br resulted in decreased anti apoptotic MCL expression In
resulted in decreased anti-apoptotic MCL1 expression. In contrast, the expression of pro-apoptotic BAX and BIM were increased upon metformin treatment. Part of the eﬀects of metformin may be due to miR-26a expression as metformin increased its expression. miR-26a mimics also decreased MCL1 expression and increased the induction of apoptosis in these Midostaurin (PKC412) (Wang et al., 2016).
Metformin administration may have eﬀects on miR-155/miR-144 which is important in the regulation of mitochondrially-bound hexokinase II (mtHKII). mtHKII has important roles in the resistance of cancer cells in avoiding cell death as well as protecting against ischemia-reperfusion (IR) injury. Some of the eﬀects of the cardioprotection against reperfusion were shown to be due to the mi-tochondria, such as preventing outer mitochondrial membrane breakage and subsequent cytochrome C release (Nederlof et al., 2014).
BBR is consumed by millions of people as it is contained in fruits. It is also prepared as a dietary supplement and present in capsules. As a nutraceutical, BBR is consumed for the prevention and suppression of various diseases. Various attempts by multiple investigators have been performed to increase BBR's eﬀectiveness. In our studies, we examined the eﬀects of modified BBRs (NAX compounds) on colony formation in four diﬀerent PDAC cell lines. Some of the NAX compounds inhibited proliferation more than other NAX compounds and the parental BBR both in terms of MTT assays and IC50 determination and in colony formation abilities (Abrams et al., 2018b). Interestingly, the eﬀects of NAX038 and NAX060 compounds were similar on all four PDAC cell lines in terms of colony formation assays. Metformin increased the ability of some, but not all, NAX compounds to inhibit proliferation.
MIA-PaCa-2 cells have a gain of function mutation at TP53 (Deer et al., 2010) and an activating mutation at KRAS. We have examined previously the eﬀects of metformin and BBR on these cells (Abrams et al., 2018a; Abrams et al., 2018b; Candido et al., 2108; McCubrey et al., 2018). AsPC-1 and BxPC3 cells also have TP53 mutations. AsPC-1 has mutations at KRAS while BxPC3 is reported to have WT-KRAS. miR pathways would be predicted to be dysregulated in these PDAC cells as TP53 is mutated and miRs can also influence the KRAS pathway. Since TP53 and KRAS are frequently mutated in PDAC, elucidating the eﬀectiveness of drugs and nutraceuticals which can function in the presence of these mutations is important.
The increased eﬀectiveness of certain NAX compounds in the presence of metformin in inhibiting proliferation could be due to multiple mechanisms. Some of the more eﬀective NAX compounds could induce more DNA damage which could synergize with the eﬀects of metformin and result in cell death. Some of the NAX compounds could induce more ROS or WT-TP53 than others which would stimulate the induction of miRs and apoptosis. The induction of autophagy is also a possible mechanism which could explain the diﬀerences of the NAX compounds eﬀectiveness. Elucidation of the mechanisms by which the diﬀerent NAX compounds in
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Fig. 15. Eﬀects of the NAX077 and NAX111 Compounds on the Proliferation and IC50s of MIA-PaCa-2 + pLXSN (Adh.) and MIA-PaCa-2 + WT-TP53 (Adh.) Cells in the Absence and Presence of 250 nM Metformin. Panel A) MTT analysis of MIA-PaCa-2 + pLXSN (Adh.) cells plated with diﬀerent concentrations of NAX077 (solid red squares) or diﬀerent concentrations of NAX077 and 250 nM metformin (solid blue triangles). Panel B) MTT analysis of MIA-PaCa-2 + WT-TP53 (Adh.) cells plated with diﬀerent concentrations of NAX077 (solid red squares) or diﬀerent concentrations of NAX077 and 250 nM metformin (solid blue triangles). Panel C) MTT analysis of MIA-PaCa-2 + pLXSN (Adh.) cells plated with diﬀerent concentrations of NAX111 (solid red squares) or diﬀerent concentrations of NAX111 and 250 nM metformin (solid blue triangles). Panel D) MTT analysis of MIA-PaCa-2 + WT-TP53 (Adh.) cells plated with diﬀerent concentrations of NAX111 (solid red squares) or diﬀerent concentrations of NAX111 and 250 nM metformin (solid blue triangles). All the experiments indicated in this figure were performed on the same day. These experiments were repeated 3 times and similar results were obtained.