Reference intervals for plasma, urinary, and salivary concentrations of free metanephrines in dogs:Relevance to the diagnosis of pheochromocytoma

Background: Measurement of free metanephrines is recommended for screening of pheochromocytoma (PCC) but requires appropriate reference intervals (RIs). Hypothesis/Objectives: To report RIs for plasma, urinary and salivary concentrations of free metanephrines and to determine the diagnostic performance of plasma free normetanephrine (pNMN) and metanephrine (pMN) concentrations in dogs with PCC, hypercortisolism (HC), and nonadrenal illness (NAI). Animals: Eighty healthy dogs, 11 PCC dogs, 25 HC dogs, 6 NAI dogs. Methods: Plasma, urine, and saliva were collected prospectively from healthy dogs, and free metanephrine concentrations were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, medical records of dogs that had plasma free metanephrine concentrations measured by LC-MS/MS between 2018-2021 were studied retrospectively. Results: The RIs for free metanephrines in plasma, urine and saliva are reported. Dogs with PCC had significantly higher pNMN than dogs with HC (P &lt;.001) and NAI (P =.002). The PCC dogs had significantly higher pMN than HC dogs (P &lt;.001), but not higher than NAI dogs (P =.29). Using the upper reference limit, pNMN (&gt;3.56 nmol/L) showed high sensitivity (100%, 95% confidence interval [CI]: 72-100) and specificity (94%, 95% CI: 79-99) for diagnosis of PCC, whereas pMN (&gt;2.49 nmol/L) showed moderate sensitivity (73%, 95% CI: 39-94) and high specificity (94%, 95% CI: 79-99). Conclusions and Clinical Importance: With establishment of these RIs, biochemical testing for PCC in dogs can be substantially improved. Measurement of pNMN is superior to pMN in dogs with PCC.</p

Reference intervals for plasma, urinary, and salivary concentrations of free metanephrines in dogs:Relevance to the diagnosis of pheochromocytoma

Background: Measurement of free metanephrines is recommended for screening of pheochromocytoma (PCC) but requires appropriate reference intervals (RIs). Hypothesis/Objectives: To report RIs for plasma, urinary and salivary concentrations of free metanephrines and to determine the diagnostic performance of plasma free normetanephrine (pNMN) and metanephrine (pMN) concentrations in dogs with PCC, hypercortisolism (HC), and nonadrenal illness (NAI). Animals: Eighty healthy dogs, 11 PCC dogs, 25 HC dogs, 6 NAI dogs. Methods: Plasma, urine, and saliva were collected prospectively from healthy dogs, and free metanephrine concentrations were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, medical records of dogs that had plasma free metanephrine concentrations measured by LC-MS/MS between 2018-2021 were studied retrospectively. Results: The RIs for free metanephrines in plasma, urine and saliva are reported. Dogs with PCC had significantly higher pNMN than dogs with HC (P &lt;.001) and NAI (P =.002). The PCC dogs had significantly higher pMN than HC dogs (P &lt;.001), but not higher than NAI dogs (P =.29). Using the upper reference limit, pNMN (&gt;3.56 nmol/L) showed high sensitivity (100%, 95% confidence interval [CI]: 72-100) and specificity (94%, 95% CI: 79-99) for diagnosis of PCC, whereas pMN (&gt;2.49 nmol/L) showed moderate sensitivity (73%, 95% CI: 39-94) and high specificity (94%, 95% CI: 79-99). Conclusions and Clinical Importance: With establishment of these RIs, biochemical testing for PCC in dogs can be substantially improved. Measurement of pNMN is superior to pMN in dogs with PCC.</p

Reference intervals for plasma, urinary, and salivary concentrations of free metanephrines in dogs:Relevance to the diagnosis of pheochromocytoma

Background: Measurement of free metanephrines is recommended for screening of pheochromocytoma (PCC) but requires appropriate reference intervals (RIs). Hypothesis/Objectives: To report RIs for plasma, urinary and salivary concentrations of free metanephrines and to determine the diagnostic performance of plasma free normetanephrine (pNMN) and metanephrine (pMN) concentrations in dogs with PCC, hypercortisolism (HC), and nonadrenal illness (NAI). Animals: Eighty healthy dogs, 11 PCC dogs, 25 HC dogs, 6 NAI dogs. Methods: Plasma, urine, and saliva were collected prospectively from healthy dogs, and free metanephrine concentrations were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, medical records of dogs that had plasma free metanephrine concentrations measured by LC-MS/MS between 2018-2021 were studied retrospectively. Results: The RIs for free metanephrines in plasma, urine and saliva are reported. Dogs with PCC had significantly higher pNMN than dogs with HC (P &lt;.001) and NAI (P =.002). The PCC dogs had significantly higher pMN than HC dogs (P &lt;.001), but not higher than NAI dogs (P =.29). Using the upper reference limit, pNMN (&gt;3.56 nmol/L) showed high sensitivity (100%, 95% confidence interval [CI]: 72-100) and specificity (94%, 95% CI: 79-99) for diagnosis of PCC, whereas pMN (&gt;2.49 nmol/L) showed moderate sensitivity (73%, 95% CI: 39-94) and high specificity (94%, 95% CI: 79-99). Conclusions and Clinical Importance: With establishment of these RIs, biochemical testing for PCC in dogs can be substantially improved. Measurement of pNMN is superior to pMN in dogs with PCC.</p

Reference intervals for plasma, urinary, and salivary concentrations of free metanephrines in dogs:Relevance to the diagnosis of pheochromocytoma

Background: Measurement of free metanephrines is recommended for screening of pheochromocytoma (PCC) but requires appropriate reference intervals (RIs). Hypothesis/Objectives: To report RIs for plasma, urinary and salivary concentrations of free metanephrines and to determine the diagnostic performance of plasma free normetanephrine (pNMN) and metanephrine (pMN) concentrations in dogs with PCC, hypercortisolism (HC), and nonadrenal illness (NAI). Animals: Eighty healthy dogs, 11 PCC dogs, 25 HC dogs, 6 NAI dogs. Methods: Plasma, urine, and saliva were collected prospectively from healthy dogs, and free metanephrine concentrations were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, medical records of dogs that had plasma free metanephrine concentrations measured by LC-MS/MS between 2018-2021 were studied retrospectively. Results: The RIs for free metanephrines in plasma, urine and saliva are reported. Dogs with PCC had significantly higher pNMN than dogs with HC (P &lt;.001) and NAI (P =.002). The PCC dogs had significantly higher pMN than HC dogs (P &lt;.001), but not higher than NAI dogs (P =.29). Using the upper reference limit, pNMN (&gt;3.56 nmol/L) showed high sensitivity (100%, 95% confidence interval [CI]: 72-100) and specificity (94%, 95% CI: 79-99) for diagnosis of PCC, whereas pMN (&gt;2.49 nmol/L) showed moderate sensitivity (73%, 95% CI: 39-94) and high specificity (94%, 95% CI: 79-99). Conclusions and Clinical Importance: With establishment of these RIs, biochemical testing for PCC in dogs can be substantially improved. Measurement of pNMN is superior to pMN in dogs with PCC.</p

Laparoscopic vs. open adrenalectomy: perioperative data and survival analysis in 70 dogs with an adrenal tumor

Adrenalectomy is the treatment of choice in case of functional adrenal tumors and malignant adrenal incidentalomas. Laparoscopic adrenalectomy (LA) in dogs has gained popularity in recent years, however, clinical studies on large patient populations are scarce. This retrospective study describes perioperative and recurrence data, survival, and prognostic factors in 70 dogs that underwent LA or open adrenalectomy (OA) in our hospital between 2008 and 2022. Diagnosis was based on history, clinical signs, endocrine function tests and advanced diagnostic imaging. Laparoscopic adrenalectomy was performed in 42 dogs (n = 27 naturally occurring hypercortisolism, n = 4 pheochromocytoma, n = 1 pheochromocytoma with concurrent hypercortisolism, n = 10 incidentaloma) and OA in 28 dogs (n = 22 hypercortisolism, n = 3 pheochromocytoma, n = 3 incidentaloma). Bilateral adrenalectomy was performed in 8/70 dogs. Surgical duration of LA and OA did not differ significantly in unilateral and bilateral procedures (P = 0.108 and P = 0.101, respectively). Systemic hypertension occurred in 7/41 and 1/28 dogs during LA and OA, respectively (P = 0.130). Hypotension occurred in 2/41 and 4/28 dogs during LA and OA, respectively (P = 0.214). A total of 40/42 dogs in the LA group and 27/28 in the OA group survived to discharge (P = 0.810). Mean hospital stay was significantly shorter (P = 0.006) after LA (1.5 days, range 1–3) than after OA (2.2 days, range 1–4). No significant differences were demonstrated between LA and OA groups in recurrence of adrenal-dependent endocrine disease (P = 0.332), disease-free period (P = 0.733) and survival time (P = 0.353). The disease-specific 1-, 2- and 3-year survival rates were 95, 89, and 89% after LA and 92, 88, and 81% after OA. Tumor size was significantly associated with the occurrence of a recurrence. In addition, tumor size had a negative effect on the disease-free period and survival time. This study shows a favorable outcome of both LA and OA in dogs. Based on low perioperative complication rate, short hospitalization time and long-term outcomes comparable to OA in selected cases, the less invasive laparoscopic approach is considered the preferred technique

Transcriptome sequencing reveals two subtypes of cortisol-secreting adrenocortical tumours in dogs and identifies CYP26B1 as a potential new therapeutic target

Cushing's syndrome (CS) is a serious endocrine disorder that is relatively common in dogs, but rare in humans. In ~15%–20% of cases, CS is caused by a cortisol-secreting adrenocortical tumour (csACT). To identify differentially expressed genes that can improve prognostic predictions after surgery and represent novel treatment targets, we performed RNA sequencing on csACTs (n = 48) and normal adrenal cortices (NACs; n = 10) of dogs. A gene was declared differentially expressed when the adjusted p-value was 2 or < −2. Between NACs and csACTs, 98 genes were differentially expressed. Based on the principal component analysis (PCA) the csACTs were separated in two groups, of which Group 1 had significantly better survival after adrenalectomy (p =.002) than Group 2. Between csACT Group G1 and Group 2, 77 genes were differentially expressed. One of these, cytochrome P450 26B1 (CYP26B1), was significantly associated with survival in both our canine csACTs and in a publicly available data set of 33 human cortisol-secreting adrenocortical carcinomas. In the validation cohort, CYP26B1 was also expressed significantly higher (p =.012) in canine csACTs compared with NACs. In future studies it would be interesting to determine whether CYP26B1 inhibitors could inhibit csACT growth in both dogs and humans

Reference intervals for plasma, urinary, and salivary concentrations of free metanephrines in dogs: Relevance to the diagnosis of pheochromocytoma

Background Measurement of free metanephrines is recommended for screening of pheochromocytoma (PCC) but requires appropriate reference intervals (RIs). Hypothesis/Objectives To report RIs for plasma, urinary and salivary concentrations of free metanephrines and to determine the diagnostic performance of plasma free normetanephrine (pNMN) and metanephrine (pMN) concentrations in dogs with PCC, hypercortisolism (HC), and nonadrenal illness (NAI). Animals Eighty healthy dogs, 11 PCC dogs, 25 HC dogs, 6 NAI dogs. Methods Plasma, urine, and saliva were collected prospectively from healthy dogs, and free metanephrine concentrations were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, medical records of dogs that had plasma free metanephrine concentrations measured by LC-MS/MS between 2018-2021 were studied retrospectively. Results The RIs for free metanephrines in plasma, urine and saliva are reported. Dogs with PCC had significantly higher pNMN than dogs with HC (P 3.56 nmol/L) showed high sensitivity (100%, 95% confidence interval [CI]: 72-100) and specificity (94%, 95% CI: 79-99) for diagnosis of PCC, whereas pMN (>2.49 nmol/L) showed moderate sensitivity (73%, 95% CI: 39-94) and high specificity (94%, 95% CI: 79-99). Conclusions and Clinical Importance With establishment of these RIs, biochemical testing for PCC in dogs can be substantially improved. Measurement of pNMN is superior to pMN in dogs with PCC

Laparoscopic vs. open adrenalectomy: perioperative data and survival analysis in 70 dogs with an adrenal tumor

Adrenalectomy is the treatment of choice in case of functional adrenal tumors and malignant adrenal incidentalomas. Laparoscopic adrenalectomy (LA) in dogs has gained popularity in recent years, however, clinical studies on large patient populations are scarce. This retrospective study describes perioperative and recurrence data, survival, and prognostic factors in 70 dogs that underwent LA or open adrenalectomy (OA) in our hospital between 2008 and 2022. Diagnosis was based on history, clinical signs, endocrine function tests and advanced diagnostic imaging. Laparoscopic adrenalectomy was performed in 42 dogs (n = 27 naturally occurring hypercortisolism, n = 4 pheochromocytoma, n = 1 pheochromocytoma with concurrent hypercortisolism, n = 10 incidentaloma) and OA in 28 dogs (n = 22 hypercortisolism, n = 3 pheochromocytoma, n = 3 incidentaloma). Bilateral adrenalectomy was performed in 8/70 dogs. Surgical duration of LA and OA did not differ significantly in unilateral and bilateral procedures (P = 0.108 and P = 0.101, respectively). Systemic hypertension occurred in 7/41 and 1/28 dogs during LA and OA, respectively (P = 0.130). Hypotension occurred in 2/41 and 4/28 dogs during LA and OA, respectively (P = 0.214). A total of 40/42 dogs in the LA group and 27/28 in the OA group survived to discharge (P = 0.810). Mean hospital stay was significantly shorter (P = 0.006) after LA (1.5 days, range 1–3) than after OA (2.2 days, range 1–4). No significant differences were demonstrated between LA and OA groups in recurrence of adrenal-dependent endocrine disease (P = 0.332), disease-free period (P = 0.733) and survival time (P = 0.353). The disease-specific 1-, 2- and 3-year survival rates were 95, 89, and 89% after LA and 92, 88, and 81% after OA. Tumor size was significantly associated with the occurrence of a recurrence. In addition, tumor size had a negative effect on the disease-free period and survival time. This study shows a favorable outcome of both LA and OA in dogs. Based on low perioperative complication rate, short hospitalization time and long-term outcomes comparable to OA in selected cases, the less invasive laparoscopic approach is considered the preferred technique

Whole transcriptome analysis of canine pheochromocytoma and paraganglioma

Pheochromocytomas and paragangliomas (PPGLs) are neuroendocrine tumors arising from the chromaffin cells in the adrenal medulla and extra-adrenal paraganglia, respectively. Local invasion, concurrent disorders, and metastases prevent surgical removal, which is the most effective treatment to date. Given the current lack of effective medical treatment, there is a need for novel therapeutic strategies. To identify druggable pathways driving PPGL development, we performed RNA sequencing on PPGLs (n = 19) and normal adrenal medullas (NAMs; n = 10) of dogs. Principal component analysis (PCA) revealed that PPGLs clearly clustered apart from NAMs. In total, 4,218 genes were differentially expressed between PPGLs and NAMs. Of these, 232 had a log2 fold change of &gt;3 or &lt; −3, of which 149 were upregulated in PPGLs, and 83 were downregulated. Compared with NAMs, PPGLs had increased expression of genes related to the cell cycle, tumor development, progression and metastasis, hypoxia and angiogenesis, and the Wnt signaling pathway, and decreased expression of genes related to adrenal steroidogenesis. Our data revealed several overexpressed genes that could provide targets for novel therapeutics, such as Ret Proto-Oncogene (RET), Dopamine Receptor D2 (DRD2), and Secreted Frizzled Related Protein 2 (SFRP2). Based on the PCA, PPGLs were classified into 2 groups, of which group 1 had significantly higher Ki67 scores (p = 0.035) and shorter survival times (p = 0.04) than group 2. Increased expression of 1 of the differentially expressed genes between group 1 and 2, pleiotrophin (PTN), appeared to correlate with a more aggressive tumor phenotype. This study has shed light on the transcriptomic profile of canine PPGL, yielding new insights into the pathogenesis of these tumors in dogs, and revealed potential novel targets for therapy. In addition, we identified 2 transcriptionally distinct groups of PPGLs that had significantly different survival times

Molecular alterations in dog pheochromocytomas and paragangliomas

Recently, genetic alterations in the genes encoding succinate dehydrogenase subunit B and D (SDHB and SDHD) were identified in pet dogs that presented with spontaneously arising pheochromocytomas (PCC) and paragangliomas (PGL; together PPGL), suggesting dogs might be an interesting comparative model for the study of human PPGL. To study whether canine PPGL resembled human PPGL, we investigated a series of 50 canine PPGLs by immunohistochemistry to determine the expression of synaptophysin (SYP), tyrosine hydroxylase (TH) and succinate dehydrogenase subunit A (SDHA) and B (SDHB). In parallel, 25 canine PPGLs were screened for mutations in SDHB and SDHD by Sanger sequencing. To detect large chromosomal alterations, single nucleotide polymorphism (SNP) arrays were performed for 11 PPGLs, including cases for which fresh frozen tissue was available. The immunohistochemical markers stained positive in the majority of canine PPGLs. Genetic screening of the canine tumors revealed the previously described variants in four cases; SDHB p.Arg38Gln (n = 1) and SDHD p.Lys122Arg (n = 3). Furthermore, the SNP arrays revealed large chromosomal alterations of which the loss of chromosome 5, partly homologous to human chromosome 1p and chromosome 11, was the most frequent finding (100% of the six cases with chromosomal alterations). In conclusion, canine and human PPGLs show similar genomic alterations, suggestive of common interspecies PPGL-related pathways