Lauren Fishbein

Pheochromocytoma and paraganglioma: understanding the complexities of the genetic background

Pheochromocytomas and paragangliomas (PCC/PGL) are tumors derived from the adrenal medulla or extra-adrenal ganglia, respectively. They are rare and often benign tumors that are associated with high morbidity and mortality due to mass effect and high circulating catecholamines. Although most PCCs and PGLs are thought to be sporadic, over one third are associated with 10 known susceptibility genes. Mutations in three genes causing well characterized tumor syndromes are associated with an increased risk of developing PCCs and PGLs, including VHL (von Hippel-Lindau disease), NF1 (Neurofibromatosis Type 1), and RET (Multiple Endocrine Neoplasia Type 2). Mutations in any of the succinate dehydrogenase (SDH) complex subunit genes (SDHA, SDHB, SDHC, SDHD) can lead to PCCs and PGLs with variable penetrance, as can mutations in the subunit cofactor, SDHAF2. Recently, two additional genes have been identified, TMEM127 and MAX. Although these tumors are rare in the general population, occurring in two to eight per million people, they are more commonly associated with an inherited mutation than any other cancer type. This review summarizes the known germline and somatic mutations leading to the development of PCC and PGL, as well as biochemical profiling for PCCs/PGLs and screening of mutation carriers.

Whole-exome sequencing identifies somatic ATRX mutations in pheochromocytomas and paragangliomas

Pheochromocytomas and paragangliomas (PCC/PGL) are the solid tumor type most commonly associated with an inherited susceptibility syndrome. However, very little is known about the somatic genetic changes leading to tumorigenesis or malignant transformation. Here we perform whole exome sequencing on a discovery set of 21 PCC/PGL and identify somatic ATRX mutations in two SDHB-associated tumors. Targeted sequencing of a separate validation set of 103 PCC/PGL identifies somatic ATRX mutations in 12.6% of PCC/PGL. PCC/PGLs with somatic ATRX mutations are associated with alternative lengthening of telomeres and clinically aggressive behavior. This finding suggests that loss of ATRX, a SWI/SNF chromatin remodeling protein, is important in the development of clinically aggressive pheochromocytomas and paragangliomas.

Pheochromocytoma/Paraganglioma: Review of Perioperative Management of Blood Pressure and Update on Genetic Mutations Associated With Pheochromocytoma

Pheochromocytomas and paragangliomas are rare tumors with high morbidity rates caused by excessive catecholamine secretion, even though the majority of tumors are benign. The use of perioperative blockade regimens, together with improved surgical techniques, has greatly impacted the perioperative morbidity associated with these tumors. The old dogma of the “tumor of tens” no longer holds true. For example, at least one third of all pheochromocytomas and paragangliomas are hereditary, with mutations in 1 of 10 well‐characterized susceptibility genes, and one quarter of all tumors are malignant. This review focuses on the perioperative management of pheochromocytoma and paragangliomas and the clinical implications of the associated genetic mutations.

Pheochromocytoma and Paraganglioma: Genetics, Diagnosis, and Treatment

Pheochromocytomas (PCCs) and paragangliomas (PGLs) are rare but unique neuroendocrine tumors. The hypersecretion of catecholamines from the tumors can be associated with high morbidity and mortality, even when tumors are benign. Up to 40% of PCCs/PGLs are associated with germline mutations in susceptibility genes. About one-quarter are malignant, defined by the presence of distant metastases. Treatment options for unresectable metastatic disease, including chemotherapy, (131)I-MIBG, and radiation, can offer limited tumor and hormone control, although none are curative. This article reviews the inherited genetics, diagnosis, and treatment of PCCs and PGLs.

External beam radiation therapy (EBRT) for patients with malignant pheochromocytoma and non-head and -neck paraganglioma: combination with 131I-MIBG.

In patients with malignant pheochromocytoma and paraganglioma, 131I-MIBG radiotherapy can achieve an objective response rate of 30-50% with the dose limiting toxicity being hematologic. Patients with disseminated disease, who also have a few index bulky or symptomatic lesions, may benefit from the addition of targeted external beam radiotherapy alone or in combination with systemic 131I-MIBG. The records of patients with malignant paraganglioma who were treated with external beam radiotherapy at the University of Pennsylvania from February 1973 to February 2011 were reviewed in an institutional review board approved retrospective study. Of the 17 patients with tumors in the thorax, abdomen, or pelvis, 76% had local control or clinically significant symptomatic relief for at least 1 year or until death. As expected, the predominant toxicity was due to irradiation of tumor-adjacent normal tissues without clinically significant hematologic toxicity. Due to widespread systemic metastases with areas of bulky, symptomatic tumor, 5 of the 17 patients were treated with sequential 131I-MIBG (2 mCi/kg per treatment) and external beam radiotherapy to 9 sites. In these patients, all areas that were irradiated with external beam radiotherapy showed durable objective response despite all patients eventually experiencing out-of-field systemic progression requiring other treatment. Four of these patients remain alive with excellent performance status 16, 18, 23, and 24 months after external beam radiotherapy. External beam radiotherapy can be highly effective in local management of malignant paraganglioma and can be used in conjunction with 131I-MIBG due to nonoverlapping toxicities with excellent control of locally bulky tumors.

SDHB mutation carriers with malignant pheochromocytoma respond better to CVD

Pheochromocytomas and paragangliomas (PCCs/PGLs) are tumors in the adrenal medulla and extra-adrenal ganglia, respectively. About a quarter are metastatic, often with a long latency period. Some PCC/PGLs are defined as clinically aggressive given extensive local invasion into adjacent normal tissue. These tumors, like metastatic PCC/PGL, cannot be surgically cured, and patients can have continued secretion of excessive catecholamines with all related sequelae. For patients with metastatic or aggressive tumors, treatment options are limited. External beam radiation can provide local control (Fishbein et al. 2012, Vogel et al. 2014). Systemic therapy includes radionucleotide treatment with 131I-MIBG or chemotherapy with cyclophosphamide, vincristine and dacarbazine (CVD), both of which have a limited progression-free survival (PFS) and limited complete or partial regression rates based on meta-analyses (Niemeijer et al. 2014, van Hulsteijn et al. 2014). Because 131I-MIBG is not uniformly available, CVD is the mainstay of systemic treatment, although predictors of response remain limited. Our objectives were to describe the clinical characteristics of a cohort of patients with malignant PCC/PGL and to determine CVD-specific PFS in order to make recommendations regarding clinical management. We performed a retrospective IRB-approved study of all patients with PCC/PGL seen at the Hospital of the University of Pennsylvania Neuroendocrine Tumor (NET) Center between January 2000 and August 2016. Malignant PCC/PGL was defined as the presence of distant metastases or extensive unresectable local invasion into adjacent organs or tissues. Demographic information was collected including dates of initial diagnosis, metastatic disease, treatment initiation and clinical progression. When dates with only month or year were available, data was imputed to the mid-day or month. Not all patients had imaging studies performed at our institution; therefore, we could not use Response Evaluation Criteria In Solid Tumors (RECIST) criteria to define progression. Instead, progression was defined as clinical disease progression, initiation of subsequent treatment modality or death due to disease. Our clinical practice is to refer all patients with PCC/PGL seen in the NET Center for genetic counseling and testing (Fishbein et al. 2013). However, not all patients had clinical genetic testing, especially in the first half of the time period. Demographic and clinical characteristics were described with summary statistics. T-tests or ANOVA with Bonferroni correction were used to detect differences between group means. Categorical variables were evaluated using a Fisher’s exact test. Kaplan–Meier analysis with a log-rank test for P values was used for overall survival and PFS analyses. P values less than 0.05 were considered statistically significant. All statistics were performed using STATA IC12 (StataCorp, College View, TX, USA). We retrospectively reviewed the charts of 71 consecutive patients with metastatic PCC/PGL who presented between January 2000 and August 2016 (Table 1). The cohort consisted of 48% women with a mean age at initial diagnosis of 41 years (range 9–73). Of the 60 patients (85%) who had clinical genetic testing, 37 (62%) had an identified susceptibility gene mutation, 32 of which were in SDHB. The entire cohort had a mean follow-up time of 11.23 years. Eighteen patients (25%) were diagnosed with metastatic disease within three months of initial diagnosis of PCC/PGL, and of the fourteen tested, nine had SDHB mutations and five had no identified mutation. For the cohort of 71 patients, the mean time from initial diagnosis of PCC/PGL to development of metastatic disease was 6.2 years (range 0–44.77) and did not differ based on SDHx mutation status (SDHx carriers 6.80 years vs other 6.39 years; P = 0.8632) or SDHB mutation status (SDHB carriers 5.99 years vs other 248

Muscle oxidative phosphorylation quantitation using creatine chemical exchange saturation transfer (CrCEST) MRI in mitochondrial disorders

Systemic mitochondrial energy deficiency is implicated in the pathophysiology of many age-related human diseases. Currently available tools to estimate mitochondrial oxidative phosphorylation (OXPHOS) capacity in skeletal muscle in vivo lack high anatomic resolution. Muscle groups vary with respect to their contractile and metabolic properties. Therefore, muscle group-specific estimates of OXPHOS would be advantageous. To address this need, a noninvasive creatine chemical exchange saturation transfer (CrCEST) MRI technique has recently been developed, which provides a measure of free creatine. After exercise, skeletal muscle can be imaged with CrCEST in order to make muscle group-specific measurements of OXPHOS capacity, reflected in the recovery rate (τCr) of free Cr. In this study, we found that individuals with genetic mitochondrial diseases had significantly (P = 0.026) prolonged postexercise τCr in the medial gastrocnemius muscle, suggestive of less OXPHOS capacity. Additionally, we observed that lower resting CrCEST was associated with prolonged τPCr, with a Pearsons correlation coefficient of -0.42 (P = 0.046), consistent with previous hypotheses predicting that resting creatine levels may correlate with 31P magnetic resonance spectroscopy-based estimates of OXPHOS capacity. We conclude that CrCEST can noninvasively detect changes in muscle creatine content and OXPHOS capacity, with high anatomic resolution, in individuals with mitochondrial disorders.

Development of new preclinical models to advance adrenocortical carcinoma research

Adrenocortical cancer (ACC) is an orphan malignancy that results in heterogeneous clinical phenotypes and molecular genotypes. There are no curative treatments for this deadly cancer with 35% survival at five years. Our understanding of the underlying pathobiology and our ability to test novel therapeutic targets has been limited due to the lack of preclinical models. Here, we report the establishment of two new ACC cell lines and corresponding patient-derived xenograft (PDX) models. CU-ACC1 cell line and PDX were derived from a perinephric metastasis in a patient whose primary tumor secreted aldosterone. CU-ACC2 cell line and PDX were derived from a liver metastasis in a patient with Lynch syndrome. Short tandem repeat profiling confirmed consistent matches between human samples and models. Both exomic and RNA sequencing profiling were performed on the patient samples and the models, and hormonal secretion was evaluated in the new cell lines. RNA sequencing and immunohistochemistry confirmed the expression of adrenal cortex markers in the PDXs and human tumors. The new cell lines replicate two of the known genetic models of ACC. CU-ACC1 cells had a mutation in CTNNB1 and secreted cortisol but not aldosterone. CU-ACC2 cells had a TP53 mutation and loss of MSH2 consistent with the patients known germline mutation causing Lynch syndrome. Both cell lines can be transfected and transduced with similar growth rates. These new preclinical models of ACC significantly advance the field by allowing investigation of underlying molecular mechanisms of ACC and the ability to test patient-specific therapeutic targets.

Chromaffin cell biology: inferences from The Cancer Genome Atlas

Pheochromocytomas and paragangliomas (PCC/PGLs) are rare neuroendocrine tumors that are unusually diverse in metabolic profiles, in classes of molecular alterations and across a large number of altered genes. The Cancer Genome Atlas (TCGA) comprehensively profiled the molecular landscape of PCC/PGLs and identified novel genomic alterations and a new molecular classification of PCC/PGLs. In this review, we discuss the significant clinico-molecular findings of this integrated profiling study. We then review the molecular data of the TCGA cohort centering around known markers of sympathoadrenal cell lineage to better understand chromaffin cell biology. This analysis adds a new layer, that of chromaffin cell type, onto the published molecular classifications and in doing so provides inferences about underlying chromaffin cell biology and diversity.

Paclitaxel is necessary for improved survival in epithelial ovarian cancers with homologous recombination gene mutations

PURPOSE To investigate the impact of somatic mutations in homologous recombination (HR) genes on the chemotherapeutic response and survival of patients with epithelial ovarian cancer (EOC). EXPERIMENTAL DESIGN We performed targeted massively parallel sequencing of tumor DNA from 158 patients with EOC. We associated adjuvant chemotherapy and clinical outcome with mutations in selected genes, focusing on those encoding HR proteins. RESULTS HR mutations were found in 47 (30%) tumors. We did not detect an overall survival (OS) difference in advanced stage patients whose tumors had HR mutations compared to those without (median OS of 49.6 months (95% CI 29.9-57.7) vs. 43.3 months (95% CI 31.9-75.47), p = 0.87). However, when stratified by chemotherapy regimen, patients whose tumors had TP53 and HR mutations demonstrated a marked survival advantage when treated with platinum and paclitaxel vs. platinum +/− cyclophosphamide (median OS of 90 months (95% CI 50-NA) vs. 29.5 months (95% CI 17.7-50.5), p = 0.0005). CONCLUSIONS Previous studies demonstrating a survival advantage for EOC patients with somatic HR mutations have been conducted with almost universal use of both platinum and paclitaxel. Our study is the first to our knowledge to compare cohorts with somatic HR gene mutations treated with and without paclitaxel containing platinum regimens. The survival benefit attributed to the platinum sensitivity of HR deficient ovarian cancers may depend upon the combined use of paclitaxel.