Spike mutation resilient scFv76 antibody counteracts SARS-CoV-2 lung damage upon aerosol delivery.

The uneven worldwide vaccination coverage against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and emergence of variants escaping immunity call for broadly effective and easily deployable therapeutic agents. We have previously described the human single-chain scFv76 antibody, which recognizes SARS-CoV-2 Alpha, Beta, Gamma and Delta variants. We now show that scFv76 also neutralizes the infectivity and fusogenic activity of the Omicron BA.1 and BA.2 variants. Cryoelectron microscopy (cryo-EM) analysis reveals that scFv76 binds to a well-conserved SARS-CoV-2 spike epitope, providing the structural basis for its broad-spectrum activity. We demonstrate that nebulized scFv76 has therapeutic efficacy in a severe hACE2 transgenic mouse model of coronavirus disease 2019 (COVID-19) pneumonia, as shown by body weight and pulmonary viral load data. Counteraction of infection correlates with inhibition of lung inflammation, as observed by histopathology and expression of inflammatory cytokines and chemokines. Biomarkers of pulmonary endothelial damage were also significantly reduced in scFv76-treated mice. The results support use of nebulized scFv76 for COVID-19 induced by any SARS-CoV-2 variants that have emerged so far.

Carnitine palmitoyl transferase-1A (CPT1A): a new tumor specific target in human breast cancer.

Transcriptional mechanisms epigenetically-regulated in tumoral tissues point out new targets for anti-cancer therapies. Carnitine palmitoyl transferase I (CPT1) is the rate-limiting enzyme in the transport of long-chain fatty acids for ß-oxidation. Here we identified the tumor specific nuclear CPT1A as a product of the transcript variant 2, that doesn't retain the classical transferase activity and is strongly involved in the epigenetic regulation of cancer pro-survival, cell death escaping and tumor invasion pathways. The knockdown of CPT1A variant 2 by small interfering RNAs (siRNAs), was sufficient to induce apoptosis in MCF-7, SK-BR3 and MDA-MB-231 breast cancer cells. The cell death triggered by CPT1A silencing correlated with reduction of HDAC activity and histone hyperacetylation. Docking experiments and molecular dynamics simulations confirmed an high binding affinity of the variant 2 for HDAC1. The CPT1A silenced cells showed an up-regulated transcription of pro-apoptotic genes (BAD, CASP9, COL18A1) and down-modulation of invasion and metastasis related-genes (TIMP-1, PDGF-A, SERPINB2). These findings provide evidence of the CPT1 variant 2 involvement in breast cancer survival, cell death escape and invasion. Thus, we propose nuclear CPT1A as a striking tumor specific target for anticancer therapeutics, more selective and effective as compared with the well-known HDAC inhibitors.

Platelet-rich plasma greatly potentiates insulin-induced adipogenic differentiation of human adipose-derived stem cells through a serine/threonine kinase Akt-dependent mechanism and promotes clinical fat graft maintenance.

The potential plasticity and therapeutic utility in tissue regeneration of human adipose-derived stem cells (ASCs) isolated from adult adipose tissue have recently been highlighted. The use of autologous platelet-rich plasma (PRP) represents an alternative strategy in regenerative medicine for the local release of multiple endogenous growth factors. Here we investigated the signaling pathways and effects of PRP and human recombinant insulin on proliferation and adipogenic differentiation of ASCs in vitro. PRP stimulated proliferation (EC(50) = 15.3 ± 1.3% vol/vol), whereas insulin's effect was the opposite (IC(50) = 3.0 ± 0.5 µM). Although PRP alone did not increase adipogenesis, in association with insulin it prevented ASC proliferative arrest, greatly enhanced intracytoplasmic lipid accumulation, strongly increased serine/threonine kinase Akt phosphorylation and mouse monoclonal anti-sterol regulatory element binding protein-1 accumulation, and downregulated Erk-1 activity; adipogenic effects were markedly prevented by the Akt inhibitor wortmannin. PRP with insulin synergistically upregulated fibroblast growth factor receptor (FGFR) and downregulated epidermal growth factor receptor (ErbB) expression; moreover, PRP in association prevented insulin-induced insulin-like growth factor-1 receptor and insulin receptor downregulation. The inhibition of FGFR-1, epidermal growth factor receptor (EGFR), and epidermal growth factor receptor-2 (ErbB2) activity reduced ASC proliferation, but only that of FGFR-1 reduced adipogenesis and Akt phosphorylation, whereas the ErbB2 inhibition effects were the opposite. However, EGFR activity was needed for ErbB2-mediated inhibition of ASC adipogenesis. Clinically, the injection of insulin further ameliorated patients' 1-year PRP-induced fat graft volume maintenance and contour restoring. Our results ascertain that PRP in association with insulin greatly potentiates adipogenesis in human ASCs through a FGFR-1 and ErbB2-regulated Akt mechanism. The ameliorated clinical fat graft maintenance suggests additional useful translational applications of combined PRP-insulin treatment in regenerative medicine.

MiR-203 controls proliferation, migration and invasive potential of prostate cancer cell lines.

Prostate cancers show a slow progression from a local lesion (primary tumor) to a metastatic and hormone-resistant phenotype. After an initial step of hyperplasia, in a high percentage of cases a neoplastic transformation event occurs that, less frequently, is followed by epithelial to mesenchymal transition and invasion of healthy tissues (usually bones). MicroRNA-203 (miR-203) is a tumor suppressor microRNA often silenced in different malignancies. Here, we show that miR-203 is downregulated in clinical primary prostatic tumors compared to normal prostate tissue, and in metastatic prostate cancer cell lines compared to normal epithelial prostatic cells. Overexpression of miR-203 in brain or bone metastatic prostate cell lines (DU145 and PC3) is sufficient to induce a mesenchymal to epithelial transition with inhibition of cell proliferation, migration and invasiveness. We have identified CKAP2, LASP1, BIRC5, WASF1, ASAP1 and RUNX2 as new miR-203 direct target mRNAs involved in these events. Therefore, miR-203 could be a potentially new prognostic marker and therapeutic target in metastatic prostate cancer.

Cardiac sarcomas: an update.

Primary cardiac sarcomas are rare and represent 20% of all primary cardiac tumors. Symptoms depend on the chambers and the cardiac structures involved. Transthoracic echocardiography is commonly used to identify a cardiac mass. The diagnosis of cardiac sarcoma requires adequate sampling and the careful use of ancillary diagnostic techniques. In the most recent histologic classification, angiosarcoma is the most common malignant tumor of the heart with recognizable differentiation. Undifferentiated sarcomas account for one-third of all cardiac sarcomas and have been incorporated in the malignant fibrous histiocytoma/pleomorphic sarcoma subgroup. Elective cardiac sarcoma therapy includes complete surgical excision when possible, followed by radio and chemotherapeutic regimen, the latter preferably containing anthracyclines, ifosfamide, or taxanes. Prognosis of cardiac sarcomas is very poor, with mean survival ranging from 9.6 to 16.5 months. A less-aggressive course seems related to the left atrium location, a low histologic grading with scarce cellular pleomorphism and low-mitotic activity, absence of necrosis, myxoid tumor appearance, and absence of metastasis at diagnosis.

A pathobiologic link between risk factors profile and morphological markers of carotid instability.

OBJECTIVE: Although cardiovascular risk factors have been strongly linked to carotid intimal-media thickness, their association with plaque progression towards instability is poorly understood. We evaluated a large database of endarterectomy specimens removed from symptomatic and asymptomatic patients to determine the correlation between major cardiovascular risk factors and carotid plaque morphology. METHODS: Incidence of thrombotic, vulnerable and stable plaques together with the degree of plaque inflammatory infiltration was evaluated in 457 carotid atherosclerotic lesions. Clinical records were reviewed in all cases for risk factors profile. RESULTS: Thrombotic plaques were more frequently observed in patients affected by stroke (66.9%) as compared to TIA (36.1%) and asymptomatic patients (26.8%, p<0.001). Out of 457 carotid plaques removed during carotid endarterectomy, 181 (39.6%) were represented by thrombotic plaques, 72 (15.8%) by vulnerable plaques (thin cap fibroateroma) and 204 (44.6%) by stable plaques. At the multivariate analysis, a strong association was observed between hypertension, low HDL-cholesterol (HDL-C) and ratio of total to HDL-C >5 with vulnerable and thrombotic carotid plaques. Hypertension (p=0.001), hypercholesterolemia (p=0.05) and low HDL-C (p=0.001) significantly also correlated with the presence of high inflammatory infiltrate of the plaque. When multivariate analysis was restricted to asymptomatic patients, hypertension (p=0.009, OR 2.29), low HDL-cholesterol (p=0.01 OR 2.21) and the ratio of total to HDL-C >5 (p=0.03, OR 2.07) were confirmed to be the risk factors most significantly associated to unstable plaques. The relative risk to carry an unstable plaque for asymptomatic patients with high Framingham Risk Score as compared with those with low risk score was 2.06 (95% C.I., 1.26-3.36). CONCLUSIONS: The present histopathological study identifies risk factors predictive of increased risk of carotid plaque rupture and thrombosis. Asymptomatic patients with high risk factors profile may constitute a specific target to reduce the likelihood of cerebrovascular accidents even in the presence of non-flow-limiting plaque.

Clusterin in stool: a new biomarker for colon cancer screening?

OBJECTIVES: The identification of useful markers for early diagnosis of human colon cancer is a major goal still in progress. Clusterin is a pleiotropic protein with a broad range of functions. It has recently drawn much attention because of its association with cancer promotion and metastasis. It is involved in prosurvival and apoptosis processes that are carried out by two different isoforms. Secreted clusterin isoform (sCLU) is cytoprotective and its prosurvival function is the basis of the current phase I/II clinical trials against prostate, lung, and breast cancers. We have already shown that in colorectal cancer (CRC) there is an increased expression of sCLU. In this report, we investigated whether sCLU is released in the blood and stool of colon cancer patients in order to study sCLU as a potential diagnostic molecular marker for colon cancer screening. METHODS: The quantitative expression of sCLU was determined by dot blot immunodosage in the serum and stool of CRC patients (n=63) and age-matched controls without clinical history of neoplasia, CRC, or systemic or bowel inflammatory disease (n=50). Unpaired t-tests and Mann-Whitney U-tests were used for continuous variables. The diagnostic performance of clusterin was appraised by means of receiver operating characteristic (ROC) curves. RESULTS: We found a significant increase of sCLU in the serum and stool of CRC patients (P=0.0002 and P<0.000, respectively) as compared with controls. ROC curves provided cutoff points showing a trade-off between sensitivity and specificity. With a cutoff point of 88.5 microg/ml, sCLU in blood showed a 55.6% sensitivity and 100% specificity, and with a cutoff point of 34.6 microg/g, the stool test reached 66.7% sensitivity and 84% specificity in discriminating between nonneoplastic and colorectal neoplastic lesions. Human cancer xenografts in nude mice indicated a positive correlation between increasing serum clusterin level and tumor size. CONCLUSIONS: This study highlights the potential of clusterin detection in stool to be a valuable tool to improve the effectiveness and efficiency of large-scale clinical cancer screening.

Comparative clinicopathologic and immunohistochemical analysis of uterine sarcomas diagnosed using the World Health Organization classification system.

Uterine sarcomas are rare tumors that account for 3% to 7% of uterine cancers. Their histopathologic classification was revised by the World Health Organization (WHO) in 2003. The objectives of this study were to determine the frequency of different subtypes of uterine sarcoma applying the WHO criteria to a series of cases, compare the outcome of patients with different subtypes, and compare their immunoprofiles using a panel of immunomarkers. Thirty-four uterine sarcomas were identified for a 20-year period (1988-2008). Eighteen benign tumors of smooth muscle or endometrial stromal origin served as a comparison group. A tissue microarray was prepared and immunostaining performed for 10 selected oncoproteins involved in cell proliferation (Ki-67, P53, p16, and phosphatase and tensin homolog [PTEN]), cell differentiation (CD10, h-caldesmon, estrogen receptor, and progesterone receptor), and apoptosis (bcl-2 and Twist). Hierarchical clustering analysis of the immunohistochemical results was performed. The uterine sarcomas were classified as follows: 20 leiomyosarcomas, 9 endometrial stromal sarcomas, and 5 undifferentiated endometrial sarcomas. The outcome for patients with uterine sarcoma was poor, irrespective of histologic type, even for those with stage I tumors. Of the patients with follow-up available, 12 (67%) of 18 with leiomyosarcoma, 4 of 5 with undifferentiated sarcoma, and 4 of 7 with endometrial stromal sarcoma experienced recurrence and 8 patients with high-grade sarcomas died of tumor. In our series, most uterine sarcomas were leiomyosarcomas. Comparison was made between leiomyosarcomas that recurred and those with a favorable outcome and 3 patients with leiomyosarcoma without evidence of recurrence on long-term follow-up had tumors that were negative/low expressors of Ki-67, p53, p16, and Twist, with strong expression of bcl-2. A subset of undifferentiated endometrial sarcomas composed of cells with uniform nuclei may be a separate entity from those with nuclear anaplasia and may be related to low-grade endometrial stromal sarcomas. It may be possible to identify a subset of leiomyosarcomas with a favorable prognosis based on staining with a panel of immunomarkers for cell proliferation and apoptosis.

Interleukin-6 affects cell death escaping mechanisms acting on Bax-Ku70-Clusterin interactions in human colon cancer progression.

Activation of pro-survival pathways and apoptotic cell death escape are considered hallmarks of oncogenic cell transformation. Tissue microenvironment strongly influences tumorigenesis, redirecting some pathways versus a persisting pro-survival state. Here, we report evidence on the role of interleukin 6 (IL-6) in affecting pro-survival pathways in colon cancer progression, modulating the expression and the molecular interactions among the pro-apoptotic factor Bax, the DNA repair proteins Ku70/86 and Clusterin isoforms. In human colorectal carcinomas (n = 50) at different stages of disease, we found an increased IL-6 production, the loss of Ku86 and Clusterin 50-55 kDa pro-apoptotic isoform. Conversely, we observed the overexpression of Bax and the 40 kDa prosurvival sClusterin (sCLU) isoform. Bax co-localized with Ku70 that was found atypically expressed in the cytoplasm of advanced stage colon cancers (Dukes'C-D; n = 22). IL-6 treatment of a colon cancer cell line, Caco-2, modulated the expression of genes involved in tumor invasion and apoptosis, as observed by microarrays. In particular, IL-6 downmodulated Bax expression at mRNA level. Concomitantly, IL-6 exposure influenced Bax also at protein level acting on the Bax-Ku70-sCLU physical interactions in the cytoplasm, by affecting the Ku70 acetylation and phosphorylation state, thus leading to the inhibition of Bax pro-apoptotic activity. In addition, we found that IL-6 treatment induced a significant downregulation of Ku86 and a strong increase of sCLU, confirming tumor biopsies data. In contrast Somatostatin treatment of Caco-2 cells was able to restore apoptosis, demonstrating that Ku70-Bax-CLU interactions could be dynamically modulated. Hence, IL-6 could favor tumor expansion, promoting cell survival and apoptosis escape throughout the different stages of tumor evolution. Uncovering the molecular mechanisms of action of these factors may offer strategies for selectively manipulate the cancer cells sensitivity to therapy.

Training and mobility: a priority for the Organisation of the European Cancer Institutes. How a national mobility initiative could enhance EU cooperation in cancer research contributing to the development of an European Research Area: the example of The Italian Comprehensive Cancer Centers' Network "Alleanza Contro il Cancro".

It is widely recognized that productivity gains, sustained economic growth and employment are largely determined by technological progress, innovation and human capital. The 2000 Lisbon strategy to make Europe a competitive knowledge-based economy by 2010 and, more specifically, the Barcelona objectives agreed upon in 2002 to increase R&D investment in the EU to approach 3% of GDP, ensuring that there are sufficient human resources for research, are a preliminary step in this direction. If we want to reach this goal we have to succeed in retaining the best researchers, creating the right environment where they can perform their activities and develop their careers. To this aim the Organization of European Cancer Institutes (OECI) has set up a working group on Education and Training with the mandate to encourage continuing education in cancer research and applications and to verify the feasibility to promote mobility programs inside the network and in association with industries. Until now only few OECI training programs have been launched and a full mobility program has not been developed yet due to limited budget resources. The Italian Network of Comprehensive Cancer Centers, Alleanza Contro il Cancro, has planned the launch of a mobility program awarding 70 annual fellowships over a period of 36 months. This program, which will be open to the world research community, could represent a first interaction through mobility among the members of the OECI network also involving industries. The program is a tangible approach to sustain the translational process needed for the development of an European Research Area in the field of cancer and its related biomedical disciplines, thus providing a practical answer to the 2005 renewed Lisbon Strategy.

Fhit interaction with ferredoxin reductase triggers generation of reactive oxygen species and apoptosis of cancer cells.

Fhit protein is lost in most cancers, its restoration suppresses tumorigenicity, and virus-mediated FHIT gene therapy induces apoptosis and suppresses tumors in preclinical models. We have used protein cross-linking and proteomics methods to characterize a Fhit protein complex involved in triggering Fhit-mediated apoptosis. The complex includes Hsp60 and Hsp10 that mediate Fhit stability and may affect import into mitochondria, where it interacts with ferredoxin reductase, responsible for transferring electrons from NADPH to cytochrome P450 via ferredoxin. Viral-mediated Fhit restoration increases production of intracellular reactive oxygen species, followed by increased apoptosis of lung cancer cells under oxidative stress conditions; conversely, Fhit-negative cells escape apoptosis, carrying serious oxidative DNA damage that may contribute to an increased mutation rate. Characterization of Fhit interacting proteins has identified direct effectors of the Fhit-mediated apoptotic pathway that is lost in most cancers through loss of Fhit.

Cellular retinol binding protein-1 expression in endometrial hyperplasia and carcinoma: diagnostic and possible therapeutic implications.

Cellular retinol binding protein-1 (CRBP-1) contributes to the maintenance of the differentiative state of endometrial glandular cells through the regulation of bioavailability of retinol and derivatives, but its role in endometrial oncogenetic process remains unclear. Antibodies to CRBP-1, estrogen and progesterone receptors (ER and PR) were applied to paraffin sections of proliferative (n = 10) and secretory endometrium (n = 9), and to endometrial polyps (n = 6), simple (n = 7), complex (n = 3) and atypical endometrial hyperplasias (n = 9) as well as to 47 endometrioid carcinomas of different histological grade (G) (G1, n = 18; G2, n = 19; G3, n = 10). Four serous and two clear cell carcinomas were also examined. In glandular cells, CRBP-1 positivity was mainly cytoplasmic and rarely in the nuclei. CRBP-1 immunodetection was weakly positive in proliferative and low and focal in secretory endometrium and higher in atypical as compared to simple and complex hyperplasias. CRBP-1 expression in G1 endometrioid carcinomas was similar to that in atypical hyperplasias. In the latter, the highest CRBP-1 expression was observed in areas of squamous differentiation. Semiquantitative evaluation revealed a significant decrease of cytoplasmic CRBP-1 immunoreactivity with the increase of tumor grade. Among G3 endometrioid carcinomas, 60% were CRBP-1 negative, whereas the remaining cases showed a very low and focal positivity. Serous carcinomas were also CRBP-1 negative. When areas of different grading were present within the same tumor, less differentiated areas retained a lower CRBP-1 immunoreaction. The progressive decrease of CRBP-1 paralleled that of ER and PR immunodetection. RT-PCR in eight endometrioid carcinomas suggested a decrease of CRBP-1 with the increase of tumor grade also at transcriptional level. Our results indicate that CRBP-1 immunodetection may constitute an additional tool for histological grading of endometrial carcinoma. The CRBP-1 loss during the progression of endometrial cancer suggests a new potential target for pharmacological strategies aimed to counteract its progression by increased intracellular retinol bioavailability.

99mTc-interleukin-2 scintigraphy for the in vivo imaging of vulnerable atherosclerotic plaques.

PURPOSE: Several histopathological studies have demonstrated that vulnerable plaques are enriched in inflammatory cells. The aims of this study were: (1a) to test the ability of 99mTc-labelled interleukin-2 (99mTc-IL2) to bind to IL2R-positive (IL2R+) cells in carotid plaques and (1b) to correlate the plaque uptake of 99mTc-IL2, measured in vivo, with the number of IL2R+ cells within the plaque, measured ex vivo by histology (transversal study, TS), and (2) to evaluate changes in 99mTc-IL2 uptake in plaques, before and after treatment with a statin or a hypocholesterolaemic diet (longitudinal study, LS). METHODS: Ultrasound scan was performed for plaque characterisation and localisation. Fourteen patients (16 plaques) eligible for endoarterectomy were recruited for the TS and underwent 99mTc-IL2 scintigraphy before surgery. Nine patients (13 plaques) were recruited for the LS; these patients received atorvastatin or a standard hypocholesterolaemic diet and 99mTc-IL2 scintigraphy was performed before and after 3 months of treatment. RESULTS: The degree of 99mTc-IL2 uptake was expressed as the plaque/background (T/B) ratio. In patients from TS, T/B ratios correlated with the percentage of IL2R+ cells at histology (r = 0.707; p = 0.002) and the number of IL2R+ cells at flow cytometry (r = 0.711; p = 0.006). No correlations were observed between ultrasound scores and either scintigraphic or histological findings. In patients from the LS, the mean 99mTc-IL2 uptake decreased in statin-treated patients (1.75+/-0.50 vs 2.16+/-0.44; p = 0.012), while it was unchanged in the patients on the hypocholesterolaemic diet (2.33+/-0.45 vs 2.34+/-0.5). CONCLUSION: 99mTc-IL2 accumulates in vulnerable carotid plaques; this accumulation is correlated with the amount of IL2R+ cells and is influenced by lipid-lowering treatment with a statin.

Widespread myocardial inflammation and infarct-related artery patency.

BACKGROUND: Diffuse coronary vascular inflammation is associated with acute coronary syndromes. However, it is unknown whether inflammation also occurs within the myocardium. Therefore, this study was aimed at assessing the presence of activated cells in unaffected remote myocardium of patients with acute myocardial infarction (AMI), in comparison to the peri-infarct region from the same cases, and in comparison to myocardial specimens from control hearts. METHODS AND RESULTS: Sixteen patients dying 1 to 12 weeks after AMI and 16 control subjects were selected at autopsy. Myocardial specimens were taken at remote unaffected viable regions and at peri-infarct regions in cases with AMI. Confocal microscopy was performed to measure the number of activated cells (DR+), T-lymphocytes (CD3+), and activated T-lymphocytes (CD3+/DR+). Activated cells and activated T-lymphocytes were found in remote unaffected regions in 11 of 16 cases (69%), in peri-infarct zone in all cases (100%), and in none of the control hearts (0%, P<0.001 versus others). A greater myocardial inflammatory burden in remote regions but not in peri-infarct regions was associated with persistent infarct-related artery occlusion (P<0.05). CONCLUSIONS: This study for the first time shows the presence of activated T-lymphocytes in remote unaffected myocardial regions in approximately two thirds of patients with recent AMI. Because these cells are associated with persistent infarct-related artery occlusion, our data may suggest that an antigenic stimulus present also in the myocardium triggers an immune response that may be critical to precipitate artery occlusion.

Sphingosine 1-phosphate induces antimicrobial activity both in vitro and in vivo.

Sphingosine 1-phosphate (S1P), a polar sphingolipid metabolite, is involved in a wide spectrum of biological processes, including Ca(++) mobilization, cell growth, differentiation, motility, and cytoskeleton organization. Here, we show a novel role of S1P in the induction of antimicrobial activity in human macrophages that leads to the intracellular killing of nonpathogenic Mycobacterium smegmatis and pathogenic M. tuberculosis. Such activity is mediated by host phospholipase D, which favors the acidification of mycobacteria-containing phagosomes. Moreover, when it was intravenously injected in mycobacteria-infected mice, S1P reduced mycobacterial growth and pulmonary tissue damage. These results identify S1P as a novel regulator of the host antimicrobial effector pathways.

Metastatic Crohn's disease of the forehead.

BACKGROUND: Metastatic Crohn's disease (CD) involves the presence of cutaneous granuloma distant from the intestinal lesions related to the disease, usually observed in colonic CD. CASE HISTORY: A 35-year-old female with a permanent ileostomy following proctocolectomy for CD presented in 1999 with a 2-month history of an unusual skin lesion of the forehead. A diagnosis of CD of the ileum, colon, and rectum had been made in 1994. In 1997, a proctocolectomy with ileostomy was performed due to fistulizing severe refractory disease. Microscopic aspects of the intestinal lesions showed deep and fissuring ulcers. After surgery, she went into remission, and a small bowel follow-up in 1999 showed no recurrence, when she presented with the skin lesion of the forehead. MICROSCOPIC DATA: Histological analysis of endoscopical and surgical intestinal specimens showed chronic granulomatous inflammation of the ileum, colon, and rectum, confirming the diagnosis of CD. The forehead skin biopsy was examined by three independent histopathologists. The lesion was composed of numerous small granulomas (Ziehl-Nielsen negative), with no foreign bodies, mainly composed of CD68-positive and periodic acid Schiff-negative monocytes. Despite the low number of lymphocytes, the macroscopical and microscopical aspect of the forehead lesion, together with the clinical history, led to a diagnosis of rare metastatic CD of the forehead. CONCLUSIONS: This case report describes the development of an unusual granulomatous skin lesion of the forehead in a patient with established CD showing no postoperative recurrence.