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BACKGROUND AND AIMS Complete loss of histone H3 lysine 27 trimethylation (H3K27me3) has recently emerged as a biomarker for malignant peripheral nerve sheath tumours (MPNST). Loss of H3K27me3 staining has also been reported in post-radiation MPNST; however, it has not been evaluated in a large series of radiation-associated sarcomas of different histological subtypes. The aim of this study was to assess H3K27me3 labelling by immunohistochemistry in radiation-associated sarcomas and to determine the prevalence of H3K27me3 loss in these tumours. METHODS AND RESULTS Radiation-associated sarcomas (n = 119) from two tertiary care referral centres were evaluated for loss of H3K27me3, defined as complete loss of staining within tumour cells in the presence of a positive internal control. Twenty-three cases (19%) showed H3K27me3 loss, including nine of 10 (90%) MPNST, seven of 77 (9%) undifferentiated spindle cell/pleomorphic sarcomas, five of 25 (20%) angiosarcomas, one of five (20%) leiomyosarcomas and one of two (50%) osteosarcomas. CONCLUSIONS Complete H3K27me3 loss was present in 19% of radiation-associated sarcomas in our series. Our findings demonstrate that loss of H3K27me3 is not specific for radiation-associated MPNST and may also occur in other histological subtypes of RAS, including radiation-associated undifferentiated spindle cell/pleomorphic sarcoma, angiosarcoma, leiomyosarcoma and osteosarcoma.

Aims MYC is a proto-oncogene that is frequently dysregulated in various malignancies, through translocation or amplification. Radiation-associated angiosarcoma frequently shows MYC amplification, and immunohistochemical expression has been shown to be a reliable surrogate marker for amplification, but less is known about MYC expression in other sarcoma types, despite reports of MYC amplification in some undifferentiated/unclassified radiation-associated sarcomas (RASs). Distinguishing putative RAS from non-radiation-associated sarcoma or sarcomatoid carcinoma can be difficult. The aim of this study was to determine the prevalence and potential diagnostic utility of MYC in this context, by evaluating MYC expression in a cohort of RASs, non-radiation-associated sarcomas, and sarcomatoid carcinomas. Methods and results Three hundred and eighty-five neoplasms were evaluated, including 81 RASs (18 angiosarcomas; 57 undifferentiated sarcomas; three leiomyosarcomas; and three malignant peripheral nerve sheath tumours), 267 non-radiation-associated sarcomas, and 37 sarcomatoid carcinomas. Immunohistochemistry was performed with a monoclonal anti-MYC antibody. Staining in tumour cells was scored on the basis of extent (focal, 1-4%; multifocal, 5-49%; and diffuse, ≥50%) and intensity (strong, moderate, and weak). One hundred percent of radiation-associated angiosarcomas expressed MYC diffusely. Expression was infrequent among other types of RAS (9.5%), and the frequency was similar to that in non-radiation-associated sarcomas (9.7%). MYC expression was more common in sarcomatoid carcinomas, occurring in 43%. The extent and intensity of staining were variable in all groups. Conclusion MYC expression is infrequent among RASs other than angiosarcoma, and has a similar prevalence in sporadic sarcomas. Given the frequency of expression in sarcomatoid carcinomas, MYC expression outside the context of radiation-associated angiosarcoma is of limited diagnostic utility, and should be interpreted with caution after exclusion of sarcomatoid carcinoma where relevant.

PURPOSE Radiation-associated sarcomas (RAS) are considered to have a poor prognosis. Although the incidence is anticipated to rise, contemporary data regarding predictors of outcomes are few. We performed a retrospective analysis to identify RAS prognostic factors and subset analyses for radiation-associated angiosarcoma arising after treatment for breast cancer (RAAB) and other RAS subtypes (other-RAS). METHODS AND MATERIALS Patients with localized RAS evaluated at an institutional multidisciplinary sarcoma clinic were identified. Clinical and histologic review was performed, and outcomes were assessed to identify prognostic features. A subset of cases underwent molecular analysis by next-generation sequencing. RESULTS Among 176 patients, histologic subtypes of RAS included angiosarcoma (41%), undifferentiated/unclassified sarcoma (40%), leiomyosarcoma (8%), malignant peripheral nerve sheath tumor (6%), and osteosarcoma (2%). Sixty-seven patients (38%) had RAAB, and 109 (62%) had other-RAS. RAAB had significantly shorter latency from time of initial radiation compared with other-RAS (8 vs. 15 years; P

Approximately half of all cancer patients receive radiation therapy as part of their oncologic treatment. Radiation-associated sarcomas occur in fewer than 1% of patients who receive radiation therapy but account for up to 5% of all sarcomas. As the use of radiation has increased in the past few decades and overall oncologic outcomes are improving, the incidence of radiation-associated sarcomas is also expected to increase. Historically, radiation-associated sarcomas have been associated with poor outcomes but recent data suggest the prognosis is improving. Distinguishing the sarcoma from the primary malignancy is a major diagnostic criterion.

Background The characterization of poorly differentiated neoplasms in fine-needle aspiration (FNA) and small biopsy specimens usually requires immunohistochemistry (IHC) with a panel of markers. Because of an increasing need to preserve limited diagnostic material for tumor genotyping and a mounting demand for cost containment, the authors investigated the usefulness of dual-color IHC with antibodies directed against broad-spectrum keratins and SOX10, a neuroectodermal transcription factor consistently expressed in melanoma, in the workup of epithelioid malignant neoplasms. Methods A total of 107 cases of FNA cell blocks (49 cases) and small biopsies (58 cases) were selected, including 34 melanomas, 31 epithelioid/pleomorphic sarcomas, and 42 carcinomas. IHC was performed on all specimens using a peroxidase-based brown chromogen for SOX10 and an alkaline phosphatase-based red chromogen for keratins AE1/AE3. The presence or absence of staining in lesional cells was scored. Results The majority of tumors demonstrated 1 of 3 distinct patterns: 1) malignant melanomas with nuclear SOX10 (sensitivity of 94% and specificity of 95%); 2) epithelioid/pleomorphic sarcomas negative for both SOX10 and AE1/AE3 (sensitivity of 84% and specificity of 88%); and 3) carcinomas with cytoplasmic AE1/AE3 (sensitivity of 76% and specificity of 98%). In addition, a fourth pattern with cytoplasmic AE1/AE3 and nuclear SOX10 was observed in a subset of carcinomas, most notably triple-negative breast cancers. Conclusions SOX10/keratin dual-color IHC appears to be an effective, sensitive, and specific test to distinguish between melanoma, sarcoma, and carcinoma. This approach can identify melanoma, prioritize additional studies, and limit the number of markers needed to workup an epithelioid malignant neoplasm, thereby potentially reducing costs and preserving valuable tissue for ancillary studies with which to guide therapy. Cancer Cytopathol 2018;126:179-89. © 2018 American Cancer Society.

BACKGROUND The reclassification of noninvasive follicular variant of papillary thyroid carcinoma as noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) has created diagnostic and management issues for thyroid fine-needle aspiration (FNA). In response to these challenges, the authors' laboratory adopted a NIFTP policy including 1) stringent criteria (requiring pseudo-inclusions, papillae, and/or psammoma bodies) for a malignant diagnosis of papillary carcinoma to limit false-positive results due to NIFTP and 2) the use of explanatory notes in cases with cytomorphologic features suggestive of possible NIFTP to encourage lobectomy over thyroidectomy. This study examined the effects of this policy on FNA classification and subsequent surgical management. METHODS All thyroid FNAs performed at Brigham and Women's Hospital (n = 1300) during a 1-year period were evaluated for changes in the use of diagnostic categories, explanatory NIFTP notes, and surgical follow-up in comparison with historical controls. RESULTS The use of specific Bethesda categories did not significantly change. Only a single case of NIFTP was mistakenly classified as malignant. NIFTP was seldom suspected prospectively (17 of 1300; 1.3%); when NIFTP was suspected, cases were reported as suspicious for a follicular neoplasm/follicular neoplasm (n = 10) or suspicious for malignancy (SUS; n = 7). Five of the 7 SUS cases (71%) underwent partial thyroidectomy, compared to 19% of those classified as SUS without an explanatory NIFTP note (P

Purpose: Recurrent or metastatic adenoid cystic carcinoma (R/M ACC) is a rare cancer often arising from the salivary glands of the head and neck. While effective therapies are lacking, preclinical models have suggested that retinoic acid agonists may inhibit ACC growth by blocking MYB binding at translocated gene enhancers. We conducted a phase II trial evaluating retinoic acid as a potential novel therapy for R/M ACC. Patients and methods: Patients with histologically confirmed R/M ACC of any primary site with measurable disease (RECIST v1.1), clinical or radiographic progression within 12 months prior to enrollment, and any number of prior lines of therapy were eligible. Cohort 1 (CH1) received ATRA 45 mg/m2 split oral daily dosing on days 1-14 of a 28-day cycle; cohort 2 (CH2) received the same dosing continuously; treatment continued until disease progression. The primary endpoint was best overall response: ≥5 patients in CH1 with disease in response (CR+PR) among N=25 (assuming ≥2 of N=14 accrued in first stage of a two-stage design had disease in response) provided 85% power to target a >10% response rate (one-sided 9% binomial test). Secondary endpoints: safety, progression-free survival (PFS), overall survival. Exploratory analyses: ATRA impact on MYB expression, define resistance mechanisms, and monitor circulating tumor DNA. All had targeted tumor sequencing prior to enrollment. Results: Between 8/2019 and 2/2020, N=14 enrolled in stage 1 CH1. Primary endpoint of response to continue accrual into stage 2 in CH1 was not met; by 5/2020, N=4 enrolled in CH2 when the trial closed to accrual (budget constraints). Among 18 patients, median age: 58, 61% (11/18) women; each patient had a median of 3 organs (range, 1-4) with metastatic disease. 39% had 2+ prior lines of therapy. Best overall response: CR+PR 0%; SD 61% (11/18); PD 28% (5/18); unevaluable 11% (2/18). Median duration of stability 3.5 months (range, 1-12.3+). One patient (CH1) remains on drug with SD >1 year. Half of those who received prior VEGFR therapy achieved SD (4/8). At a median follow-up of 7.9 months, median PFS was 3.2 months (95% CI, 1.8-3.9). N=1 required dose adjustment; N=1 came off drug for toxicity. There were no grade 3-4 adverse events (headache, dry skin were common); no deaths due to treatment. Median tumor mutational burden was 0 (range, 0-5). NOTCH1 was the most frequent alteration (4, 22%) with 2 evaluable NOTCH1-mutant patients exhibiting SD. Mutations in the PI3K pathway, TP53, and TERT promoter were common. Low MYB protein expression was associated with longer duration of stability (p Citation Format: Glenn J. Hanna, Anne O'Neill, Jennifer M. Cutler, Michelle Flynn, Tushara Vijaykumar, John R. Clark, Lori J. Wirth, Jochen H. Lorch, Jong C. Park, Jeffrey Mito, Jens G. Lohr, Jeffrey Kaufman, Leonard I. Zon, Robert I. Haddad. A phase II trial of all-trans retinoic acid (ATRA) in advanced adenoid cystic carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr CT165.

The goal of resection of soft tissue sarcomas located in the extremity is to preserve limb function while completely excising the tumor with a margin of normal tissue. With surgery alone, one-third of patients with soft tissue sarcoma of the extremity will have local recurrence due to microscopic residual disease in the tumor bed. Currently, a limited number of intraoperative pathology-based techniques are used to assess margin status; however, few have been widely adopted due to sampling error and time constraints. To aid in intraoperative diagnosis, we developed a quantitative optical microscopy toolbox, which includes acriflavine staining, fluorescence microscopy, and analytic techniques called sparse component analysis and circle transform to yield quantitative diagnosis of tumor margins. A series of variables were quantified from images of resected primary sarcomas and used to optimize a multivariate model. The sensitivity and specificity for differentiating positive from negative ex vivo resected tumor margins was 82 and 75%. The utility of this approach was tested by imaging the in vivo tumor cavities from 34 mice after resection of a sarcoma with local recurrence as a bench mark. When applied prospectively to images from the tumor cavity, the sensitivity and specificity for differentiating local recurrence was 78 and 82%. For comparison, if pathology was used to predict local recurrence in this data set, it would achieve a sensitivity of 29% and a specificity of 71%. These results indicate a robust approach for detecting microscopic residual disease, which is an effective predictor of local recurrence.

Objectives Accurate diagnosis of malignant peripheral nerve sheath tumor (MPNST) is often challenging on fine-needle aspiration (FNA) or core needle biopsy. Recurrent mutations in EED and SUZ12, which encode subunits of polycomb repressive complex 2 (PRC2), have been identified in 70% to 92% of MPNSTs; PRC2 inactivation leads to loss of trimethylation of lysine 27 of histone H3 (H3K27me3). We evaluated the utility of H3K27me3 immunohistochemistry for distinguishing MPNST from its cytomorphologic mimics. Methods H3K27me3 immunohistochemistry was performed on 180 cases of spindle cell neoplasms sampled by FNA (n = 66) and needle biopsy (n = 114), and loss of nuclear staining was scored. Tumor types included MPNST, dedifferentiated liposarcoma, schwannoma, solitary fibrous tumor, leiomyosarcoma, melanoma, synovial sarcoma, sarcomatoid carcinoma, gastrointestinal stromal tumor, desmoid fibromatosis, low-grade fibromyxoid sarcoma, and unclassified spindle cell sarcoma/undifferentiated pleomorphic sarcoma. Results Complete loss of H3K27me3 was observed in 54% (13/24) of MPNSTs. In contrast, only two (of 156) histologic mimics showed complete loss of H3K27me3. Partial loss of H3K27me3 was present in a subset of cases (26/180), including both MPNST and non-MPNSTs. Conclusions Complete loss of H3K27me3 is a highly specific (98.7%) marker of MPNST that can distinguish MPNST from cytomorphologic mimics in FNA cell block and small biopsy specimens.

Purpose: Cathepsin-activated fluorescent probes can detect tumors in mice and in canine patients. We previously showed that these probes can detect microscopic residual sarcoma in the tumor bed of mice during gross total resection. Many patients with soft tissue sarcoma (STS) and other tumors undergo radiation therapy (RT) before surgery. This study assesses the effect of RT on the ability of cathepsin-activated probes to differentiate between normal and cancerous tissue. Methods and Materials: A genetically engineered mouse model of STS was used to generate primary hind limb sarcomas that were treated with hypofractionated RT. Mice were injected intravenously with cathepsin-activated fluorescent probes, and various tissues, including the tumor, were imaged using a hand-held imaging device. Resected tumor and normal muscle samples were harvested to assess cathepsin expression by Western blot. Uptake of activated probe was analyzed by flow cytometry and confocal microscopy. Parallel in vitro studies using mouse sarcoma cells were performed. Results: RT of primary STS in mice and mouse sarcoma cell lines caused no change in probe activation or cathepsin protease expression. Increasing radiation dose resulted in an upward trend in probe activation. Flow cytometry and immunofluorescence showed that a substantial proportion of probe-labeled cells were CD11b-positive tumor-associated immune cells. Conclusions: In this primary murine model of STS, RT did not affect the ability of cathepsin-activated probes to differentiate between tumor and normal muscle. Cathepsin-activated probes labeled tumor cells and tumor-associated macrophages. Our results suggest that it would be feasible to include patients who have received preoperative RT in clinical studies evaluating cathepsin-activated imaging probes., Damon Runyon Cancer Research Foundation (Damon Runyon-Rachleff Innovation Award)

Treatment of soft tissue sarcoma (STS) includes complete tumor excision. However, in some patients, residual sarcoma cells remain in the tumor bed. We previously described a novel hand-held imaging device prototype that uses molecular imaging to detect microscopic residual cancer in mice during surgery.To test this device in a clinical trial of dogs with naturally occurring sarcomas, we asked: (1) Are any adverse clinical or laboratory effects observed after intravenous administration of the fluorescent probes? (2) Do canine sarcomas exhibit fluorescence after administration of the cathepsin-activated probe? (3) Is the tumor-to-background ratio sufficient to distinguish tumor from tumor bed? And (4) can residual fluorescence be detected in the tumor bed during surgery and does this correlate with a positive margin?We studied nine dogs undergoing treatment for 10 STS or mast cell tumors. Dogs received an intravenous injection of VM249, a fluorescent probe that becomes optically active in the presence of cathepsin proteases. After injection, tumors were removed by wide resection. The tumor bed was imaged using the novel imaging device to search for residual fluorescence. We determined correlations between tissue fluorescence and histopathology, cathepsin protease expression, and development of recurrent disease. Minimum followup was 9 months (mean, 12 months; range, 9-15 months).Fluorescence was apparent from all 10 tumors and ranged from 3 × 10(7) to 1 × 10(9) counts/millisecond/cm(2). During intraoperative imaging, normal skeletal muscle showed no residual fluorescence. Histopathologic assessment of surgical margins correlated with intraoperative imaging in nine of 10 cases; in the other case, there was no residual fluorescence, but tumor was found at the margin on histologic examination. No animals had recurrent disease at 9 to 15 months.These initial findings suggest this imaging system might be useful to intraoperatively detect residual tumor after wide resections.The ability to assess the tumor bed intraoperatively for residual disease has the potential to improve local control.

BACKGROUND: The goal of limb-sparing surgery for a soft tissue sarcoma of the extremity is to remove all malignant cells while preserving limb function. After initial surgery, microscopic residual disease in the tumor bed will cause a local recurrence in approximately 33% of patients with sarcoma. To help identify these patients, the authors developed an in vivo imaging system to investigate the suitability of molecular imaging for intraoperative visualization. METHODS: A primary mouse model of soft tissue sarcoma and a wide field-of-view imaging device were used to investigate a series of exogenously administered, near-infrared (NIR) fluorescent probes activated by cathepsin proteases for real-time intraoperative imaging. RESULTS: The authors demonstrated that exogenously administered cathepsin-activated probes can be used for image-guided surgery to identify microscopic residual NIR fluorescence in the tumor beds of mice. The presence of residual NIR fluorescence was correlated with microscopic residual sarcoma and local recurrence. The removal of residual NIR fluorescence improved local control. CONCLUSIONS: The authors concluded that their technique has the potential to be used for intraoperative image-guided surgery to identify microscopic residual disease in patients with cancer. Cancer 2012. © 2012 American Cancer Society.

Recent advances in sarcoma genomics have identified novel mutations in the PI3K pathway in human sarcomas. Here, we use a mouse model of primary soft-tissue sarcoma for preclinical testing of doxorubicin and inhibitors of the PI3K pathway: BKM120 (PI3K inhibitor) and BEZ235 (a dual PI3K/mTOR inhibitor). Doxorubicin-treated tumors (n=15) showed a partial response rate of 6.6%, just as the majority of human sarcomas do not respond to doxorubicin. Treatment with BKM120 elicited a partial response in 50% of tumors (n=10), which was also seen in combination with doxorubicin (n=10). Additionally, BKM120 treatment produced a robust delay in tumor growth kinetics. BEZ235-treated tumors (n=9) showed a complete response rate of 11.1%. Combining BEZ235 with doxorubicin (n=10) increased the complete response rate to 50% (P=0.035). These studies demonstrate that PI3K pathway inhibition is a viable and attractive target for soft-tissue sarcomas.

Several experiments were performed to elucidate the origin of the second‐order optical nonlinearity χ(2) induced in the corona‐poled Corning 7059 glass films. The values and the time decay of the second‐harmonic coefficient d33 were measured for the glass films poled on several different kinds of substrates. The value of d33 was found to be dependent on the kinds of substrates. It was also observed that the glass films poled on a soda‐lime substrate showed faster time decay of d33 than those poled on a Pyrex glass substrate. The glass films were also poled at room temperature, and the values of d33 were measured as a function of poling time. As a result, the value of d33 increased with the poling time and approached a value which was close to that obtained for the glass films poled at 100 °C. From the experimental results, it is speculated that the space charge formed in the vicinity of the interface between the glass film and the substrate contributes to the creation of the χ(2). The planar charge densit...

Soft tissue tumors are composed of a large heterogenous group of mesenchymal neoplasms that are classified accordingly to the normal tissue counterpart. However, morphologic overlap between tumors may happen, and neoplasms may be further divided into histologic broad categories: round cell tumors, spindle cell tumors, epithelioid tumors, myxoid tumors, and pleomorphic tumors. In addition, soft tissue neoplasms have been shown to have reproducible numerical and structural chromosomal abnormalities that can be very promptly recognized by conventional cytogenetics and/or by other molecular tests, such as fluorescence in situ hybridization (FISH). The diagnosis of these neoplasms is based on the histologic recognition of the lesion as well as expression of immunoperoxidase studies in certain tumors, and characteristic cytogenetic findings. In contrast to most other solid tumors, in particular epithelial neoplasms, where the American Joint Committee on Cancer (AJCC) staging plays a pivotal role in the prognosis, in soft tissue neoplasms is dictated by natural history of the tumors and treatment.

A fluorescence-based imaging system has been developed for real-time, in vivo tumor margin assessment with microscopic sensitivity. Intra-operative imaging results have matched post-surgical histo-pahtological margin assessment in all the sarcoma mice cases tested so far.

DNA fibers were prepared by melt spinning method from DNA-CTMA powder. A hemicyanine dye, trans-4-{4-(dibutylamino)-styryl}-1-methylpyridinium iodide (DBASMPI) doped DNA-CTMA fiber with core diameter of 1 mm and dye concentration of 3.6 wt% was obtained by soaking it in an aqueous dye solution. Laser (532 nm) pumped amplified spontaneous emission (ASE) at 610 nm was observed in the dye-doped DNA-CTMA fiber. The ASE occurred at energy density 50 mW. The amplification of optical signals at 607 nm wavelength was confirmed. The results from ASE emphasize that DBASMPI doped DNA-CTMA fiber is appealing as a good candidate for optical amplifiers and superfluorescence sources in a variety of communication and sensor applications.

TPS11135 Background: Local recurrence is a common mode of failure for patients with cancer. Intra-operative detection of microscopic residual disease in the tumor bed could be used to decrease the ...

Second‐order optical nonlinearity χ(2) is induced in glass films by corona poling. The glass films are fabricated by radio‐frequency sputtering Corning7059 onto Pyrex glass substrates. The values of second‐order nonlinear coefficient d33 of the corona‐poled glass films are estimated to be ∼0.5 pm/V. Using the corona‐poled glass films as a waveguide, phase‐matched second‐harmonic generation has been demonstrated in the glass films for the first time. The origin of the χ(2) is considered to be related to the defects in the glass films.

Corning 7059 glass films were deposited onto 1mm Pyrex glass substrates by rf sputtering using gas mixture of Ar and O2. Corona discharge for poling the glass films was used between a tungsten wire and the electrically grounded Pyrex glass substrates apart 10 mm each other. The poling was carried out for five hours at room temperature and at 150 °C.

Limb-sparing surgery for extremity soft tissue sarcoma removes all cancer cells at the primary tumor site in the majority of patients. However, without radiation therapy, microscopic residual sarcoma cells left behind in the tumor bed will cause a tumor recurrence in approximately one-third of patients. Therefore, adjuvant radiation therapy is delivered to most patients, even when the tumor bed lacks residual cancer. Here, we present an imaging system for residual cancer assessment, consisting of a novel wide field-of-view imaging device and a protease-activated fluorescent probe. We demonstrate that this system directly images microscopic residual sarcoma cells in the tumor bed of mice when primary soft tissue sarcomas are resected. Moreover, this system can detect single tumor cells that have activated the fluorescent probe in vivo. This technology has the potential to be used as an intra-operative tool to identify microscopic residual disease for soft tissue sarcoma and other cancers with the goal of reducing rates of local recurrence, re-operation for positive margins, and adjuvant radiation therapy to tumor beds that lack residual cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4316.