR in solid epithelial cancers has two distinct mechanisms: either a kinase-activating mutation in EGFR (EGFR-mutant) and/or an overexpression of wild-type EGFR (wt-EGFR). The underlying mechanism of EGFR dysregulation influences the efficacy of anti-EGFR therapy as well as the nature of resistance patterns and secondary mutations. This review will critically analyse the mechanisms of EGFR expression in NSCLC, its relevance to currently approved targeted treatment options, and the complex nature of secondary mutations and intrinsic and acquired resistance patterns in NSCLC.

Vismodegib for Preservation of Visual Function in Patients with Advanced Periocular Basal Cell Carcinoma: The VISORB Trial.

BACKGROUND: Basal cell carcinoma (BCC) is a common skin cancer often curable by excision; however, for patients with BCC around the eye, excision places visual organs and function at risk. In this article, we test the hypothesis that use of the hedgehog inhibitor vismodegib will improve vision-related outcomes in patients with orbital and extensive periocular BCC (opBCC). MATERIALS AND METHODS: In this open-label, nonrandomized phase IV trial, we enrolled patients with globe- and lacrimal drainage system-threatening opBCC. To assess visual function in the context of invasive periorbital and lacrimal disease, we used a novel Visual Assessment Weighted Score (VAWS) in addition to standard ophthalmic exams. Primary endpoint was VAWS with a score of 21/50 (or greater) considered successful, signifying globe preservation. Tumor response was evaluated using RECIST v1.1. Surgical specimens were examined histologically by dermatopathologists. RESULTS: In 34 patients with opBCC, mean VAWS was 44/50 at baseline, 46/50 at 3 months, and 47/50 at 12 months or postsurgery. In total, 100% of patients maintained successful VAWS outcome at study endpoint. Compared with baseline, 3% (95% confidence interval [CI], 0.1-15.3) experienced major score decline (5+ points), 14.7% (95% CI, 5 to 31.1) experienced a minor decline (2-4 points), and 79.4% experienced a stable or improved score (95% CI, 62.1-91.3). A total of 56% (19) of patients demonstrated complete tumor regression by physical examination, and 47% (16) had complete regression by MRI/CT. A total of 79.4% (27) of patients underwent surgery, of which 67% (18) had no histologic evidence of disease, 22% (6) had residual disease with clear margins, and 11% (3) had residual disease extending to margins. CONCLUSION: Vismodegib treatment, primary or neoadjuvant, preserves globe and visual function in patients with opBCC. Clinical trail identification number.NCT02436408. IMPLICATIONS FOR PRACTICE: Use of the antihedgehog inhibitor vismodegib resulted in preservation of end-organ function, specifically with regard to preservation of the eye and lacrimal apparatus when treating extensive periocular basal cell carcinoma. Vismodegib as a neoadjuvant also maximized clinical benefit while minimizing toxic side effects. This is the first prospective clinical trial to demonstrate efficacy of neoadjuvant antihedgehog therapy for locally advanced periocular basal cell carcinoma, and the first such trial to demonstrate end-organ preservation.

Change in Postoperative Opioid Prescribing Patterns for Oculoplastic and Orbital Procedures Associated With State Opioid Legislation.

Importance: Understanding whether statewide legislation, such as the Michigan Opioid Laws, is associated with reduction in postoperative opioid prescriptions is informative in guiding future legislation. Objective: To identify changes in opioid prescribing patterns for oculoplastic and orbital procedures associated with the enactment of the Michigan Opioid Laws in 2017 and 2018. Design, Setting, and Participants: This cross-sectional study included 3781 patients who underwent any of 10 common oculoplastic and orbital procedures between June 1, 2016, and November 30, 2019, at a tertiary care institution. Exposures: From 2017 to 2018, Michigan enacted a series of laws to address the state's worsening opioid epidemic. Two major components of this legislation enacted on June 1, 2018, required prescribers to review a report of patients' opioid use history and obtain signed consent after educating patients on the use and disposal of opioids prior to prescribing. Main Outcomes and Measures: Demographic information, type of surgery, type and amount of opioid prescriptions, and morphine milligram equivalent (MME) were analyzed. MME was calculated as the product of dose, quantity, and opioid-specific conversion factor for each prescription. Linear interpolation spline regression was used to evaluate the association of prescription MME with time. Results: Of 3781 patients, 1614 (42.7%) were male. The mean (SD) age at the time of surgery was 63.3 (16.6) years. Of 2026 patients undergoing surgery before June 1, 2018, 1782 (88.0%) were prescribed postoperative opioids; of 1755 patients undergoing surgery after June 1, 2018, 878 (50.0%) were prescribed postoperative opioids (P < .001). There was no difference in age, sex, race/ethnicity, surgery type, or opioids prescribed between these 2 cohorts. Linear interpolation spline regression showed a decrease of 26.025 MMEs (equivalent to a 36.2% reduction of mean MME) between June 1, 2017, and September 30, 2018 (ß, -1.735; 95% CI, -0.088 to -0.024; P < .001), stabilizing at a persistently reduced rate of MME prescribed through the end of the study period (October 1, 2018, to November 30, 2019; ß, -0.005; 95% CI, -0.039 to 0.016; P = .42). Changes in MME in the 12 months before or 12 months after the period of legislation enactment were not identified. Conclusions and Relevance: In this cross-sectional study, reduction in opioid prescriptions for oculoplastic and orbital procedures was observed during the enactment period of the Michigan Opioid Laws and appeared to be sustained through the end of the study period. Similar statewide or national legislations aimed at increasing prescriber awareness and patient education on opioid use may help curtail the prescription opioid epidemic.

Antibody/Ligand-Target Receptor Internalization Assay Protocol Using Fresh Human or Murine Tumor Ex Vivo Samples.

We describe an ex vivo EGF ligand internalization assay using fresh patient tumor biopsies to determine how antigen targets will be trafficked before patients receive mAb treatment. This protocol facilitates a sensitive and reproducible indication as to mAbs surface retention times during treatment. EGF uptake protocols can also be used to analyze EGFR heterogeneity and localization of EGFR in both tumor and xenograft tissue. The technology can be adapted to analyze other receptors such as PD-L1 for which methods are provided. For complete details on the use and execution of this protocol, please refer to Joseph et al. (2019) and Chew et al. (2020).

Endocytosis Inhibition in Humans to Improve Responses to ADCC-Mediating Antibodies.

A safe and controlled manipulation of endocytosis in vivo may have disruptive therapeutic potential. Here, we demonstrate that the anti-emetic/anti-psychotic prochlorperazine can be repurposed to reversibly inhibit the in vivo endocytosis of membrane proteins targeted by therapeutic monoclonal antibodies, as directly demonstrated by our human tumor ex vivo assay. Temporary endocytosis inhibition results in enhanced target availability and improved efficiency of natural killer cell-mediated antibody-dependent cellular cytotoxicity (ADCC), a mediator of clinical responses induced by IgG1 antibodies, demonstrated here for cetuximab, trastuzumab, and avelumab. Extensive analysis of downstream signaling pathways ruled out on-target toxicities. By overcoming the heterogeneity of drug target availability that frequently characterizes poorly responsive or resistant tumors, clinical application of reversible endocytosis inhibition may considerably improve the clinical benefit of ADCC-mediating therapeutic antibodies.

Epidermal Growth Factor Receptor's Function in Cutaneous Squamous Cell Carcinoma and Its Role as a Therapeutic Target in the Age of Immunotherapies.

OPINION STATEMENT: Recent studies have evidenced the potential of combining anti-EGFR therapies with anti-PD-1/PD-L1 checkpoint therapies. Both anti-EGFR and anti-PD-1/PD-L1 have been separately tested in the treatment of cutaneous SCC (cSCC). Here, we review recent data on EGFR in the context of cancer progression, as a prognostic and as a therapeutic target in cSCC. Anti-EGFR/checkpoint immunotherapy and other combination therapy approaches are discussed. With the advent of immunotherapy, EGFR is still a valid cSCC target.

The Use of Umbilical Amnion for Conjunctival Socket, Fornix, and Eyelid Margin Reconstruction.

PURPOSE: To describe outcomes using umbilical amnion for conjunctival fornix, socket, and eyelid margin reconstruction. METHODS: A medical record review was performed to identify patients who had received umbilical amnion over a 2-year period in their department. Patient demographics, disease etiology, and data regarding surgical outcomes were collected. The primary outcome was the success rate of the surgical intervention. RESULTS: Twenty-one patients received umbilical amnion for anophthalmic socket contracture (n = 16), orbital implant exposure (n = 3), ocular surface burn (n = 1), and cicatricial entropion repair (n = 1). The primary outcome was met in 76% of patients overall. In anophthalmic socket contracture, the primary outcome was met in 86% and 0% of patients with acquired and congenital anophthalmia, respectively. The primary outcome was met in all cases of orbital implant exposure and cicatricial entropion. The primary outcome was not met in a Roper-Hall grade IV ocular surface burn. CONCLUSIONS: Umbilical amnion is an ideal substrate graft for reconstruction of the conjunctival fornix, socket, and eyelid margin. Umbilical amnion appears to be efficacious for the management of socket contracture in acquired anophthalmia, orbital implant exposure, and cicatricial entropion. Further experience is needed to determine the efficacy of umbilical amnion in ocular surface burns.