IGF1R signaling induces epithelial-mesenchymal plasticity via ITGAV in cutaneous carcinoma.

BACKGROUND: Early cutaneous squamous cell carcinomas (cSCCs) generally show epithelial differentiation features and good prognosis, whereas advanced cSCCs present mesenchymal traits associated with tumor relapse, metastasis, and poor survival. Currently, the mechanisms involved in cSCC progression are unclear, and the established markers are suboptimal for accurately predicting the clinical course of the disease. METHODS: Using a mouse model of cSCC progression, expression microarray analysis, immunofluorescence and flow cytometry assays, we have identified a prognostic biomarker of tumor relapse, which has been evaluated in a cohort of cSCC patient samples. Phosphoproteomic analysis have revealed signaling pathways induced in epithelial plastic cancer cells that promote epithelial-mesenchymal plasticity (EMP) and tumor progression. These pathways have been validated by genetic and pharmacological inhibition assays. RESULTS: We show that the emergence of epithelial cancer cells expressing integrin αV (ITGAV) promotes cSCC progression to a mesenchymal state. Consistently, ITGAV expression allows the identification of patients at risk of cSCC relapse above the currently employed clinical histopathological parameters. We also demonstrate that activation of insulin-like growth factor-1 receptor (IGF1R) pathway in epithelial cancer cells is necessary to induce EMP and mesenchymal state acquisition in response to tumor microenvironment-derived factors, while promoting ITGAV expression. Likewise, ITGAV knockdown in epithelial plastic cancer cells also blocks EMP acquisition, generating epithelial tumors. CONCLUSIONS: Our results demonstrate that ITGAV is a prognostic biomarker of relapse in cSCCs that would allow improved patient stratification. ITGAV also collaborates with IGF1R to induce EMP in epithelial cancer cells and promotes cSCC progression, revealing a potential therapeutic strategy to block the generation of advanced mesenchymal cSCCs.

Transcriptomic subtyping of gastrointestinal malignancies.

Gastrointestinal (GI) cancers are highly heterogeneous at multiple levels. Tumor heterogeneity can be captured by molecular profiling, such as genetic, epigenetic, proteomic, and transcriptomic classification. Transcriptomic subtyping has the advantage of combining genetic and epigenetic information, cancer cell-intrinsic properties, and the tumor microenvironment (TME). Unsupervised transcriptomic subtyping systems of different GI malignancies have gained interest because they reveal shared biological features across cancers and bear prognostic and predictive value. Importantly, transcriptomic subtypes accurately reflect complex phenotypic states varying not only per tumor region, but also throughout disease progression, with consequences for clinical management. Here, we discuss methodologies of transcriptomic subtyping, proposed taxonomies for GI malignancies, and the challenges posed to clinical implementation, highlighting opportunities for future transcriptomic profiling efforts to optimize clinical impact.

Diversity, Prevalence and Virulence of Colletotrichum Species Causing Anthracnose on Cassava Leaves in the Northern Region of Brazil.

Cassava (Manihot esculenta Crantz) is a staple crop widely cultivated by small farmers in tropical countries. However, despite the low level of technology required for its management, it can be affected by several diseases, with anthracnose as the main threat. There is little information about the main species of Colletotrichum that infect cassava in Brazil. Thus, the objective of this work was to study the diversity, prevalence and virulence of Colletotrichum species that cause anthracnose in cassava leaves in northern Brazil. Twenty municipalities of the Pará and Tocantins states were selected, and leaves with symptoms were collected in those locations. Pure cultures were isolated in the laboratory. Species were identified using phylogenetic analyses of multiple loci, and their pathogenicity, aggressivity and virulence levels were assessed. Our results showed the greatest diversity of Colletotrichum associated with anthracnose in cassava plants of the "Formosa" cultivar in the Tocantins and Pará states. We determined the presence of Colletotrichum chrysophilum, C. truncatum, C. siamense, C. fructicola, C. plurivorum, C. musicola and C. karsti, with C. chrysophilum as the most aggressive and virulent. Our findings provide accurate identifications of species of Colletotrichum causing anthracnose in cassava crops, which are of great relevance for cassava breeding programs (e.g., the search for genotypes with polygenic resistance since the pathogen is so diverse) and for developing anthracnose management strategies that can work efficiently against species complexes of Colletotrichum.

Branch retinal artery occlusion in a 49-year-Old woman taking phentermine.

Purpose: This report describes the presentation of a 49-year-old woman with a branch retinal artery occlusion of the right eye in the setting of taking phentermine, a commonly used weight loss medication. Observations: A 49-year-old woman presented with acute painless vision loss in her right eye and was found to have a branch retinal artery occlusion after taking prescribed dosages of phentermine for weight loss therapy. Fundus examination revealed retinal whitening in the distribution of the superior temporal branch retinal artery, and spectral domain optical coherence tomography demonstrated macular edema. Systemic evaluation was negative for cardiovascular, infectious, or autoimmune etiologies. Based on the retinal findings, the patient was diagnosed with phentermine associated branch retinal artery occlusion. She was followed for nine years with no further complications and her vision remained stable in the right eye. Conclusions and Importance: This case highlights that phentermine, a commonly used weight loss medication, could be associated with ischemic retinopathies. Thus, clinicians should be aware that retinal vascular occlusions may not only occur in those who use recreational amphetamines but also in patients taking the prescribed dosages of a weight loss medication like phentermine.

A consensus molecular subtypes classification strategy for clinical colorectal cancer tissues.

Consensus Molecular Subtype (CMS) classification of colorectal cancer (CRC) tissues is complicated by RNA degradation upon formalin-fixed paraffin-embedded (FFPE) preservation. Here, we present an FFPE-curated CMS classifier. The CMSFFPE classifier was developed using genes with a high transcript integrity in FFPE-derived RNA. We evaluated the classification accuracy in two FFPE-RNA datasets with matched fresh-frozen (FF) RNA data, and an FF-derived RNA set. An FFPE-RNA application cohort of metastatic CRC patients was established, partly treated with anti-EGFR therapy. Key characteristics per CMS were assessed. Cross-referenced with matched benchmark FF CMS calls, the CMSFFPE classifier strongly improved classification accuracy in two FFPE datasets compared with the original CMSClassifier (63.6% versus 40.9% and 83.3% versus 66.7%, respectively). We recovered CMS-specific recurrence-free survival patterns (CMS4 versus CMS2: hazard ratio 1.75, 95% CI 1.24-2.46). Key molecular and clinical associations of the CMSs were confirmed. In particular, we demonstrated the predictive value of CMS2 and CMS3 for anti-EGFR therapy response (CMS2&3: odds ratio 5.48, 95% CI 1.10-27.27). The CMSFFPE classifier is an optimized FFPE-curated research tool for CMS classification of clinical CRC samples.

VRK1 Regulates Sensitivity to Oxidative Stress by Altering Histone Epigenetic Modifications and the Nuclear Phosphoproteome in Tumor Cells.

The chromatin organization and its dynamic remodeling determine its accessibility and sensitivity to DNA damage oxidative stress, the main source of endogenous DNA damage. We studied the role of the VRK1 chromatin kinase in the response to oxidative stress. which alters the nuclear pattern of histone epigenetic modifications and phosphoproteome pathways. The early effect of oxidative stress on chromatin was studied by determining the levels of 8-oxoG lesions and the alteration of the epigenetic modification of histones. Oxidative stress caused an accumulation of 8-oxoG DNA lesions that were increased by VRK1 depletion, causing a significant accumulation of DNA strand breaks detected by labeling free 3'-DNA ends. In addition, oxidative stress altered the pattern of chromatin epigenetic marks and the nuclear phosphoproteome pathways that were impaired by VRK1 depletion. Oxidative stress induced the acetylation of H4K16ac and H3K9 and the loss of H3K4me3. The depletion of VRK1 altered all these modifications induced by oxidative stress and resulted in losses of H4K16ac and H3K9ac and increases in the H3K9me3 and H3K4me3 levels. All these changes were induced by the oxidative stress in the epigenetic pattern of histones and impaired by VRK1 depletion, indicating that VRK1 plays a major role in the functional reorganization of chromatin in the response to oxidative stress. The analysis of the nuclear phosphoproteome in response to oxidative stress detected an enrichment of the phosphorylated proteins associated with the chromosome organization and chromatin remodeling pathways, which were significantly decreased by VRK1 depletion. VRK1 depletion alters the histone epigenetic pattern and nuclear phosphoproteome pathways in response to oxidative stress. The enzymes performing post-translational epigenetic modifications are potential targets in synthetic lethality strategies for cancer therapies.

Ocular Dirofilariasis: A Clinicopathologic Case Series and Literature Review.

Purpose: Ocular dirofilariasis is an uncommon zoonotic infection that is usually associated with a carnivore host. In this case series and literature review, we investigate the clinical presentation, management, and histopathology of ocular dirofilariasis. Methods: The database at the Florida Lions Ocular Pathology Laboratory was searched for surgical specimens at the Bascom Palmer Eye Institute under approval of the Institutional Review Board. Patients with a histopathologic diagnosis of dirofilariasis between the years 1962 and 2022 from the Florida Lions Ocular Pathology Laboratory database were included (n = 3). A systematic PubMed search was conducted by two independent authors to identify published cases of ophthalmic dirofilariasis worldwide. Keywords were used to identify articles, and exclusion criteria were applied. Results: Three patients, two males and one female, were identified from the Florida Lions Ocular Pathology Laboratory database with a diagnosis of ocular dirofilariasis. The mean age was 46.7 years (with a range 33-57 years). There were two eyelid lesions (Cases 1 and 3) and one involving the subconjunctival space (Case 2). All three organisms were excised and presumptively identified as Dirofilaria tenuis. All 3 patients were managed with curative surgical removal and recovered completely. Our review of the literature identified 540 published reports and 142 published reports with 186 cases that met the exclusion criteria. Conclusion: We present a case series and literature review of ocular dirofilariasis. Knowledge of the incidence, risk factors, prevention, and diagnosis of this unique parasitic infection will help in proper management and prevent further ocular complications.

Faricimab in Previously Treated Eyes With Neovascular Age-Related Macular Degeneration: An Assessment of Durability and Treatment Outcomes.

BACKGROUND AND OBJECTIVE: This study evaluated the efficacy and durability of faricimab in patients with neovascular age-related macular degeneration (nAMD) who were previously treated with anti-vascular endothelial growth factor (anti-VEGF) agents. PATIENTS AND METHODS: This retrospective case series was conducted at a single tertiary center in the United States. It focused on nAMD patients who transitioned to faricimab after initial anti-VEGF therapy, with a follow-up period of at least 9 months. "Complete dryness" was defined as the absence of intra- and/or subretinal fluid on optical coherence tomography. Durability was gauged by the extension of treatment intervals relative to the injection frequency of the previous agent. RESULTS: Sixty-two eyes from 62 patients were included. Treatment interval ranged from 5 to 10 weeks; 10 (16%) patients were able to be extended by 2 or more weeks compared to their previous regimen. Median (interquartile range [IQR]) central field thickness was 310 µm (254, 376) on initiating faricimab and declined by the ninth month (P values at 3, 6, and 9 months were 0.01, 0.02, and 0.07, respectively). Median (IQR) visual acuity at initiation of faricimab was 0.4 (0.20, 0.50) and did not change by the ninth month. Complete anatomical dryness was present in 10 (16%) eyes before switching; 90% remained dry at 9 months. Of 52 (84%) incompletely dry eyes before switching, 15% achieved complete dryness by 9 months on faricimab. CONCLUSIONS: Faricimab modestly improved the treatment intervals for a small proportion of previously treated patients on anti-VEGF therapy. [Ophthalmic Surg Lasers Imaging Retina 2024;55:XX-XX.].

Protein homeostasis maintained by HOOK1 levels promotes the tumorigenic and stemness properties of ovarian cancer cells through reticulum stress and autophagy.

BACKGROUND: Ovarian cancer has a high mortality rate mainly due to its resistance to currently used therapies. This resistance has been associated with the presence of cancer stem cells (CSCs), interactions with the microenvironment, and intratumoral heterogeneity. Therefore, the search for new therapeutic targets, particularly those targeting CSCs, is important for improving patient prognosis. HOOK1 has been found to be transcriptionally altered in a substantial percentage of ovarian tumors, but its role in tumor initiation and development is still not fully understood. METHODS: The downregulation of HOOK1 was performed in ovarian cancer cell lines using CRISPR/Cas9 technology, followed by growth in vitro and in vivo assays. Subsequently, migration (Boyden chamber), cell death (Western-Blot and flow cytometry) and stemness properties (clonal heterogeneity analysis, tumorspheres assay and flow cytometry) of the downregulated cell lines were analysed. To gain insights into the specific mechanisms of action of HOOK1 in ovarian cancer, a proteomic analysis was performed, followed by Western-blot and cytotoxicity assays to confirm the results found within the mass spectrometry. Immunofluorescence staining, Western-blotting and flow cytometry were also employed to finish uncovering the role of HOOK1 in ovarian cancer. RESULTS: In this study, we observed that reducing the levels of HOOK1 in ovarian cancer cells reduced in vitro growth and migration and prevented tumor formation in vivo. Furthermore, HOOK1 reduction led to a decrease in stem-like capabilities in these cells, which, however, did not seem related to the expression of genes traditionally associated with this phenotype. A proteome study, along with other analysis, showed that the downregulation of HOOK1 also induced an increase in endoplasmic reticulum stress levels in these cells. Finally, the decrease in stem-like properties observed in cells with downregulated HOOK1 could be explained by an increase in cell death in the CSC population within the culture due to endoplasmic reticulum stress by the unfolded protein response. CONCLUSION: HOOK1 contributes to maintaining the tumorigenic and stemness properties of ovarian cancer cells by preserving protein homeostasis and could be considered an alternative therapeutic target, especially in combination with inducers of endoplasmic reticulum or proteotoxic stress such as proteasome inhibitors.

Exploiting a subtype-specific mitochondrial vulnerability for successful treatment of colorectal peritoneal metastases.

Peritoneal metastases (PMs) from colorectal cancer (CRC) respond poorly to treatment and are associated with unfavorable prognosis. For example, the addition of hyperthermic intraperitoneal chemotherapy (HIPEC) to cytoreductive surgery in resectable patients shows limited benefit, and novel treatments are urgently needed. The majority of CRC-PMs represent the CMS4 molecular subtype of CRC, and here we queried the vulnerabilities of this subtype in pharmacogenomic databases to identify novel therapies. This reveals the copper ionophore elesclomol (ES) as highly effective against CRC-PMs. ES exhibits rapid cytotoxicity against CMS4 cells by targeting mitochondria. We find that a markedly reduced mitochondrial content in CMS4 cells explains their vulnerability to ES. ES demonstrates efficacy in preclinical models of PMs, including CRC-PMs and ovarian cancer organoids, mouse models, and a HIPEC rat model of PMs. The above proposes ES as a promising candidate for the local treatment of CRC-PMs, with broader implications for other PM-prone cancers.

Phacolytic Glaucoma Diagnosed by Cytopathology: A Clinicopathologic Case Series.

PURPOSE: The aim of this study was to describe the clinical presentation, histopathologic characteristics, and management of phacolytic glaucoma. PATIENTS AND METHODS: The database at the Florida Lions Ocular Pathology Laboratory was searched for surgical specimens at the Bascom Palmer Eye Institute. Patients with a diagnosis of "phacolytic glaucoma" on cytopathology between the years of 1997 and 2023 were included (n = 15). Patient demographics, anatomic site, laterality, clinical features, and ocular examination findings were obtained from available medical records. RESULTS: The diagnosis of phacolytic glaucoma was established by light microscopic examination of cytology specimens in 15 eyes (Right = 8, Left = 6, not specified = 1). Between 1997 and 2023, there were 32 cytopathology cases with a preoperative diagnosis of phacolytic glaucoma. From this group of 15 cases, there were nine males and six females. Mean age was 64.2 years (Range: 39-87). Thirteen samples were from the anterior chamber and two were obtained from the vitreous. All 15 cases (100%) demonstrated histiocytes with engulfed foamy and/or granular presumed lenticular material. CD68 immunohistochemistry was positive within histiocytes in four cases that were stained (100%). Of the available medical records, clinical features, and ocular examination findings included: eye redness, decreased vision, eye pain, anterior chamber inflammation, dislocated or subluxed crystalline lens, cataract, and elevated intraocular pressure (average = 41.3 ± 8.67 mm Hg). CONCLUSIONS: Phacolytic glaucoma is a rare complication of mature/hypermature cataracts that presents with ocular pain, decreased vision, and anterior chamber inflammation. AC paracentesis with cytopathologic evaluation is a minimally invasive, rapid technique that can aid in the diagnosis and management of this disease.

Kinase activities in pancreatic ductal adenocarcinoma with prognostic and therapeutic avenues.

Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with a limited number of known driver mutations but considerable cancer cell heterogeneity. Phosphoproteomics provides a direct read-out of aberrant signaling and the resultant clinically relevant phenotype. Mass spectrometry (MS)-based proteomics and phosphoproteomics were applied to 42 PDAC tumors. Data encompassed over 19 936 phosphoserine or phosphothreonine (pS/T; in 5412 phosphoproteins) and 1208 phosphotyrosine (pY; in 501 phosphoproteins) sites and a total of 3756 proteins. Proteome data identified three distinct subtypes with tumor intrinsic and stromal features. Subsequently, three phospho-subtypes were apparent: two tumor intrinsic (Phos1/2) and one stromal (Phos3), resembling known PDAC molecular subtypes. Kinase activity was analyzed by the Integrative iNferred Kinase Activity (INKA) scoring. Phospho-subtypes displayed differential phosphorylation signals and kinase activity, such as FGR and GSK3 activation in Phos1, SRC kinase family and EPHA2 in Phos2, and EGFR, INSR, MET, ABL1, HIPK1, JAK, and PRKCD in Phos3. Kinase activity analysis of an external PDAC cohort supported our findings and underscored the importance of PI3K/AKT and ERK pathways, among others. Interestingly, unfavorable patient prognosis correlated with higher RTK, PAK2, STK10, and CDK7 activity and high proliferation, whereas long survival was associated with MYLK and PTK6 activity, which was previously unknown. Subtype-associated activity profiles can guide therapeutic combination approaches in tumor and stroma-enriched tissues, and emphasize the critical role of parallel signaling pathways. In addition, kinase activity profiling identifies potential disease markers with prognostic significance.

Therapeutic Response to Treatment of a Papillomatous Ocular Surface Squamous Neoplasia With Intramuscular Human Papillomavirus Vaccine.

PURPOSE: The purpose of this study was to describe the response of a papillomatous ocular surface squamous neoplasia (OSSN) to the intramuscular (IM) 9-valent human papillomavirus (HPV) vaccine after failed medical and surgical interventions. METHODS: A 79-year-old White man with a conjunctival lesion underwent a biopsy which revealed OSSN and positivity for high-risk HPV. Initially treated with medical therapy and surgical excisions, the patient developed a recurrence and refused further surgery. He was given 4 doses of IM HPV vaccine at the 6-week interval. RESULTS: A dramatic reduction in lesion size and reduced epithelial thickening and hyperreflectivity was noted on slitlamp examination and high-resolution anterior segment optical coherence tomography after receiving the IM HPV vaccine. Although lesion size was markedly reduced, the therapy did not achieve total resolution, resulting in further treatment with topical 1% 5-fluorouracil (5-FU) eye drops and later 0.04% mitomycin C eye drops. The patient then elected to discontinue further treatment and solely observe. CONCLUSIONS: This case report adds to the growing literature demonstrating the potential therapeutic use of vaccines in cancer treatment. Although HPV vaccination is currently approved for prophylaxis, the use of HPV vaccines as a therapeutic option for various HPV-mediated diseases, including OSSN, should be further explored. The HPV vaccine yielded significant initial improvement in this patient who refused further surgical interventions. The use of IM HPV vaccine as an adjunctive treatment of papillomatous OSSN may represent a potential therapeutic option in cases refractory to standard treatment modalities.

A Comparative Biomechanical Study of Alternative Medial Collateral Ligament Reconstruction Techniques.

BACKGROUND: There is little evidence of the biomechanical performance of medial collateral ligament (MCL) reconstructions for restoring stability to the MCL-deficient knee regarding valgus, external rotation (ER), and anteromedial rotatory instability (AMRI). HYPOTHESIS: A short isometric reconstruction will better restore stability than a longer superficial MCL (sMCL) reconstruction, and an additional deep MCL (dMCL) graft will better control ER and AMRI than single-strand reconstructions. STUDY DESIGN: Controlled laboratory study. METHODS: Nine cadaveric human knees were tested in a kinematics rig that allowed tibial loading while the knee was flexed-extended 0° to 100°. Optical markers were placed on the femur and tibia and displacements were measured using a stereo camera system. The knee was tested intact, and then after MCL (sMCL + dMCL) transection, and loaded in anterior tibial translation (ATT), ER, varus-valgus, and combined ATT + ER (AMRI loading). Five different isometric MCL reconstructions were tested: isolated long sMCL, a short construct, each with and without dMCL addition, and isolated dMCL reconstruction, using an 8 mm-wide synthetic graft. RESULTS: MCL deficiency caused an increase in ER of 4° at 0° of flexion (P = .271) up to 14° at 100° of flexion (P = .002), and valgus laxity increased by 5° to 8° between 0° and 100° of flexion (P < .024 at 0°-90°). ATT did not increase significantly in isolated MCL deficiency (P > .999). All 5 reconstructions restored native stability across the arc of flexion apart from the isolated long sMCL, which demonstrated residual ER instability (P≤ .047 vs other reconstructions). CONCLUSION: All tested techniques apart from the isolated long sMCL graft are satisfactory in the context of restoring the valgus, ER, and AMRI stability to the MCL-deficient knee in a cadaveric model. CLINICAL RELEVANCE: Contemporary MCL reconstruction techniques fail to control ER and therefore AMRI as they use a long sMCL graft and do not address the dMCL. This study compares 5 MCL reconstruction techniques. Both long and short isometric constructs other than the long sMCL achieved native stability in valgus and ER/AMRI. Double-strand reconstructions (sMCL + dMCL) tended to provide more stability. This study shows which reconstructions demonstrate the best biomechanical performance, informs surgical reconstruction techniques for AMRI, and questions the efficacy of current popular techniques.

LaSap vaccine: Immunotherapy and immunochemotherapy associated with allopurinol in dogs naturally infected with Leishmania infantum.

Canine visceral leishmaniasis is a parasitic zoonosis that has a profound impact on public health in countries where it is endemic. Chemotherapeutic treatments cannot keep dogs stable for long periods, and the risk of generating parasitic resistance must be considered. Forty-four symptomatic and naturally infected dogs with Leishmania infantum were tested with two treatment protocols (i) immunotherapy with LaSap vaccine and (ii) immunochemotherapy with LaSap vaccine plus allopurinol. At 90 days after the end of the treatment, it was verified that, although both protocols had generated significant clinical improvements with a greater production of IFN-γ/IL-10, in relation to the parasite load, mainly in the skin, the dogs treated only with immunotherapy maintained the same profile. These results indicate that LaSap is a good strategy to control dog parasitism.

Protein kinase D drives the secretion of invasion mediators in triple-negative breast cancer cell lines.

The protein kinase D (PKD) family members regulate the fission of cargo vesicles at the Golgi complex and play a pro-oncogenic role in triple-negative breast cancer (TNBC). Whether PKD facilitates the secretion of tumor-promoting factors in TNBC, however, is still unknown. Using the pharmacological inhibition of PKD activity and siRNA-mediated depletion of PKD2 and PKD3, we identified the PKD-dependent secretome of the TNBC cell lines MDA-MB-231 and MDA-MB-468. Mass spectrometry-based proteomics and antibody-based assays revealed a significant downregulation of extracellular matrix related proteins and pro-invasive factors such as LIF, MMP-1, MMP-13, IL-11, M-CSF and GM-CSF in PKD-perturbed cells. Notably, secretion of these proteins in MDA-MB-231 cells was predominantly controlled by PKD2 and enhanced spheroid invasion. Consistently, PKD-dependent secretion of pro-invasive factors was more pronounced in metastatic TNBC cell lines. Our study thus uncovers a novel role of PKD2 in releasing a pro-invasive secretome.