Human Effectors of Acute and Chronic GVHD Overexpress CD83 and Predict Mortality.

PURPOSE: Acute and chronic GVHD remain major causes of transplant-related morbidity and mortality (TRM) after allogeneic hematopoietic cell transplantation (alloHCT). We have shown CD83 chimeric antigen receptor (CAR) T cells prevent GVHD and kill myeloid leukemia cell lines. In this pilot study, we investigate CD83 expression on GVHD effector cells, correlate these discoveries with clinical outcomes, and evaluate critical therapeutic implications for transplant recipients. EXPERIMENTAL DESIGN: CD83 expression was evaluated among circulating CD4+ T cells, B-cell subsets, T follicular helper (Tfh) cells, and monocytes from patients with/without acute or chronic GVHD (n = 48 for each group), respectively. CD83 expression was correlated with survival, TRM, and relapse after alloHCT. Differential effects of GVHD therapies on CD83 expression was determined. RESULTS: CD83 overexpression on CD4+ T cells correlates with reduced survival and increased TRM. Increased CD83+ B cells and Tfh cells, but not monocytes, are associated with poor posttransplant survival. CD83 CAR T eliminate autoreactive CD83+ B cells isolated from patients with chronic GVHD, without B-cell aplasia as observed with CD19 CAR T. We demonstrate robust CD83 antigen density on human acute myeloid leukemia (AML), and confirm potent antileukemic activity of CD83 CAR T in vivo, without observed myeloablation. CONCLUSIONS: CD83 is a promising diagnostic marker of GVHD and warrants further investigation as a therapeutic target of both GVHD and AML relapse after alloHCT.

Genomic and Single-Cell Landscape Reveals Novel Drivers and Therapeutic Vulnerabilities of Transformed Cutaneous T-cell Lymphoma.

ABSTRACT: Cutaneous T-cell lymphoma (CTCL) is a rare cancer of skin-homing T cells. A subgroup of patients develops large cell transformation with rapid progression to an aggressive lymphoma. Here, we investigated the transformed CTCL (tCTCL) tumor ecosystem using integrative multiomics spanning whole-exome sequencing (WES), single-cell RNA sequencing, and immune profiling in a unique cohort of 56 patients. WES of 70 skin biopsies showed high tumor mutation burden, UV signatures that are prognostic for survival, exome-based driver events, and most recurrently mutated pathways in tCTCL. Single-cell profiling of 16 tCTCL skin biopsies identified a core oncogenic program with metabolic reprogramming toward oxidative phosphorylation (OXPHOS), cellular plasticity, upregulation of MYC and E2F activities, and downregulation of MHC I suggestive of immune escape. Pharmacologic perturbation using OXPHOS and MYC inhibitors demonstrated potent antitumor activities, whereas immune profiling provided in situ evidence of intercellular communications between malignant T cells expressing macrophage migration inhibitory factor and macrophages and B cells expressing CD74. SIGNIFICANCE: Our study contributes a key resource to the community with the largest collection of tCTCL biopsies that are difficult to obtain. The multiomics data herein provide the first comprehensive compendium of genomic alterations in tCTCL and identify potential prognostic signatures and novel therapeutic targets for an incurable T-cell lymphoma. This article is highlighted in the In This Issue feature, p. 1171.

FGD4 (Frabin) Overexpression in Pancreatic Neuroendocrine Neoplasms.

OBJECTIVE: The pathogenesis of pancreatic neuroendocrine tumors (PNETs) is still unclear. We propose Frabin as a new molecular alteration in PNETs. Frabin is a guanine nucleotide exchange factor playing a role in mediating actin cytoskeleton changes during cell migration, morphogenesis, polarization, and division. METHODS: Patients with PNETs of different grades were assessed for Frabin expression using immunohistochemistry and tissue microarray. The tissue microarray included 12 grade 1 and 3 grade 2 PNETs and 14 grade 3 pancreatic neuroendocrine carcinomas (PECAs). Frabin immunostain was scored with Allred system. Statistical analysis used SAS and R software. Immunohistochemistry scores were correlated with tumor grade and stage. The Spearman correlation coefficient was calculated with P values. RESULTS: Pancreatic neuroendocrine tumors were graded according to the World Health Organization 2017 guidelines. Frabin was expressed by 24 (82.7%) of the PNET/PECA studied. Only 5 (17.2%) of the 29 PNETs/PECA evaluated were Frabin negative. Frabin expression was cytoplasmic in all cases. We found a significant positive correlation (ρ = 0.47) between Frabin immunohistochemistry score and tumor grade (P = 0.01). No correlation was found between Frabin expression and tumor stage (P = 0.91). CONCLUSIONS: We report Frabin overexpression as a novel molecular alteration occurring in PNETs/PECAs.

Frequency of Mismatch Repair Protein Deficiency in a Puerto Rican Population with Colonic Adenoma and Adenocarcinoma.

BACKGROUND/AIM: Microsatellite instability (MSI) results from genetic alterations involving the mismatch repair (MMR) genes MLH1, PSM2, MSH2, and MSH6. MSI has been implicated in both sporadic CRC and Lynch syndrome. The aim of the study was to assess the frequency of alterations in MMR protein expression in both primary colorectal cancer and precursor lesions among Puerto Rican patients. PATIENTS AND METHODS: A retrospective study of 84 Puerto Rican patients was performed to assess the frequency of MMR protein expression alterations in both primary CRC and precursor lesions using tissue microarray and immunohistochemistry. RESULTS: The loss of expression of both MLH1 and PMS2 proteins was present in 6.3% of adenomas, 9.1% of adenomas with high-grade dysplasia and 9.4% of colon adenocarcinomas. Negative nuclear staining for both MSH2 and MSH6 proteins was found in 2.4% of colon adenocarcinomas. CONCLUSION: When compared to prior reports, this study suggests a lower frequency of MSI among the Puerto Rican population. The higher prevalence of MLH1 mutations correlates with previous studies of protein expression among the Hispanic community including Colombian, Uruguay and Brazilian populations.

Cancer Antigen 125 (CA125, MUC16) Protein Expression in the Diagnosis and Progression of Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive carcinoma, with most patients diagnosed at an advanced stage, with a 5-year survival rate of around 5%. An urgent need exists for identifying better diagnostic, prognostic, and therapeutic markers for this lethal disease. Recently, CA125 has been identified in PDAC, and the aim of this research is to study the changes in CA125 expression during the progression of benign pancreatic tissue (BPT) to PDAC and to assess its value as a biomarker of tumor growth. To address these questions, the cellular levels of CA125 in BPT and PDAC were measured using immunohistochemistry and compared on the basis of tumor staging, and the tissue microarray technology were constructed using resected pancreatic tissues. The staining reactions for each case were evaluated semiquantitatively using the histologic score system. Our investigation demonstrates a consistent and significant upregulation of CA125 during the transition from BPT to PDAC. We also found a direct correlation between CA125 immunohistochemistry score and tumor stage (P=0.02). In conclusion, our data indicate that CA125 plays a direct role in pancreatic carcinogenesis and suggests that it may eventually be used as a diagnostic and/or prognostic biomarker of pancreatic cancer. Prospective studies are recommended to evaluate further the diagnostic and prognostic capabilities of CA125 in PDAC, and further studies are warranted to assess the use of CA125 as a therapeutic marker.

Her-2 Expression in Gastroesophageal Intestinal Metaplasia, Dysplasia, and Adenocarcinoma.

Overexpression of human epidermal growth factor receptor 2 protein (Her-2) in Barrett neoplasia is significant for targeted therapy with trastuzumab. Here, we studied the frequency of Her-2 overexpression in Barrett adenocarcinoma and precursor lesions. Retrospective formalin-fixed paraffin-embedded tissue samples of 25 normal (NM) esophageal mucosa, 50 Barrett esophagus (BE) without dysplasia, 49 BE with low-grade dysplasia (LGD), 50 BE with high-grade dysplasia (HGD), and 50 invasive adenocarcinoma (ICA) were used. A BE tissue microarray was built and analyzed by Her-2 immunohistochemistry (IHC) and Her-2 dual in situ hybridization (DISH). Her-2 IHC expression was negative in NM and low in 26% of BE (IHC score: 1+) and in 24.5% of LGD (IHC score: 1 to 2+). Her-2 overexpression was seen in 28% of HGD and in 24% of ICA (IHC score: 2 to 3+). Her-2 DISH was negative in NM and BE but positive in 6% of LGD, 20% of HGD, and 18% of ICA. Differences in Her-2 DISH positivity between NM and HGD or ICA were statistically significant (P=0.02), but those between NM and LGD or HGD and ICA were not (P=0.2). Although Her-2 overexpression results in ICA were similar to previous reports, the finding of 28% in HGD was unexpected and may have clinical implications. Positive Her-2 DISH in 6% of LGD is novel, suggesting a role of Her-2 during BE progression.

Expression of CXCR4, E-cadherin, Bcl-2, and survivin in Merkel cell carcinoma: an immunohistochemical study using a tissue microarray.

Merkel cell carcinoma (MCC) is a rare but highly aggressive cutaneous malignancy with a mortality rate exceeding that of melanoma. Although smaller studies of markers of progression have been performed, large-scale investigation has been difficult due to the rarity of this tumor. Investigation of 4 potential immunohistochemical progression markers using an MCC tissue microarray was performed. An immunohistochemical analysis of CXCR4, E-cadherin, Bcl-2, and Survivin was performed on a tissue microarray of two hundred twenty-seven 0.6-mm tumor cores-110 primary, 73 local/regional metastatic, and 44 distant metastatic-from 87 patients, 23 of which were sampled 2 or more times. There was a statistically significant increase in immunoreactivity to CXCR4 and Survivin in local/regional nodal MCC metastases compared with primary and distant metastatic lesions. No significant differences by disease location were found for either Bcl-2 or E-cadherin. These results suggest a potential role for CXCR4 and Survivin in MCC tumor progression. However, previous data from other studies suggesting a role for Bcl-2 and E-cadherin in MCC progression are not confirmed in this larger sample. Further discovery of additional markers are needed to better characterize this rare but deadly malignancy.

Bax-interacting factor-1 expression in prostate cancer.

BACKGROUND: Bax-interacting factor (Bif)-1 protein is a member of the endophilin B family that binds to and activates the proapoptotic Bax protein in response to apoptotic signals. Loss of Bif-1 suppresses the intrinsic pathway of apoptosis and promotes tumorigenesis. We examined the expression levels of Bif-1 protein in human prostate cancer. MATERIALS AND METHODS: Thirty-nine archival tissue specimens of human prostate cancer, and a human prostate cancer tissue microarray containing 19 samples of normal prostate, 26 samples of benign prostatic hyperplasias (BPHs), 30 samples of high-grade prostatic intraepithelial neoplasia (PIN), and 153 samples of prostate cancer, were selected for immunohistochemical staining with Bif-1 antibody. The slides were scored by 2 independent observers. RESULTS: Nontissue microarray samples: moderate to strong Bif-1 staining was identified in 38 of 39 prostate cancer samples. In 32 cases, foci of PIN were identified adjacent to prostate cancer samples. Of these, 29 samples (90.6%) showed strong and diffuse Bif-1 staining. Benign prostatic hyperplasias, identified in 27 cases, was weakly Bif-1 positive in 88.9% of cases. Tissue microarray samples: 38.6% (59 of 153) of prostate cancer samples showed moderate to strong Bif-1 expression, and 21.6% (33 of 153) were Bif-1 negative. Bif-1 expression was moderate to strong in 76.7% (23 of 30) of PIN. Bif-1 was weak to moderate in 53.8% (14 of 26) of BPH and negative in 46.2% (12 of 26) of them. Low to moderate Bif-1 was seen in 89.5% of normal prostate samples. CONCLUSION: The loss of Bif-1 expression in a subset of prostate cancer samples is in agreement with the proapoptotic function of Bif-1. The significance of the increased Bif-1 in a subgroup of prostate cancer samples and in PIN remains to be determined. It seems that Bif-1 has a role in prostate cancer, providing the rationale for using Bif-1 as a target for prostate anticancer therapy.

Activation of the serine/threonine protein kinase AKT during the progression of colorectal neoplasia.

BACKGROUND: AKT has been identified as a major regulator of cell proliferation, tumorigenesis, and apoptosis. In this study, we evaluated changes in the activity of AKT during colorectal cancer (CRC) progression. MATERIALS AND METHODS: We used stage-oriented human CRC tissue microarrays, including 99 invasive carcinomas, 28 tubular adenomas, and 18 samples of normal colonic mucosa. The tissue array slides were stained with a mouse monoclonal antiphospho-AKT antibody using the avidin-biotin complex method. RESULTS: Activation of AKT was detected mostly in the invasive carcinomas. Sixty-three percent of carcinomas demonstrated strong to moderate AKT activity. Seven percent of carcinomas were phospho-AKT (p-AKT) negative, and 30% (30 of 99) were p-AKT weakly positive. Conversely, 78% of normal colonic mucosas were p-AKT negative, and only 4 samples stained weakly for p-AKT. Eighty-two percent of adenomas were weakly positive for p-AKT, 1 was p-AKT negative, and none exhibited strong or moderate p-AKT stain. At a significance level of .05, we found that the distribution of p-AKT stain scores for cancer was shifted to the right of adenoma (P < .0001) and normal (P < .0001) and for adenoma was shifted to the right of normal (P < .0001). AKT activation did not correlate with tumor stage (P = .28), lymph node metastasis (P = .45), lymphatic invasion (P = .46), or distant metastasis (P = .34). CONCLUSION: This study shows increasing activation of AKT during CRC progression. This finding suggests a role of p-AKT in colorectal carcinogenesis and provides a rationale for using p-AKT inhibitor API-2/triciribine, which is currently under clinical investigation for the treatment of CRC.