OR35-04 Anti-Müllerian Hormone Variants identified in Patients with Polycystic Ovary Syndrome Affect Secretion and Immunoactivity

Disclosure: L. Meng: None. A. McLuskey: None. A.E. Dunaif: None. J.A. Visser: None. Recently rare heterozygous AMH variants were identified in women with PCOS, causing reduced AMH signaling. However, the exact functional mechanism remains unknown. Here, we analyzed the processing, secretion and signaling of these AMH variants. Six PCOS-specific AMH variants (V(12)G, P(151)S, P(270)S, P(352)S, P(362)S, H(506)Q) and one control-specific variant (A(519)V) were introduced in an AMH expression vector. Signaling activity was measured using the BRE-Luc reporter in the mouse granulosa cell-line KK-1. AMH cleavage and secretion was analyzed by Western Blot, confocal microscopy and ELISA. Serum AMH levels in PCOS cases harboring these variants were measured by ELISA. hAMH-(151)S and hAMH-(506)Q have ∼90% decreased AMH signaling (P<0.001). The other five variants were comparable to wild-type (wt)-AMH signaling. Coexpression of hAMH-(151)S or hAMH-(506)Q with wt-hAMH at equal amounts inhibited wt-hAMH signaling by ∼30% (P<0.001). Western blot analysis revealed that hAMH-(151)S and hAMH-(506)Q proteins were only detected in the cell lysate but not in the supernatant, while wt-hAMH and the hAMH-(352)S and hAMH-(362)S variants were detected in both the cell lysate and the supernatant. Confocal analysis showed that cells expressing hAMH-(151)S and hAMH-(506)Q had higher cellular AMH protein levels with ER retention compared to cells expressing wt-hAMH and hAMH-(352)S. In agreement, using two commercial AMH ELISA’s, hAMH-(151)S was readily detected in the cell lysate, while only very low levels were detected in the supernatant. Both hAMH-(362)S and hAMH-(519)V were detectable using the Fujirebio-AMH ELISA, but showed severely reduced immunoactivity measured by the pico-AMH ELISA. Surprisingly, hAMH-(506)Q was undetectable in both the cell lysate and supernatant by both ELISA’s. However, in PCOS cases, heterozygous carriers of the P(151)S and H(506)Q variants still had detectable AMH in both assays. In conclusion, our results show that P(151)S and H(506)Q disrupt normal processing and secretion of AMH, leading to hampered secretion of wt-AMH. Additionally, AMH variants impair the AMH immunoactivity with (H(506)Q) or without (P(362)S and A(519)V) influencing their bioactivities. Presentation Date: Sunday, June 18, 2023