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. 2017 Dec 18;36(16):1594–1602. doi: 10.1200/JCO.2017.76.6915

Fig 4.

Fig 4.

Pharmacodynamic assessment of γH2AX formation in plucked hair bulbs and peripheral blood mononuclear cells (PBMCs) after topotecan and M6620. (A) Diagram depicting the collection of PBMCs and hair follicles. All patients received topotecan on days 1, 2, and 3, with patients 7 to 21 receiving M6620 on day 2. Samples were collected before infusions of topotecan and M6620 (D1-pre), 5 to 30 minutes after day 2 topotecan (D2-post 1), 30 minutes to 1 hour after day 2 M6620 (D2-post 2), before day 3 topotecan (D3-post 1), and 5 minutes after day 3 topotecan (D3-post 2). (B) γH2AX in plucked hairs with all data plotted as γH2AX signal intensities. γH2AX induction in patients who received topotecan only (patients 1 to 6) shows DNA double-stranded breaks after topotecan alone (left; n = 6). Patients who received M6620 plus topotecan on day 2 (patients 7 to 21) had increased DNA damage levels on day 3 compared with day 2, which was not the case for those who received topotecan alone on day 2 (right; n = 12). To understand the kinetics of γH2AX formation further, we obtained hair follicles at additional time points: 30 minutes to 1 hour after day 2 M6620 infusion (D2-post 2) and before day 3 topotecan (D3-post 1). (C) γH2AX in plucked hairs with all data plotted as γH2AX signal intensities in six patients who had additional sampling. Although follicles obtained 30 to 60 minutes after M6620 infusion on day 2 showed a decrease in γH2AX signal, an increase in H2AX phosphorylation was observed in follicles obtained 24 hours after M6620 infusion (before any additional topotecan). (D) Representative images of γH2AX staining in plucked hair bulbs collected from patient 14 (green, γH2AX; red, DNA). Original magnification x63. (E) γH2AX in PBMCs, with all data plotted as γH2AX foci per cells in patients who received topotecan only (left; n = 6) and in patients who received both topotecan and M6620 (right; n = 15). (F) Representative images of γH2AX staining in PBMCs (green, γH2AX; red, DNA). Original magnification x63. P values for the paired differences appear on the plots. The γH2AX signal intensities are expressed in arbitrary unit.