kD = -12(5) mL/g

kD = -58(5) mL/g

kD = -52(9) mL/g

kD = -59(5) mL/g

Cetuximab (trade name Erbitux®, Eli Lilly) is chimeric murine/human mAb that inhibits

EGFR. It was purchased from a clinical supplier. The cetuximab formulation was stressed

in the presence of the ReForm surfactant alternatives by agitating for 40 hours on an

orbital shaker at 275 rpm at ambient temperature. Particle formation was quantified after

shaking the samples using a FlowCam fluid imaging instrument. The particle counts

obtained in the presence of SR1, SR2, SR3, SR4, and SR5 were normalized to those

obtained in the presence of an equivalent quantity of PS80.

ReForm has developed five classes polysorbate alternatives, SR1, SR2, SR3, SR4, and SR5,

that displace protein from air-water interfaces, effectively preventing particle formation.

Importantly, none of the ReForm surfactant replacement excipient classes contain unstable

ester bonds.

A Fresh Perspective on Biologic FormulationsDaniel G. Greene, Philip Wuthrich, Rosa Casado Portilla, Marie Herring,

Robert P. Mahoney, Sharon Webb, John Sorvillo, and David S. Soane

Formulating biologics at high concentrations is challenging due to their large sizes and complex structures. These features can impart high solution viscosities at elevated

concentrations and make the molecules prone to degradation, which can impact the clinical behavior of the formulations and affect the manufacturing process itself.

However, few formulation strategies exist to address these viscosity and stability problems satisfactorily. As an additional problem, biologic formulations often contain

polysorbate surfactants, which have been implicated in product degradation. Here we offer innovative approaches to both of these formulation issues: we present data

around innovative viscosity modifiers and alternatives to conventional polysorbate surfactants. Importantly, we have selected molecules that have not been used in

biologic formulations previously, but that have well-known toxicology and safety profiles. Our additions to the biologic formulation toolkit can enable alternate

administration routes, improve molecule stability, and provide solutions to important challenges in formulating biologics.

Viscosity and stability

Surfactant replacement

Attractive protein-protein interactions (PPIs) give rise to elevated solution viscosities, which

prohibit self-administration by subcutaneous injection. ReForm Biologics has developed several

chemical structures that minimize PPIs and thus lower viscosity.

Biologics are prone to degradation via adsorption to air-water interfaces, which can lead

to particle formation (grey circles below). Surfactants are added to displace biologics

from interfaces and prevent particle formation. Polysorbates are the most common

surfactants used in industry, but they contain a hydrolysable ester bond (red circle) that

can either thermally hydrolyze or be degraded via enzymatic cleavage. In both cases,

cleavage results in a water-soluble sorbitol derivative and a water-insoluble fatty acid.

The insoluble fatty acid agglomerates into particles, which can be immunogenic.

We don’t make biologics – we make biologics better.

Infliximab and adalimumab model mAb systems

Bovine gamma globulin model system

ReForm Biologics identified four chemical classes, V1, V2,

V3, and V4, that are useful in reducing the viscosity of

highly concentrated protein solutions. Bovine gamma

globulin (BGG) was reconstituted into histidine buffer at

pH 6, containing different excipients to a final

concentration of 280 mg/mL. The viscosity was measured

at 25°C using either a Brookfield cone and plate

rheometer or a RheoSense μVISC viscometer. The

viscosities are normalized to the viscosity of an excipient-

free solution.

Processing benefits

Elevated viscosity can hinder processing steps such as

tangential flow filtration and ultra/diafiltration.

Introducing a viscosity modifier, such as ReForm A, has

the potential to improve these processing steps. In this

example, BGG is concentrated with a centrifugal

concentrator in the presence and absence of 75 mM

ReForm A excipient. The filtrate flux was monitored as

a function of time and the bulk concentration was

calculated from the mass balance. Formulating with

ReForm A improves the final concentration by 46%

(Figure F). This improvement leads to faster

processing times and higher product recoveries.

kD = -5(1) mL/g

kD = 16.7(9) mL/g

ConclusionsReForm Biologics has leveraged its fundamental knowledge of protein-protein and

protein-interface interactions to identify excipients that can decrease viscosity, improve

stability and processability, and replace problematic polysorbates. Viscosity

improvements are achieved by decreasing attractive, while the surfactant replacements

out-compete protein for air-water interfaces. These excipients expand the options for

formulation technologies by solving common and vexing industry problems and can

enable bio-better formulations.

AirAir

PBS + ReForm

SR1, SR2, SR3, SR4, SR5

PBS SR1 SR2 SR3PS80 SR5SR4

Infliximab (trade name Remicade®, Janssen) and adalimumab (trade name Humira®, AbbVie)

are TNFα inhibitors. Infliximab is a chimeric murine/human mAb, while adalimumab is a fully

humanized mAb.

Four infliximab formulations were prepared: 1) a control formulation containing 10 mM acetate,

10 mM phosphate buffer at pH 6, 2) 75 mM ReForm A, 3) 100 mM arginine, and 4) 100 mM NaCl.

Formulations 2, 3, and 4 were prepared in the control buffer. Samples were concentrated using an

Amicon centrifugal concentrator with 30 kDa cutoff and measured for viscosity and diffusivity.

ReForm A belongs to the V1 class of viscosity modifiers and effectively lowered the viscosity

compared to the control, while arginine and NaCl did not (Figure A). ReForm A acts by reducing

attractive PPIs as indicated by the diffusion interaction parameter, kD, which changed in value

from -60 mL/g to -12 mL/g (Figure B).

Adalimumab was formulated in 20 mM

histidine, pH 5 with 77 mM ReForm A (red data

points). Humira® was used as received for the

control curve (black data points). Again, the

samples were measured for viscosity and

diffusivity. As for infliximab, ReForm A

excipient lowers the viscosity of adalimumab

formulations (Figure C) as a result of

decreasing attractive PPIs (Figure D).

Additionally, the ReForm A excipient improved

the stability at 40°C during an accelerated

aging study at elevated protein concentration

(Figure E).

Visit us at reformbiologics.com

for more information and to

download a copy of this poster.