Timing Is Everything In Protein Formulation by Angelo DePalma, Ph.D.

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Feature Articles : Sep 15, 2011 (Vol. 31, No. 16)

Timing Is Everything in Protein
Formulation
Choosing When and How to Proceed Depends on Available Resources and
Competition
Angelo DePalma, Ph.D.

Formulation can make or break a development-stage protein drug, and add years of IP
protection and value for post-patent molecules. Yet significant disagreement exists on
the best time to undertake the formulation investment.

Robert E. Zoubek, Ph.D., head of protein characterization and preformulation at
Formycon, a business unit of Scil Technology, advises customers to complete final
formulation before Phase I.

He recognizes the dilemma of investing in formulation development before knowing if
the drug will even make it to Phase II. “However, the reason many monoclonal
antibodies fail is specifically because they are improperly formulated in the first place.”

Moreover, the moving-target approach to formulation, whereby tweaks are introduced
over time, can be risky from a regulatory perspective. “Regulators have been known to
ask for more tests on suboptimal formulations,” Dr. Zoubek notes.

“I know of one company that was asked to redo preclinical studies just as they were
preparing to enter Phase II. Regulators wanted proof that the product as it was
formulated at the time would not harm pregnant animals.”

He also cites ethical reasons for formulating from the start. It’s not right to subject Phase
I subjects to a substandard formulation, he says, “just to save money.”

Dr. Zoubek believes the time for considering reformulation as a lifecycle-extending
strategy depends on the therapeutic area and level of competition. Suboptimal
formulations put companies at a competitive disadvantage, so it is never too early to
consider reformulation.

However, all things being equal, companies need to factor in time-to-market, including
any human studies that will be required. One and a half years before patent expiration is
not too early, he says.

Sponsors should add time, possibly more than a year, when lifecycle extension includes

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more exotic, specialty-type extenders including nanoparticles, liposomes, and
bioresorbable ceramics. These formulations will require more testing and more
sophisticated analytics than straight-up liquid formulations. The same is true for
lyophilization.

Since formulation development typically occurs during early clinical stages (or, optimally,
even earlier)—when sponsors are scrambling to manufacture product for testing—
material limitations can pose serious challenges. This is particularly true for products
that will be formulated at high concentration.

High Throughput or "Rational"?
From his perspective as principal scientist at Cook Pharmica, Zachary Yim, Ph.D., is
seeing more combination products than ever before, such as Fc fragments coupled with
enzymes, small molecules, or peptides linked to proteins. “Formulation of these products
is complex and multifaceted because you have to deal with both components.”

As a CMO/CRO, Cook sees formulation-development strategies of every type, duration,
budget, and timeline. Dr. Yim says that formulation is often defined by when toxicology
studies or clinical trial manufacturing must be completed. Cash-strapped sponsors do
the best they can to meet minimal stability requirements.

“On the other hand,” says Dr. Yim, “those with more resources can think longer-term,
because ideally you don’t want to change the formulation unless you have to.”

Whatever the strategy, many formulation developers today employ some type of design
of experiment and high-throughput method to arrive at formulation candidates and,
eventually, to select among and optimize these choices. Yet high-throughput methods
can still take considerable time and always generate hundreds or thousands of
formulations.

Dr. Yim suggests taking a tiered or stepwise approach that might begin with testing
critical factors like pH or ionic strength, identifying optimized values and trouble spots,
then tackling other conditions. “Doing a pH screen at the beginning could save a
considerable amount of work.”

Then there is the issue of breadth vs. depth. When evaluating a very large number of
conditions at once through high-throughput methods, it’s difficult to devote as much time
per condition as, say, through a more rational or stepwise approach to formulation
development.

“Detailed observation is often lost with robotics,” Dr. Yim explains. “Something can occur
transiently and you miss it. It becomes a tradeoff. In addition, since all parameters are
linked, you cannot change just one parameter at a time.”

High-Concentration Proteins
The majority of protein therapeutics are formulated for injectable or infusible
applications, explains Vadim Klyushnichenko, Ph.D., vp for preclinical services and
process development at Paragon Bioservices.


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“Since alternative delivery methods like transdermal, oral, and inhalable are limited by
protein size, stability, and bioavailability, and many biotech drugs are administered at
high dose, formulations must accommodate very high drug concentrations.”

Formulation developers need to navigate the thin line between concentration and
aggregation or precipitation, both of which are common with monoclonal antibodies.
High-dose therapeutic proteins can also cause side effects, for example infusion
reactions.

Paragon originally focused on development and manufacturing for therapeutic proteins,
monoclonal antibodies, viruses, and virus-like particles. Formulation development was
part of these services.

When their clients began asking for full biologics development, including formulation
development of drug substance and drug product, short- and long-term stability studies,
and fill/finish, the company doubled the size of its facilities to more than 50,000 square
feet. The new space, which includes new formulation labs, stability chamber area, and
cGMP fill/finish, is expected to come online next month.

From Paragon’s perspective, approaches for achieving stable, high, and effective
concentrations for protein drugs lie in sustained-release technologies such as micro-
and nanoparticles, liposomal complexes, or biodegradable matrices for implantable
depot delivery.

“Development of such systems for targeted delivery of therapeutic proteins is the goal of
formulation groups across the biopharmaceutical industry,” Dr. Klyushnichenko adds.

Formulation development should be initiated as early as possible, he says, if only
through preliminary preformulation studies or protein characterization, analytical
development, degradation studies, short-term stability studies, and development of a
preliminary drug substance formulation.

According to this model, developers will investigate several “preformulations” for each
potential formulation type and delivery mechanism. Based on data derived from actual
formulation, long-term stability studies, and preclinical data, the number of candidate
formulations is reduced by the time the product enters Phase I studies.

But Dr. Klyushnichenko warns companies to avoid a common catch-22. “If you do not
know the properties of the protein, you cannot define the formulation and calculate
accurately the quantity of drug substance required for preclinical or early clinical trials.”

This will lead to a “simple and reliable” formulation for Phase I, and perhaps Phase II as
well, he says. The formulation changes subsequently as data from human studies
become available.

Formulation development continues through the entire development period and, in many
cases, throughout the drug’s entire lifecycle. Ideally the same protein could be
formulated to cover several drug forms, administration routes, or indications. Each
additional formulation provides extra IP protection and market coverage.


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“There is always room for improvement in terms of additional patient safety and stable
revenue for the company,” Dr. Klyushnichenko adds.

Crystalline Work-Around
Althea’s protein crystal technology supports the switch from intravenous to highly
concentrated subcutaneous therapeutic protein delivery, and with it self-administration,
long-acting formulation, and lifecycle extension. Protein crystals are related to but
should not be confused with CLECs (crosslinked enzyme crystals), which Althea
inherited through its acquisition of Altus Biologics.

Protein crystal formulations are delivered as suspensions of 5–10 micron particles, not
solutions. When formulated as suspended solids, proteins are far less prone to stability
and aggregation issues. Since the drug is significantly less viscous than highly
concentrated protein solutions, it may be delivered through a fine-gauge needle.

For example, it takes less than one minute to load a 1 mL syringe with crystalline
infliximab (Remicade) at a nominal concentration of 200 mg/mL. By comparison, a 1 mL
syringe of soluble infliximab at only 150 mg/mL takes 20 minutes to load. Similarly,
injection of the crystalline product takes 20 seconds compared with 100 seconds for the
solubilized protein.

John Hicks, director of corporate development at Althea, is quick to note that alternative
subcutaneous injection formulations exist, “although there aren’t many, and few achieve
high drug concentrations.” Most notable is Halozyme’s partnership with Roche to
develop subcutaneous Rituxan and Herceptin using a recombinant hyaluronidase
technology.

“It appears that they can deliver subcutaneously,” Hicks says. “However with Herceptin,
Roche had to invest $185 million on a proprietary delivery device to target 5–10 minute
administration of injections as high as 12 mL. Pain becomes a serious issue with
injections significantly larger than 1.5 mL.”

The switch to subcutaneous administration involves an IP or lifecycle component—
companies hurrying to switch before their IV patents expire. But healthcare economics is
undoubtedly the true driving force. “Why bring a patient into a hospital or an infusion
center when the treatment can be administered in an office or even self-administered?”

Crystalline suspensions don’t necessarily require longer development times than
conventional platforms, but they do require a somewhat different skill set. Chemists may
use any reagent they like to produce crystals, whereas bioprocessors are restrained to
pharmaceutically acceptable buffers under conditions that promote rapid crystallization
at high yield. Althea claims yields of greater than 90% in 24 hours.

Protein crystallization does not change the state of the native protein, yet Althea has met
with resistance to the technology.

“There’s a knee-jerk reaction against injecting crystals, most likely based on the
potential for immunogenicity. We haven’t seen any in the weekly hGH trials so far,” says
Hicks. “Besides, insulin crystals have been used for years on a daily basis, for example


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Lilly’s Humalog, which is a mix of soluble and crystalline insulin.”

The Super-Excipient
Albumin is a critical ingredient for cell culture, formulated proteins, and
other biotech products. It binds to and stabilizes important molecules
and ions, prevents aggregation and nonspecific adsorption onto glass,
and serves as an antioxidant.

Aralast (Baxter), Kogenate (Abbott), Intron A (Merck), and Avonex
(Biogen Idec) are just a few of the products formulated with albumin as
an excipient.

Because of cost, bovine albumin was at one time preferred over human
albumin. However, the trend toward serum-free and animal
component-free media, and injected/infused products, has led to
significant demand for recombinant human albumin (rAlbumin).

“Albumin is an extremely useful excipient in protein drug formulations,
particularly for drugs that are self-administered,” says Dermot Pearson,
marketing director at Novozymes Biopharma.

“There is no such thing as a universal excipient, but albumin achieves
several formulation objectives compared with SADs.” Pearson is
referring here to the acronym for common bioformulation excipients:
sugars, amino acids, and detergents.

SADs are known to stabilize products in lyophilized forms, but their
ability to stabilize proteins formulated as liquids has yet to be proved on
a broad basis, according to Pearson.

Novozymes specializes in recombinant human albumin manufactured
in yeast. Recombumin®, the firm’s lead product, and albucult® are
used in drug and vaccine manufacture.

Recombinant human albumin can assist in both initial formulation and
lifecycle-management reformulations. Novozymes’ customers are
becoming interested in fine-tuning their formulations at earlier stages in
the clinical development process.

For companies producing monoclonal antibodies, recombinant albumin
can help resolve formulation issues earlier, according to Pearson,
particularly when initial efforts using SADs fail.

Similarly, he maintains that the excipient “can help build security and
maintain market share and exclusivity” for products nearing patent
expiration.


Timing Is Everything In Protein Formulation by Angelo DePalma, Ph.D.

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Timing Is Everything In Protein Formulation by Angelo DePalma, Ph.D.