High expression of AMAP1, an ARF6 effector, is associated with elevated levels of PD-L1 and fibrosis of pancreatic cancer

Abstract

Background: Not merely the onset of immune evasion, but other factors, such as acidosis and fibrosis, are also major barriers in cancer therapeutics. Dense fibrosis is a hallmark of pancreatic ductal carcinoma (PDAC), in which hyperactivation of focal adhesion kinase (FAK) in tumor cells was shown to be crucial. Double mutations of KRAS/ TP53 are characteristic to PDAC. We previously showed that high protein expression of ARF6 and its downstream effector AMAP1, as well as processes involved in the ARF6 activation by cell surface tyrosine kinase receptors, are major targets of the KRAS/TP53 mutations to promote PDAC invasion, metastasis, and immune evasion. This notion was recaptured by KPC mouse model of human PDAC (LSL-Kras(G12D/+); LSL-Trp53(R172H/+)); Pdx-1-Cre). Mechanistically, the ARF6-AMAP1 pathway is primarily involved in cellular dynamics of PD-L1, β1-integrins, and Ecadherin; and hence modulates cell-adhesion properties when ARF6 is activated. Here, with an aim to understand whether the ARF6-AMAP1 pathway is critically involved in the elevated levels of PD-L1 and fibrosis of PDAC, we analyzed relationship between AMAP1 and these malignant phenotypes. Moreover, because the ARF6 pathway may closely be related to focal adhesion dynamics and hence to FAK, we also investigated whether AMAP1 employs FAK in fibrosis. Methods: Clinical specimens, as well as KPC cells/tumors and their shAMAP1 or shFAK derivatives were analyzed. Results: Elevated levels of PD-L1 and fibrosis correlated with poor outcome of our patient cohort, to be consistent with previous reports; in which high AMAP1 expression statistically correlated with the elevated PD-L1 and fibrosis. To be consistent, silencing of AMAP1 (shAMAP1) in KPC cells resulted in reduced PD-L1 expression and fibrosis in their tumors. On the other hand, shAMAP1 only slightly affected FAK activation in KPC cells, and phosphorylated FAK did not correlate with enhanced fibrosis or with poor outcome of our patients. Conclusions: Together with our previous data, our results collectively indicated that the ARF6-AMAP1 pathway, empowered by the KRAS/TP53 mutations, is closely associated with elevated PD-L1 expression and fibrosis of human PDACs, to be recaptured in the KPC mouse model. The ARF6 pathway may promote fibrosis independent of FAK.

Inhibition of mutant KRAS-driven overexpression of ARF6 and MYC by an eIF4A inhibitor drug improves the effects of anti-PD-1 immunotherapy for pancreatic cancer

“…In addition, oncogenic TP53 mutations have been shown to facilitate processes activating ARF6 by external ligands, via their known functions in promoting the expression of platelet-derived growth factor receptor and several enzymes involved in the mevalonate pathway [ 12 ]. Consistently, tumor cells arising in the representative mouse model of human PDAC, namely, KPC mice ( LSL-Kras(G12D/ + ); LSL-Trp53(R172H/ + ); Pdx-1-Cre ), express Arf6 and Amap1 at high levels, and use them to drive processes involved in malignancy, including invasion, immune evasion, and fibrosis in vivo [ 12 , 15 ].…”

“…The ARF6-AMAP1 pathway regulates intracellular dynamics of the β1-integrins, E-cadherin, and PD-L1, and hence modulates cell adhesion to the stroma, as well as to other cells, including immune cells, to promote invasion, metastasis, and immune evasion [12][13][14]. The ARF6-AMAP1 pathway also increases cell-surface levels of the β1-integrins and PD-L1, and also the fibrosis caused by PDAC, which is another barrier for immunotherapy [12,13,15]. This pathway furthermore regulates the intracellular distribution of mitochondria, and is hence indispensable for avoiding mitochondria-based oxidative catastrophe during cell invasion into narrow paths [16].…”

FAK PROTAC Inhibits Ovarian Tumor Growth and Metastasis by Disrupting Kinase Dependent and Independent Pathways

“…FAK was associated with ASAP1 in gastric cancer and associated with poor patient survival, indicating that FAK may play a synergistic role in gastric tumorigenesis and metastasis ( 25 ). ASAP1 was also associated with PD-L1 expression in pancreatic cancer fibrosis ( 33 ). The interaction between the SH3 domain of ASAP1 with the second prolin-rich domain of FAK has been found to contribute to focal adhesion assembly ( 23 ).…”

ARF6-AMAP1 pathway induces pancreatic cancer cells to express immunosuppressive chemokines

“…Moreover, the stimulation of KPC cells by PDGF was not enough to promote the chemokine switching (our unpublished data); and it remains unclear as to what external stimuli and intracellular mechanisms are involved in the chemokine switching, in addition to the ARF6-AMAP1 pathway. Furthermore, as this pathway is associated with several different functions, including enhancement of the cell surface expression of b1-integrins, CA9, and PD-L1 (6)(7)(8)18,19), whether chemokine switching plays a central role in significantly altering the immune properties of the TME requires further investigation.…”