Early Organ Metastasis in Granulomatous Mycosis Fungoides: A Systematic Review.

BACKGROUND: Granulomatous mycosis fungoides (GMF) is a rare form of cutaneous T-cell lymphoma characterized by a granulomatous inflammatory infiltrate. OBJECTIVE: The impact of granulomatous inflammation on the prognosis of the disease remains controversial as there have been both favorable and unfavorable outcomes documented. METHODS: We performed a systematic review of 116 GMF cases previously described in the literature. RESULTS: In contrast to the classic Alibert-Bazin type of mycosis fungoides (MF), cutaneous lesions in GMF tend to involve distal extremities (lower legs, feet, hands) early in the disease course. In the literature, 30% of GMF patients developed organ metastasis, most frequently to the lung. The median time to stage progression was 25 months. CONCLUSION: GMF is an aggressive form of MF. Therefore, screening for distant metastases should be considered at presentation and repeated during follow-up.

Deep learning-enabled realistic virtual histology with ultraviolet photoacoustic remote sensing microscopy.

The goal of oncologic surgeries is complete tumor resection, yet positive margins are frequently found postoperatively using gold standard H&E-stained histology methods. Frozen section analysis is sometimes performed for rapid intraoperative margin evaluation, albeit with known inaccuracies. Here, we introduce a label-free histological imaging method based on an ultraviolet photoacoustic remote sensing and scattering microscope, combined with unsupervised deep learning using a cycle-consistent generative adversarial network for realistic virtual staining. Unstained tissues are scanned at rates of up to 7 mins/cm2, at resolution equivalent to 400x digital histopathology. Quantitative validation suggests strong concordance with conventional histology in benign and malignant prostate and breast tissues. In diagnostic utility studies we demonstrate a mean sensitivity and specificity of 0.96 and 0.91 in breast specimens, and respectively 0.87 and 0.94 in prostate specimens. We also find virtual stain quality is preferred (P = 0.03) compared to frozen section analysis in a blinded survey of pathologists.

Primary testicular lymphoma demonstrates overexpression of the Wilms tumor 1 gene and different mRNA and miRNA expression profiles compared to nodal diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) shows a high degree of clinical and biological heterogeneity. Primary testicular lymphoma (PTL) is an extranodal variant of DLBCL associated with a higher risk of recurrence, including contralateral testicles and central nervous system sanctuary sites. Several molecular aberrations, including somatic mutation of MYD88, CD79B, and upregulation of NF-kB, PDL-1, and PDL-2, are thought to contribute to the pathogenesis and poor prognosis of PTL. However, additional biomarkers are needed that may improve the prognosis and help understand the PTL biology and lead to new therapeutic targets. RNA from diagnostic tissue biopsies of the PTL-ABC subtype and matched nodal DLBCL-ABC subtype patients was evaluated by mRNA and miRNA expression. Screening of 730 essential oncogenic genes was performed, and their epigenetic connections were examined using the nCounter PAN-cancer pathway, and Human miRNA assays with the nCounter System (NanoString Technologies). PTL and nodal DLBCL patients were comparable in age, gender, and putative cell of origin (p > 0.05). Wilms tumor 1 (WT1) expression in PTL exceeded that in nodal DLBCL (>6-fold; p = 0.01, FDR <0.01) and WT1 associated pathway genes THBS4, PTPN5, PLA2G2A, and IFNA17 were upregulated in PTL (>2.0-fold, p < 0.01, FDR <0.01). Additionally, miRNAs targeting WT1 (hsa15a-5p, hsa-miR-16-5p, has-miR-361-5p, has-miR-27b-3p, has-miR-199a-5p, has-miR-199b-5p, has-miR-132-3p, and hsa-miR-128-3p) showed higher expression in PTL compared to nodal DLBCL (≥2.0-fold; FDR 0.01). Lower expression of BMP7, LAMB3, GAS1, MMP7, and LAMC2 (>2.0-fold, p < 0.01) was observed in PTL compared to nodal DLBCL. This research revealed higher WT1 expression in PTL relative to nodal DLBCL, suggesting that a specific miRNA subset may target WT1 expression and impact the PI3k/Akt pathway in PTL. Further investigation is needed to explore WT1's biological role in PTL and its potential as a therapeutic target.

Incidence of ophthalmologic pathology and associated risk factors in orbital fractures at a level I trauma centre.

OBJECTIVE: To estimate the incidence of ophthalmologic pathology at presentation of patients with orbital fracture to a level I trauma centre and the most significant associated risk factors. METHODS: A total of 244 patients with 278 fractured orbits over a 2-year period at a level I trauma centre were reviewed. The primary outcome was the incidence of urgent ophthalmologic pathology, defined as requiring attention without delay. Patient demographics, history, findings on radiographic imaging, and physical examination findings at initial and follow-up examinations were recorded. Odds ratios with 95% confidence intervals were calculated. RESULTS: On initial examination and follow-up, 9.7% of orbits had ophthalmologic pathology. Only 3 patients (1.1%) had urgent pathology, including orbital compartment syndrome and globe rupture, whereas 22 patients (7.9%) had semiurgent pathology and 4 patients (1.4%) had nonurgent pathology. Subjective decreased vision (odds ratio [OR] = 3.5; p = 0.021), assault-related injuries (OR = 2.4; p = 0.036), work-related injuries (OR = 7.7; p = 0.004), afferent pupillary defect (OR = 19.2; p = 0.017), anisocoria (OR = 7.8; p = 0.001), and symmetrical extraocular movement limitation (OR = 5.2; p = 0.003) and fixed pupil (OR = 16.9; p < 0.001) had statistically significant odds ratios associated with pathology. Patient sex, eye involved, intoxication, anticoagulation, and antiplatelets, as well as previous ocular surgery, were not associated with pathology. CONCLUSIONS: Most orbital fractures do not present with ophthalmologic pathology. Subjective vision loss, history of assault or work trauma, and pupil abnormalities on examination were the greatest risk factors for pathology. Our results highlight the most important factors on patient presentation that should prompt first responders to seek urgent ophthalmologic consultation.

Mutations of the DNA repair gene PNKP in a patient with microcephaly, seizures, and developmental delay (MCSZ) presenting with a high-grade brain tumor.

Polynucleotide Kinase-Phosphatase (PNKP) is a bifunctional enzyme that possesses both DNA 3'-phosphatase and DNA 5'-kinase activities, which are required for processing termini of single- and double-strand breaks generated by reactive oxygen species (ROS), ionizing radiation and topoisomerase I poisons. Even though PNKP is central to DNA repair, there have been no reports linking PNKP mutations in a Microcephaly, Seizures, and Developmental Delay (MSCZ) patient to cancer. Here, we characterized the biochemical significance of 2 germ-line point mutations in the PNKP gene of a 3-year old male with MSCZ who presented with a high-grade brain tumor (glioblastoma multiforme) within the cerebellum. Functional and biochemical studies demonstrated these PNKP mutations significantly diminished DNA kinase/phosphatase activities, altered its cellular distribution, caused defective repair of DNA single/double stranded breaks, and were associated with a higher propensity for oncogenic transformation. Our findings indicate that specific PNKP mutations may contribute to tumor initiation within susceptible cells in the CNS by limiting DNA damage repair and increasing rates of spontaneous mutations resulting in pediatric glioma associated driver mutations such as ATRX and TP53.

Fast hybrid optomechanical scanning photoacoustic remote sensing microscopy for virtual histology.

A rapid scanning microscopy method for hematoxylin and eosin (H&E) like images is sought after for interoperative diagnosis of solid tumor margins. The rapid observation and diagnosis of histological samples can greatly lower surgical risk and improve patient outcomes from solid tumor resection surgeries. Photoacoustic remote sensing (PARS) has recently been demonstrated to provide images of virtual H&E stains with excellent concordance with true H&E staining of formalin-fixed, paraffin embedded tissues. By using PARS with constant velocity and 1D galvanometer mirror scanning we acquire large virtual H&E images (10mm x 5mm) of prostate tissue in less than 3.5 minutes without staining, and over two orders of magnitude faster data acquisition than the current PARS imaging speed.

Virtual hematoxylin and eosin histopathology using simultaneous photoacoustic remote sensing and scattering microscopy.

Hematoxylin and eosin (H&E) staining is the gold standard for most histopathological diagnostics but requires lengthy processing times not suitable for point-of-care diagnosis. Here we demonstrate a 266-nm excitation ultraviolet photoacoustic remote sensing (UV-PARS) and 1310-nm microscopy system capable of virtual H&E 3D imaging of tissues. Virtual hematoxylin staining of nuclei is achieved with UV-PARS, while virtual eosin staining is achieved using the already implemented interrogation laser from UV-PARS for scattering contrast. We demonstrate the capabilities of this dual-contrast system for en-face planar and depth-resolved imaging of human tissue samples exhibiting high concordance with H&E staining procedures and confocal fluorescence microscopy. To our knowledge, this is the first microscopy approach capable of depth-resolved imaging of unstained thick tissues with virtual H&E contrast.

The Clinical Spectrum of Primary Cutaneous CD4+ Small/Medium-Sized Pleomorphic T-Cell Lymphoproliferative Disorder: An Updated Systematic Literature Review and Case Series.

BACKGROUND: Primary cutaneous CD4+ small/medium pleomorphic T-cell lymphoproliferative disorder (SMPLPD) is a provisional entity within the 2016 World Health Organization classification of primary cutaneous lymphomas. The condition is currently classified as a lymphoproliferative disorder to emphasize its benign course and discourage aggressive, systemic treatment modalities. OBJECTIVE: To provide a relevant synthesis for the dermatological practitioner on the prevalence, presentation, and treatment of SMPLPD. METHODS: We conducted an updated systematic literature review and a retrospective chart review of diagnosed cases of SMPLPD from 2 Canadian academic cutaneous lymphoma centers. RESULTS: A total of 23 studies with 136 cases were extracted from the systematic review and 24 patients from our retrospective chart review. SMPLPD proved relatively common accounting for 12.5% of all cutaneous T-cell lymphomas encountered in our cutaneous lymphoma clinics, second in frequency only to mycosis fungoides. The typical clinical presentation was that of an older individual (median age 59 years) with an asymptomatic solitary lesion on their upper extremity. The most common clinical differentials were cutaneous lymphoid hyperplasia, basal cell carcinoma, and lymphoma unspecified. T follicular helper markers were reliably detected. The main treatment modalities were surgical excision, local radiation therapy, and topical or intralesional steroids. Cure was achieved in the vast majority of cases. CONCLUSIONS: SMPLPD is an underdiagnosed T-cell lymphoma with an overtly benign clinical course. The condition has an excellent prognosis and responds well to skin-directed therapies. Practitioners should be aware of this condition to avoid aggressive systemic treatments.

Quality of life considerations in uveal melanoma patients: a systematic review.

OBJECTIVE: The aim of this study is to examine the quality of life (QOL) outcomes of patients undergoing different uveal melanoma (UM) treatments and to appraise the literature on the topic. DESIGN AND PARTICIPANTS: A systematic review was conducted to address the study objective. Patients undergoing UM treatment with or without metastasis were eligible for inclusion in this review. METHODS: A literature search was performed using National Library of Medicine (PubMed), Embase, Ovid online, and Cochrane Central Register of Controlled Trials databases. We included all English, original retrospective or prospective studies published between January 1998 to September 2019 in which the primary outcome was the QOL of patients with treated UM. RESULTS: Our search strategy yielded 101 articles. Of these, 18 articles met all our inclusion criteria. The majority of included articles (61%) are cross-sectional studies. On average, each study employed 2 different QOL assessment tools. Overall, physical functioning and mental well-being are impaired in patients with UM after treatment compared with the general population. The severity of the impairment decreases as early as 3 months post-treatment; 8 of 12 studies comparing treatment options reported no statistical difference in physical functioning between treatments; 4 of 12 studies reported better visual function with radiation therapy compared with enucleation, 2 of which described no difference between the 2 options at long-term. Anxiety is more prevalent than depression, and both decrease to less than 10% at 1-year follow-up. CONCLUSIONS: Overall, there is no significant difference in long-term QOL in patients with UM from different treatment groups past 1-year follow-up. This work underscores the need for and importance of developing a standardized, complete assessment tool tailored to the challenges inherent to the diagnosis of UM.

PTHrP, A Biomarker for CNS Metastasis in Triple-Negative Breast Cancer and Selection for Adjuvant Chemotherapy in Node-Negative Disease.

BACKGROUND: Triple-negative breast cancer (TNBC) is characterized by poor prognosis and lack of targeted therapies and biomarkers to guide decisions on adjuvant chemotherapy. Parathyroid hormone-related protein (PTHrP) is frequently overexpressed in breast cancer and involved in proliferation and metastasis, two hallmarks of poor prognosis for node-negative breast cancer. We investigated the prognostic value of PTHrP with respect to organ-specific metastasis and nodal status in TNBC. METHODS: We assessed PTHrP expression using immunohistochemistry in a clinically annotated tissue microarray for a population-based study of 314 patients newly diagnosed with TNBC, then analyzed its correlation to progression and survival using Kaplan-Meier and Cox regression analyses. The Cancer Genome Atlas (TCGA) validation analysis was performed through Bioconductor. All statistical tests were two-sided. RESULTS: PTHrP overexpression (160 of 290 scorable cases, 55.2%) was statistically significantly associated in univariate analysis with decreased overall survival (OS) in our cohort (P = .0055) and The Cancer Genome Atlas (P = .0018) and decreased central nervous system (CNS)-progression-free survival (P = .0029). In multivariate analysis, PTHrP was a statistically significant independent prognostic factor for CNS-progression-free survival in TNBC (hazard ratio [HR] = 5.014, 95% confidence interval [CI] = 1.421 to 17.692, P = .0122) and for OS selectively in node-negative TNBC (HR = 2.423, 95% CI = 1.129 to 5.197, P = .0231). Strikingly, PTHrP emerged as the only statistically significant prognostic factor (HR = 2.576, 95% CI = 1.019 to 6.513, P = .0456) for OS of low-clinical risk node-negative patients who did not receive adjuvant chemotherapy. CONCLUSIONS: PTHrP is a novel independent prognostic factor for CNS metastasis and adjuvant chemotherapy selection of low-clinical risk node-negative TNBC. Its predictive value needs to be prospectively assessed in clinical trials.

Fit-For-Purpose PD-L1 Biomarker Testing For Patient Selection in Immuno-Oncology: Guidelines For Clinical Laboratories From the Canadian Association of Pathologists-Association Canadienne Des Pathologistes (CAP-ACP).

Since 2014, programmed cell death protein 1 (PD-1)/programmed cell death ligand 1 (PD-L1) checkpoint inhibitors have been approved by various regulatory agencies for the treatment of multiple cancers including melanoma, lung cancer, urothelial carcinoma, renal cell carcinoma, head and neck cancer, classical Hodgkin lymphoma, colorectal cancer, gastroesophageal cancer, hepatocellular cancer, and other solid tumors. Of these approved drug/disease combinations, a subset also has regulatory agency-approved, commercially available companion/complementary diagnostic assays that were clinically validated using data from their corresponding clinical trials. The objective of this document is to provide evidence-based guidance to assist clinical laboratories in establishing fit-for-purpose PD-L1 biomarker assays that can accurately identify patients with specific tumor types who may respond to specific approved immuno-oncology therapies targeting the PD-1/PD-L1 checkpoint. These recommendations are issued as 38 Guideline Statements that address (i) assay development for surgical pathology and cytopathology specimens, (ii) reporting elements, and (iii) quality assurance (including validation/verification, internal quality assurance, and external quality assurance). The intent of this work is to provide recommendations that are relevant to any tumor type, are universally applicable and can be implemented by any clinical immunohistochemistry laboratory performing predictive PD-L1 immunohistochemistry testing.

Posterior vitreous detachment and incidence of delayed retinal breaks: a retrospective, 2-year study at an academic centre.

OBJECTIVE: Posterior vitreous detachment (PVD) is common in older patients, particularly postcataract surgery. Patients are typically followed 4 to 6 weeks after the initial presentation. The study goal was to assess the rate of new retinal tears for patients without new symptoms on routine follow-up. METHODS: This is a multisite, retrospective cohort study of all patients presenting to the emergency departments at an academic health centre in a 2-year period diagnosed with uncomplicated PVD. Patient demographics, pastocular surgery, retinal findings at the first and second examinations, and the identity of the examiner were recorded. The primary outcome was absence of new retinal tears or pathology on follow-up. RESULTS: One hundred sixty-six patients were identified, and 105 patients were present for both an initial and follow-up examination (median, 6 weeks). No patients had any new pathology on the follow-up examination. Most patients (n = 62) were seen initially by one of 15 residents alone, and most patients (n = 80) were seen at follow-up by one of 10 staff from specialties of comprehensive, uveitis, neuro-ophthalmology, cornea, and retina. The remaining 25 patients were seen in follow-up by residents. CONCLUSION: To our knowledge, this is the first study on the incidence of delayed retinal breaks with the examination performed by a variety of residents and comprehensive and subspecialist staff. Our results suggest there may be limited benefit to routine follow-up of PVD patients without new symptoms, including when followed by nonretina ophthalmologists. These findings could allow for better health care resources management.

An integrated stress response via PKR suppresses HER2+ cancers and improves trastuzumab therapy.

Trastuzumab is integral to HER2+ cancer treatment, but its therapeutic index is narrowed by the development of resistance. Phosphorylation of the translation initiation factor eIF2α (eIF2α-P) is the nodal point of the integrated stress response, which promotes survival or death in a context-dependent manner. Here, we show an anti-tumor function of the protein kinase PKR and its substrate eIF2α in a mouse HER2+ breast cancer model. The anti-tumor function depends on the transcription factor ATF4, which upregulates the CDK inhibitor P21CIP1 and activates JNK1/2. The PKR/eIF2α-P arm is induced by Trastuzumab in sensitive but not resistant HER2+ breast tumors. Also, eIF2α-P stimulation by the phosphatase inhibitor SAL003 substantially increases Trastuzumab potency in resistant HER2+ breast and gastric tumors. Increased eIF2α-P prognosticates a better response of HER2+ metastatic breast cancer patients to Trastuzumab therapy. Hence, the PKR/eIF2α-P arm antagonizes HER2 tumorigenesis whereas its pharmacological stimulation improves the efficacy of Trastuzumab therapy.

Outcome Analysis of Visual Acuity and Side Effect after Ruthenium-106 Plaque Brachytherapy for Medium-sized Choroidal Melanoma.

PURPOSE: The purpose of this study is to report on treatment outcomes for medium-sized choroidal melanoma treated with Ruthenium-106 (Ru-106) plaque brachytherapy. METHODS: A retrospective case series of 28 patients received Ru-106 brachytherapy treatment for choroidal melanoma. The prescribed tumor dose was 85 Gy to a depth of 5 mm. RESULTS: Median follow-up was 31.2 months. At 12 and 24-month postirradiation, the best corrected visual acuity ≥20/70 (LogMar ≥-0.54) was 53.8% and 64.2%, respectively. Median time to tumor regression was estimated to be 10 months (95% CI = 9-18 months), with 100% of response rate by 32 months. Radiation-induced side effects were limited, and there were no postradiation enucleations. CONCLUSIONS: The majority of patients maintained good visual acuity, with no enucleations and minimal side effects. In this cohort, the Ru-106 plaque brachytherapy proved to be an efficacious and safe treatment option for patients with medium-sized choroidal melanomas with a maximal tumor height of 5 mm.

Autocrine activation of platelet-derived growth factor receptor α in metastatic papillary thyroid cancer.

Metastatic dissemination of papillary thyroid cancer has been reported to be strongly associated with expression of platelet-derived growth factor (PDGFR) α and altered TTF1 function. However, the status of PDGF ligands in papillary thyroid cancer and the potential role of these ligands in metastatic disease are obscure. We assessed the prevalence of PDGF ligands in benign and malignant thyroid tumors to determine if ligand upregulation is associated with α-isoform (PDGF-AA or PDGF-BB) or the ß-isoform (PDGF-BB or PDGF-DD) of PDGFR in individual tumors. The immunohistochemical expression of PDGFRα, PDGF-AA, PDGF-BB, and PDGF-DD was surveyed in follicular adenomas (n=55), papillary thyroid carcinomas (103 with and 59 without nodal metastases), and lymph node metastasis (n=12). There is an augmented tendency for PDGF-AA expression in node-positive papillary thyroid cancer metastases (P<.0001). Although PDGF-BB and -DD were commonly identified, there was no relationship between the presence of these cytokines and malignant disease or metastases. Logistic regression demonstrated that PDGF-AA expression was significantly associated with the presence of PDGFRα (odds ratio=4.6, P=.004) and recurrent disease. When either PDGFRα or PDGF-AA was used to predict the presence of metastases, the sensitivity achieved was 86% and 88%, respectively, whereas specificities were lower at 71% and 61%, respectively. The augmented coexpression of PDGF-AA and PDGFRα in metastatic papillary thyroid cancers suggests that an autocrine signaling loop may contribute to nodal infiltration. Combined testing for the expression of PDGF-AA and PDGFRα may identify those patients with papillary thyroid cancer at risk of metastatic disease and resistance to therapy.

Ocular injury in orbital fractures at a level I trauma center.

OBJECTIVE: To determine the acuity of ophthalmology referral for orbital fractures. METHODS: This is a retrospective chart review of 73 cases of orbital fractures over a 1-year period at a level I trauma centre. All consultations including those for orbital fractures were included in the study. The age and sex of the patient, date and time of initial emergency room (ER) arrival, date and time of input of the ophthalmology consultation, date and time the patient actually saw ophthalmology, reasons for consultation, location of examination, mechanism, and ocular findings at initial and follow-up examination, and any orbital surgery performed were recorded. RESULTS: Of the 73 patients referred, 10 had an ocular abnormality on examination. Of the 65 routine referrals for orbital fracture received, 5 had an ocular abnormality on examination, whereas 5 of the 8 patients referred for an ocular abnormality in addition to the orbital fracture had an ocular abnormality, for an overall rate of 13.7%. Of the routine referrals, none had severe ocular injuries, such as globe rupture or retrobulbar hemorrhage, requiring immediate ocular intervention. CONCLUSIONS: Orbital fractures may be associated with severe ocular abnormalities, although routine cases of orbital fractures in the absence of suspicion of serious ocular injury may be less likely to have abnormalities requiring emergent evaluation by an ophthalmologist. A prospective study evaluating orbital fractures would help clarify this.

Pleomorphic spindle cell sarcoma (PSCS) (formerly known as malignant fibrous histiocytoma, MFH) of the spleen, mimics an atypical haemangioma on 99mTc-RBC, CT and Ultrasound: staging with 18F-FDG PET/CT.

A 63-year-old male was found to have a 7.5-cm splenic mass that had imaging appearances of an atypical haemangioma on CT, ultrasound and a 99mTc-RBC scan, and he was followed conservatively with serial ultrasounds. Sixteen months later, however, the splenic lesion grew and he developed numerous new liver masses which were biopsy confirmed to be a pleomorphic spindle cell sarcoma (PSCS), formerly known as malignant fibrous histiocytoma (MFH). A staging 18F-FDG PET/CT was performed and showed innumerable, mostly necrotic hepatic and splenic masses. The patient passed away a few days after the PET/CT, before a treatment program could be implemented. The use of 18F-FDG PET/CT in the staging of splenic PSCS has not been previously described. We present the 99mTc-RBC and 18F-FDG PET/CT image characteristics of a patient with splenic PSCS.

Cerebral and pulmonary lymphomatoid granulomatosis and EBV positive oesophageal ulcer in an immunosuppressed renal transplant patient staged and followed with serial MRI and 18F-FDG PET/CT after rituximab therapy.

Lymphomatoid granulomatosis is a rare Epstein-Barr virus-related lymphoproliferative disorder. We describe a case of a 42-year-old female with lupus nephritis and immunosuppression post renal transplant, who was diagnosed with central nervous system and lung lymphomatoid granulomatosis, as well as an Epstein-Barr virus-positive oesophageal ulcer, and was staged and followed up long term with multiple 18F-fludeoxyglucose positron emission tomography/CT scans and brain MRIs after achieving a complete metabolic response with rituximab.

Canadian anaplastic lymphoma kinase study: a model for multicenter standardization and optimization of ALK testing in lung cancer.

INTRODUCTION: Fluorescence in situ hybridization (FISH) is currently the standard for diagnosing anaplastic lymphoma kinase (ALK)-rearranged (ALK+) lung cancers for ALK inhibitor therapies. ALK immunohistochemistry (IHC) may serve as a screening and alternative diagnostic method. The Canadian ALK (CALK) study was initiated to implement a multicenter optimization and standardization of laboratory developed ALK IHC and FISH tests across 14 hospitals. METHODS: Twenty-eight lung adenocarcinomas with known ALK status were used as blinded study samples. Thirteen laboratories performed IHC using locally developed staining protocols for 5A4, ALK1, or D5F3 antibodies; results were assessed by H-score. Twelve centers conducted FISH using protocols based on Vysis' ALK break-apart FISH kit. Initial IHC results were used to optimize local IHC protocols, followed by a repeat IHC study to assess the results of standardization. Three laboratories conducted a prospective parallel IHC and FISH analysis on 411 consecutive clinical samples using post-validation optimized assays. RESULTS: Among study samples, FISH demonstrated 22 consensus ALK+ and six ALK wild type tumors. Preoptimization IHC scores from 12 centers with 5A4 and the percent abnormal cells by FISH from 12 centers showed intraclass correlation coefficients of 0.83 and 0.68, respectively. IHC optimization improved the intraclass correlation coefficients to 0.94. Factors affecting FISH scoring and outliers were identified. Post-optimization concurrent IHC/FISH testing in 373 informative cases revealed 100% sensitivity and specificity for IHC versus FISH. CONCLUSIONS: Multicenter standardization study may accelerate the implementation of ALK testing protocols across a country/region. Our data support the use of an appropriately validated IHC assay to screen for ALK+ lung cancers.