Studies in a Murine Model Confirm the Safety of Griffithsin and Advocate Its Further Development as a Microbicide Targeting HIV-1 and Other Enveloped Viruses

Kouokam, Joseph Calvin; Lasnik, Amanda B.; Palmer, Kenneth E.

Since we did not observe any harmful effects after a single dose of 10 mg/kg in vivo, we were interested in evaluating the potential toxicity of single s.c. injection of a higher dose of 50 mg/kg GRFT in mice. The survival rate in this experiment was 100%, and the animals treated with 50 mg/kg GRFT did not show any changes in behavior or body weight ( Figure 4B ). These findings suggested that single doses of GRFT at 50 mg/kg did not cause massive toxicity when injected into mice. Mice were then euthanized one, seven, and 14 days post treatment. At all time-points, heart, lung, liver, and kidney weights were unaffected by the presence of GRFT (data not shown). However, spleen size and weight were increased for the animals sacrificed one day post treatment with a mean spleen body weight ratio of 0.56% ± 0.06% for GRFT-treated mice vs. 0.36% ± 0.02% for the PBS group ( Figure 4E ). Although decreasing, the splenomegaly persisted for one week after treatment with ratios of 0.52% ± 0.08% and 0.36% ± 0.03% for GRFT and PBS groups, respectively ( Figure 4E ). Interestingly, GRFT-treated animals had spleens comparable in size and weight to those of the PBS group after two weeks of recovery (ratios of 0.38% ± 0.03% for GRFT animals and 0.35% ± 0.03% for PBS controls) as shown in Figure 4F . Blood chemistry parameters in mice revealed no significant treatment-related differences between GRFT and control groups. Likewise, the functions of kidney (BUN and CRE), pancreas (AMY, GLU), immune system (GLOB), and most indicators of liver function (ALB, TBIL, and ALT) were not adversely affected by GRFT treatment (Table S1 ). However, a non-significant increase was observed in ALP levels in samples collected one week after GRFT treatment (147.3 ± 7.8 arbitrary units (AU) vs. 100.0 ± 33.1 AU for PBS controls, p = 0.07). Two weeks after administration the same trend was observed with 144.2 ± 49.3 AU recorded in the GRFT group and 100.0 ± 5.3 AU for PBS treated animals (p = 0.1933). Furthermore, blood chemistry profiles revealed no effect of GRFT on the nutritional status (TP), calcium (Ca), phosphorous (PHOS), and plasma sodium (Na) (Table S1 ). However, a non-significant increase was noticeable in cholesterol amounts, with 107.0 ± 48.6 mg/dL in GRFT-treated animals compared with 63.3 ± 9.1 mg/dL found in the control group (p = 0.1817) after two weeks of recovery (Table S1 ). Blood samples were submitted to complete blood count (CBC) and there were no significant variations in the hematological profile due to GRFT treatment. Indeed, the values obtained after CBC for white blood cells (total leucocytes, NE, LY, MO, EO, and BA), erythrocytes (total count, HGB, HCT, MCV, MCH, MCHC, and RDW), and platelets (total count and mean platelet volume) were similar in both GRFT-and PBS-treated groups (Table S2 ).