Rajaratinam Harishini, Rasudin Nur Syahmina, Al Astani Tengku Ahmad Damitri, Mokhtar Noor Fatmawati, Yahya Maya Mazuwin, Zain Wan Zainira Wan, Asma-Abdullah Nurul, Fuad Wan Ezumi Mohd
School of Health Sciences, Health Campus, Universiti Sains Malaysia (USM), Kubang Kerian, Kelantan 16150, Malaysia.
Department of Chemical Pathology, School of Medical Sciences, Health Campus, Universiti Sains Malaysia (USM), Kubang Kerian, Kelantan 16150, Malaysia.
Oncol Lett. 2021 Feb;21(2):108. doi: 10.3892/ol.2020.12369. Epub 2020 Dec 11.
Neonatal Nav1.5 (nNav1.5) is the alternative splice variant of Nav1.5 and it has been widely associated with the progression of breast cancer. The immunological context of nNav1.5 with respect to breast cancer metastases remains unexplored. The presence of antibodies against nNav1.5 may highlight the immunogenicity of nNav1.5. Hence, the aim of the present study was to detect the presence of antineonatal Nav1.5 antibodies (antinNav1.5-Ab) in the serum of patients with breast cancer and to elucidate the effects of breast cancer therapy on its expression. A total of 32 healthy female volunteers and 64 patients with breast cancer were randomly recruited into the present study as the control and breast cancer group, respectively. Patients with breast cancer were divided equally based on their pre- and ongoing-treatment status. Serum samples were tested with in-house indirect enzyme-linked immunosorbent assay (ELISA) to detect antinNav1.5-Ab, CD25 (T regulatory cell marker) using an ELISA kit and Luminex assay to detect the expression of metastasis-associated cytokines, such as vascular endothelial growth factor (VEGF), interleukin (IL)-6, IL-10, IL-8, chemokine (C-C motif) ligand 2 and tumor necrosis factor-alpha (TNF-α) The mean difference in the expression of antinNav1.5-Ab among the three groups (control, pretreatment and ongoing-treatment) was significant (P=0.0005) and the pretreatment breast cancer group exhibited the highest expression. The concentration of CD25 was highest in the pretreatment breast cancer group compared with the control and ongoing-treatment groups. There was a significant positive correlation between antinNav1.5-Ab and IL-6 in the pretreatment group (r=0.7260; P=0.0210) and a significant negative correlation between antinNav1.5-Ab and VEGF in the ongoing-treatment group (r=-0.842; P-value=0.0040). The high expression of antinNav1.5-Ab in the pretreatment group was in accordance with the uninterrupted presence of metastasis and highlighted the immunogenicity of nNav1.5 whereas the low expression of antinNav1.5-Ab in the ongoing-treatment group reflected the efficacy of breast cancer therapy in eliminating metastases. The augmented manifestation of T regulatory cells in the pretreatment group highlighted the functional role of nNav1.5 in promoting metastasis. The parallel expression of antinNav1.5-Ab with the imbalanced expression of cytokines promoting metastasis (IL-8, IL-6 and TNF-α) and cytokines that prevent metastasis (IL-10) indicated the role of nNav1.5 in breast cancer growth. The expression of antinNav1.5-Ab in accordance to the metastatic microenvironment indicates the immunogenicity of the protein and highlights the influence of breast cancer therapy on its expression level.
新生儿Nav1.5（nNav1.5）是Nav1.5的可变剪接变体，它与乳腺癌的进展密切相关。nNav1.5在乳腺癌转移方面的免疫背景仍未得到探索。抗nNav1.5抗体的存在可能凸显了nNav1.5的免疫原性。因此，本研究的目的是检测乳腺癌患者血清中抗新生儿Nav1.5抗体（antinNav1.5-Ab）的存在，并阐明乳腺癌治疗对其表达的影响。本研究共随机招募了32名健康女性志愿者和64名乳腺癌患者，分别作为对照组和乳腺癌组。乳腺癌患者根据其治疗前和治疗中的状态平均分为两组。血清样本采用自制的间接酶联免疫吸附测定（ELISA）检测antinNav1.5-Ab，使用ELISA试剂盒检测CD25（T调节细胞标志物），并采用Luminex测定法检测转移相关细胞因子的表达，如血管内皮生长因子（VEGF）、白细胞介素（IL）-6、IL-10、IL-8、趋化因子（C-C基序）配体2和肿瘤坏死因子-α。三组（对照组、治疗前组和治疗中组）中antinNav1.5-Ab表达的平均差异具有统计学意义（P = 0.0005），且治疗前乳腺癌组的表达最高。与对照组和治疗中组相比，治疗前乳腺癌组中CD25的浓度最高。治疗前组中antinNav1.5-Ab与IL-6之间存在显著正相关（r = 0.7260；P = 0.0210），治疗中组中antinNav1.5-Ab与VEGF之间存在显著负相关（r = -0.842；P值 = 0.0040）。治疗前组中antinNav1.5-Ab的高表达与转移的持续存在一致，凸显了nNav1.5的免疫原性，而治疗中组中antinNav1.5-Ab的低表达反映了乳腺癌治疗在消除转移方面的疗效。治疗前组中T调节细胞的增强表现凸显了nNav1.5在促进转移中的功能作用。antinNav1.5-Ab与促进转移的细胞因子（IL-8、IL-6和TNF-α）以及防止转移的细胞因子（IL-10）的不平衡表达平行，表明nNav1.5在乳腺癌生长中的作用。antinNav1.5-Ab的表达与转移微环境一致，表明该蛋白具有免疫原性，并凸显了乳腺癌治疗对其表达水平的影响。
