Table 4.

Prediction of pathogenicity in novel germline CACNA1D variants using information from somatic mutations in APAs and APCCs

Classification	Criteria
Pathogenic	If absent in controls and typical gating changes (Table 2) have been observed in functional studies
	If absent in controls and has been described independently in at least two different adrenals
	If present in a single control, described independently in at least two different adrenals and typical gating changes (Table 2) have been observed in functional studies
	If absent in controls and has been described in only one adrenal but at least one additional variant (pathogenic/likely pathogenic) is described in the same position
Likely pathogenic	If present in a single control and described independently in at least two different adrenals
	If absent in controls and has been described in only one adrenal
Likely benign	If present in more than 3 controls and only one adrenal
	If the variant likely represents a null variant (loss of channel function, nonsense, frameshift, canonical ± 1 or 2 splice sites, initiation codon, single or multiexon deletion)
Uncertain	If present in one control and described in only one adrenal
	If present in more than one control and described in more than one adrenal

The criteria listed in the table were used to classify the pathogenicity of somatic mutations in APAs and APCCs assuming that some of them cannot (yet) be considered pathogenic mutations causing high risk for disease based e.g. on the frequency of them being independently reported and their occurrence in healthy control individuals. Note that this classification should aid in predicting the potential pathogenicity of germline mutations for endocrine or neurodevelopmental syndromes associated with CACNA1D variants. In the absence of a suitable guideline for classifying the potential pathogenicity for somatic tumor mutations [38], we consider the here described criteria for classification of the pathogenicity of CACNA1D APA/APCC variants