Citations

Abstract Background: Previous studies showed conflicting results regarding association between stromal tumor infiltrating lymphocytes (sTIL) and outcome in HER2+ breast cancer. While sTIL were not associated with outcome in patients (pts) treated with trastuzumab in N9831, immune functions genes are linked to outcome in these pts. TLS is an organized form of ectopic lymphoid aggregates (LA) with secondary lymphoid organ structure. Several studies showed that TLS associates with improved outcome in multiple cancers. However, distinguishing TLS and simple LA is challenging, particularly when germinal center is absent. In this study, we evaluated integrated pathological quantification and genomic data to assess functional TLS. Methods: Pathological evaluation of LA in H&E slides from pts treated in Arm A (chemotherapy alone) and Arm C (chemotherapy with concurrent trastuzumab) in N9831 was performed. NanoString was used to quantify mRNA. Wilcoxon rank sum test, Chi squared test, Kaplan-Meier method and Cox regression model were used to evaluate association between LA and baseline characteristics as well as outcomes. Results: LA was quantified in 1011 pts (526 Arm A, 485 Arm C). Greater number of LA was significantly associated with higher tumor grade, ER/PR negativity and increasing sTIL, but not age, tumor size or lymph node status. Increasing numbers of LA were associated with improved RFS in both arms combined (p 0.028). However, using multivariable Cox regression analysis in each treatment arm, LA (≥ 1 vs 0) was associated with improved RFS only in Arm A (HR 0.6, 95%CI 0.43-0.84, p 0.003) but not Arm C (HR 0.72, 95%CI 0.47-1.1, p 0.134). Further evaluation of expression of TLS-related immune genes including IFNG, ICOSLG, CXCL13, CXCR3, BCL6, IL21R, ICOS, PDCD1, CXCR5, CXCL9, TBX21, CD38, CXCL10, CLXCL11, IL21, CD200 was carried out among 252 pts in Arm C with LA ≥ 1. Among these 16 TLS-related immune genes, as a continuous variable, higher expression of BCL6 (HR 0.61, 95%CI 0.41-0.92, p 0.019) and IL21R (HR 0.78, 95%CI 0.62-0.98, p 0.03) were associated with improved RFS in Arm C pts with LA ≥ 1. However, these genes were not significantly associated with outcome in Arm C pts without LA with BCL6 HR1.07 (95%CI 0.67-1.71, p 0.776) and IL21R HR 0.96 (95%CI 0.71-1.29, p 0.775). Conclusion: Similar to sTIL, greater number of LA was associated with improved outcome in HER2+ pts treated with chemotherapy alone but not chemotherapy in combination with trastuzumab. Using histogenomic integration with the combination of pathological LA and TLS-related immune genes, we identified that pts with functional TLS with LA ≥ 1 and higher expression of BCL6 or IL21R had significantly improved outcome when treated with trastuzumab. Future studies are needed to further confirm these findings. Support: U10 CA180821, U24 CA196171, https://acknowledgments.alliancefound.org; Genentech; Clinicaltrials.gov Id: NCT00005970 Citation Format: Saranya Chumsri, Tracy R. Shachner, Zhuo Li, Nadine Norton, Alvaro Moreno-Aspitia, Gerardo Colon-Otero, Edith A. Perez, E. A. Thompson, Aziza Nassar, Keith L. Knutson. Functional tertiary lymphoid structure (TLS) and outcome in HER2-positive (HER2+) breast cancer in NCCTG N9831 (Alliance) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5627.

Abstract Objective Using a US Food and Drug Administration (FDA) emergency use authorization (EUA) reverse transcription polymerase chain reaction (RT-PCR) method, we examined the analytic performance accuracy of saliva specimens as compared to nasopharyngeal (NP) specimens in symptomatic patients. Correlation between test results and symptoms was also evaluated. Methods Over a 5-week period in 2020, 89 matched saliva and nasopharyngeal swabs were collected from individuals exhibiting symptoms consistent with SARS-CoV-2. Specimens were tested with an FDA EUA-approved RT-PCR method, and performance characteristics were compared. Results The concordance rate between saliva and nasopharyngeal testing was 93.26%. The mean cycle threshold value of saliva when compared to the NP specimen was 3.56 cycles higher. As compared to NP swab, saliva testing demonstrates acceptable agreement but lower sensitivity. Conclusion When compared to a reference method using NP swabs, the use of saliva testing proved to be a reliable method. Self-collected saliva testing for SARS-CoV-2 allows for a viable option when trained staff or collection materials are in short supply.

535 Background: Invasive lobular carcinoma (ILC) is a rarer form of breast cancer, accounting for 10% of the disease cases. HER2 overexpression in ILC is infrequent and limited data exist regarding clinical characteristics and outcome of HER2-positive (HER2+) ILC patients (pts) treated with adjuvant trastuzumab. Methods: Patient characteristics were compared between ILC and invasive ductal carcinoma (IDC) using Wilcoxon rank sum test for continuous variables and Chi-square test for categorical variables. Kaplan-Meier (KM) method was used to estimate the freedom from mortality and recurrence. Cox regression model was used to evaluate the association between ILC and outcomes adjusted for other characteristics. NanoString technology was used to quantify mRNA to develop immune-related gene signatures. Results: From a total of 3,304 pts, 122 (3.7%) pts had ILC. Pts with ILC were significantly older (median age 54 vs. 49 years), had larger tumors, lower grade, more ER and PR positive tumors, and more lymph node involvement (25.4% had N3 disease compared to 12.7% in IDC). Overall, with KM analysis, pts with ILC had significantly worse overall survival (OS, p = 0.005) and recurrence-free survival (RFS, p = 0.046) compared to IDC. The 15-year freedom from recurrence was merely 57.67% in ILC compared to 72.68% in IDC. A significant number of hormone receptor-positive (HR+) ILC pts developed late recurrence with cumulative event rates increasing from 23% at 5 years to 42% at 15 years. Nevertheless, in multivariate Cox regression analysis adjusting for other clinical characteristics, including age, tumor size, grade, ER/PR, and lymph node status, lobular histology was not significantly associated with worse outcome for OS (HR = 1.19, 95%CI 0.67-2.1, p = 0.55) and RFS (HR = 1.5, 95%CI 0.9-2.5, p = 0.12), as compared with IDC. However, ILC pts appeared to have similar degree of benefit from trastuzumab, with RFS HR = 0.58 compared to HR = 0.67 in the entire population. For immune landscape, there was no significant difference in gene signatures related to CD45, CD8, B cells, or cytotoxic cells. However, ILC had more enrichment in mast cell gene signature and fewer macrophage, NK CD56dim, and regulatory T cell signatures compared to IDC (p < 0.05). Conclusions: HER2+ ILC has distinct clinical characteristics and immune landscape compared to IDC. ILC pts appeared to have worse outcome compared to IDC likely because ILC pts often presented with more locally advanced disease. However, similar benefit of trastuzumab was observed in ILC pts. Due to high risk of late relapse in HR+ HER2+ ILC, extended adjuvant endocrine therapy should be considered in this group of high-risk pts. Clinical trial information: NCT00005970.

Abstract Context. —Papanicolaou tests with atypical squamous cells of undetermined significance (ASC-US) cytology and adjunct testing for high-risk human papillomaviruses (hr-HPV) are helpful in detecting high-grade disease. Detection of disease may be further improved with molecular markers known to be overexpressed in cervical carcinoma. ProEx C detects 2 such molecular markers, minichromosome maintenance protein 2 and topoisomerase II, which are associated with abnormal cell cycle regulation. Objective. —To determine the utility of ProEx C as a marker for high-grade cervical intraepithelial neoplasia 2+ disease when compared with hr-HPV status in Papanicolaou tests with ASC-US cytology. Design. —A SurePath slide was prepared on all ASC-US cases from the residual SurePath vial pellet and stained using the ProEx C reagent prediluted with water-bath antigen retrieval, using a Dako autostainer. Nuclear staining of cytologically atypical squamous cells was considered a positive result. Adjunct testing for hr-HPV used Digene Hybrid Capture 2. Follow-up biopsy results were available for review following the Papanicolaou test. Results. —Two hundred patients with ASC-US diagnoses were part of this study. The sensitivities of ProEx C and hr-HPV testing in detecting high-grade cervical intraepithelial neoplasia 2+ disease were 98.04% and 82.35%, respectively, whereas the specificity for detecting high-grade disease was 74.50% and 73.15%, respectively. Conclusions. —ProEx C staining is a more sensitive and specific biomarker for detecting cervical disease than adjunct testing for hr-HPV status in Papanicolaou tests with ASC-US.

Inflammatory breast cancer is a highly aggressive form of breast cancer that forms clusters of tumor emboli in dermal lymphatics and readily metastasizes. These cancers express high levels of E-cadherin, the major mediator of adherens junctions, which enhances formation of tumor emboli. Previous studies suggest that E-cadherin promotes cancer when the balance between apical and basolateral cadherin complexes is disrupted. Here, we used immunohistochemistry of inflammatory breast cancer patient samples and analysis of cell lines to determine the expression of PLEKHA7, an apical adherens junction protein. We used viral transduction to re-express PLEKHA7 in inflammatory breast cancer cells and examined their aggressiveness in 2D and 3D cultures and in vivo. We determined that PLEKHA7 was deregulated in inflammatory breast cancer, demonstrating improper localization or lost expression in most patient samples and very low expression in cell lines. Re-expressing PLEKHA7 suppressed proliferation, anchorage independent growth, spheroid viability, and tumor growth in vivo. The data indicate that PLEKHA7 is frequently deregulated and acts to suppress inflammatory breast cancer. The data also promote the need for future inquiry into the imbalance between apical and basolateral cadherin complexes as driving forces in inflammatory breast cancer.

Abstract Background Echinococcus multilocularis is a destructive zoonotic cestode with low human incidence. Hydatid disease classically presents with hepatic or lung involvement with infrequent extrahepatic bone destruction. Diagnosis is challenging due to its latency and mimicry. Fig.1: Case 1 - X-ray imaging of the pelvis shows osseous destruction of the iliac crest secondary to known osteomyelitis status post left ilium debridement. Fig.2: Case 1 - Magnetic resonance imaging demonstrates extensive osteomyelitis throughout left ilium. Stable scattered focal fluid collections seen throughout the left lower quadrant. Methods CASE 1: A 57 year-old Albanian male with diabetes, latent TB, and left iliac lytic lesion presented with 4 weeks of left flank pain and was treated with 6 weeks of IV Ceftriaxone and Flagyl. 2 years later he returned with flank pain and purulent lumbar drainage. Hip x-ray suggested chronic osteomyelitis, with left psoas fluid collections on CT. Bartonella, Q fever, Brucella, HIV, AFB and fungal serologies were negative. Hemipelvis debridement revealed structures concerning for hydatid disease. Echinococcus IgG was equivocal. Histopathology was consistent with Echinococcus multilocularis species, and albendazole was started. On follow-up, he presented with left hip tenderness and toe extensor weakness. Labs showed mild leukocytosis. CT revealed progressive destruction of the left iliac with sacroiliac extension concerning for abscess. CASE 2: A 36 year-old female presented with lung and liver cysts, progressive dyspnea, and non-productive cough. She lived in Africa, Asia, and Europe and consumed local street food and unpasteurized milk. Hobbies included spelunking and swimming in freshwater lakes. She had exposure to stray animals, but denied bites or scratches. Over 4 years dyspnea progressed to orthopnea. MR abdomen revealed a 10x6x12cm liver cyst and chest CT showed 2 fluid-attenuating lesions in the LLL and RLL, measuring 4.9 x 6.0 cm and 6.8 x 4.3 cm respectively. Echinococcus, Bartonella, Q fever, Brucella, HIV, AFB and fungal serologies were negative. Schistosomiasis serology was equivocal. Fig. 3: Case 2 - MRI abdomen demonstrating 10x6x12cm liver cyst Fig. 4: Case 2 - Chest CT showed 2 dominant fluid attenuating lesions within the LLL and RLL. The larger lesion in RLL measures 6.8 x 4.3 cm. The left lower lobe lesion measures 4.9 x 6.0 cm. Results Patient 1 underwent type I hemipelvectomy. Patient 2 underwent pulmonary segmentectomy and liver lobectomy. Both were continued on albendazole. Fig. 5: Case 1 - Photo taken during debridement of left ileac and hip. Note presence of white cysts discovered intraoperatively. Fig. 6: Case 1 - Histopathologic slides (H&E stain) demonstrating hooks and scolices consistent with Echinococcus multilocularis. A. Hooklet (100x magnification). B. Hydatid cyst with black-staining structures suggestive of degenerating hooklets. C. Zoomed detail of cyst wall. D. Degenerating hydatid cyst and hooklets. Conclusion Equivocal IgG serology does not exclude infection. History and clinical presentation are key to diagnosis, but histopathology remains the gold standard. Hydatid bone infection progresses insidiously and frequently recurs, depending upon excision and debridement. Finally, echinococcosis demands aggressive long-term therapy and surveillance. Disclosures Claudia R. Libertin, MD, Pfizer, Inc. (Grant/Research Support, Research Grant or Support)

BackgroundIn the US, 60% to 80% of oropharyngeal squamous cell carcinomas (OPSCCs) are associated with human papillomavirus (HPV). However, until recently, no consensus existed about when and how to test for HPV in patients with head and neck cancers. We aimed to evaluate the use of p16 and HPV testing at our institution because p16 immunohistochemistry is reportedly a reliable surrogate marker for HPV detection in OPSCCs.MethodsWe identified all cases at our institution of primary or metastatic squamous cell carcinoma (SCC) of the head and neck with a concurrent p16 immunostain analysis from January 1, 2013, through August 31, 2018. Patient demographic data, tumor characteristics, p16 result, and any HPV result (in situ hybridization and E6 and E7 RNA test) were captured.ResultsWe identified 104 patients. Most primary tumors (53/57 [93.0%]) and metastases (40/47 [85.1%]) were positive for p16. Thirty‐seven cases (35.6%) had reflex high‐risk HPV (HR HPV) testing performed. Of the 35 p16‐positive cases, 6 had discrepant HR HPV results (p16+/HPV−). We identified 47 p16 immunostains that were performed on lymph nodes with primary tumors of unknown origin. Most were cytology cases (34/47 [72.3%]), and most were p16 positive (40/47 [85.1%]). Neither tumor differentiation nor tumor keratinization was predictive of p16 positivity. Tumors with basaloid differentiation were universally p16 positive.Conclusionp16 immunohistochemistry accurately identifies HPV‐positive OPSCC. Cytology specimens have an important role in characterizing SCC of unknown origin. HR HPV testing is not routinely required, and results may be discrepant with p16 findings.

Abstract Background: Inflammatory breast cancer is a highly aggressive form of breast cancer that robustly forms clusters of tumor emboli in dermal lymphatics and readily metastasizes. Inflammatory breast cancers express high levels of E-cadherin, the major protein of adherens junctions, which may enhance the ability of tumor cells to form such clusters and contribute to metastasis. Seemingly contradictory, E-cadherin has both tumor-suppressing and tumor-promoting roles in cancer; previous studies suggest that this depends on the balance between apical and basolateral cadherin-catenin complexes. Methods: In the present study, we use immunohistochemistry of inflammatory breast cancer patient samples and biochemical analysis of cell lines to determine the expression of PLEKHA7, an apical adherens junction protein. We use viral transduction to ectopically express PLEKHA7 in the SUM149 inflammatory breast cancer cell line. The effect of PLEKHA7 on the aggressiveness of inflammatory breast cancer in 2D, 3D and in-vivo were examined. Results: We determined that PLEKHA7 was deregulated in inflammatory breast cancer, demonstrating improper localization or lost expression in a strong majority of patient samples and very low expression in cell line models. We found that re-expressing PLEKHA7 is sufficient to suppress proliferation, anchorage independent growth, spheroid viability, and tumor growth in-vivo. We also observed a negative-selection pressure within the xenograft tumors to lose PLEKHA7 function or expression.Conclusions: The data indicate that PLEKHA7 is frequently deregulated and acts as a suppressor of inflammatory breast cancer. They also suggest that the resulting imbalance between apical and basolateral cadherin-catenin complexes contributes to growth, survival and emboli-forming capacities of inflammatory breast cancer.

Abstract Background National guidelines recommend genomic profiling of tumor tissue to guide precision therapy. We compared the specimen adequacy for genomic profiling and yield of DNA between endoscopic ultrasound (EUS)-guided fine-needle biopsy (FNB) and EUS-guided fine-needle aspiration (FNA). Methods In our tandem, randomized controlled trial, consecutive patients undergoing EUS for evaluation of pancreatic masses underwent both conventional EUS-FNA with a 25-gauge needle and paired EUS-FNB (19 or 22-gauge needle), with the order randomized (EUS-FNA first followed by EUS-FNB, or vice versa). A minimum of one pass with each needle was obtained for histology. Second and third passes were performed to collect DNA. Specimens were evaluated by a cytopathologist blinded to the needle type. Specimen adequacy for genomic profiling was calculated based on FoundationOne clinical diagnostic (CDx) adequacy requirements. We compared the adequacy for genomic profiling DNA (quantity) and histology yields with both needles. Results Analysis included 50 patients (25 men; mean age 68 [standard deviation (SD) 13] years), with a mean lesion size of 38 (SD 17) mm; 37 lesions (74 %) were pancreatic ductal adenocarcinoma (PDAC). The mean DNA concentrations in PDAC by FNB and FNA needles were 5.930 (SD 0.881) µg/mL vs. 3.365 (SD 0.788) µg/mL, respectively (P = 0.01). The median standardized histology score per pass with EUS-FNB was 5 (sufficient for histology) and for EUS-FNA was 2 (enough for cytology). Specimen adequacy for genomic profiling and yield of DNA was significantly higher with FNB than with FNA needles. Conclusions In this study, adequacy for genomic profiling, DNA, and histology yield were considerably superior using an EUS-FNB needle compared with an EUS-FNA needle.