Aesthetic Surgery Journal
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`Injecting Puragen Plus Into the Nasolabial Folds: Preliminary Observations of FDA Trial
`Brian M. Kinney
`Aesthetic Surgery Journal
` 2006 26: 741
`DOI: 10.1016/j.asj.2006.10.008
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`The online version of this article can be found at:
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`H o t T o p i c s
`
`Injecting Puragen Plus into the Nasolabial Folds:
`Preliminary Observations of FDA Trial
`
`Based on participation in ongoing FDA trials, the
`author presents his initial impressions of Puragen Plus
`for treatment of the nasolabial folds. Puragen and
`Puragen Plus (Mentor Corp., Santa Barbara, CA) are
`double–cross-linked NASHA products. Depending on
`double cross-linking for duration of effect, instead of a
`varying particle size, may allow for use of one filler at
`all levels in the soft tissue. Other features observed by
`the author in the clinical setting included reduced
`injection pain, minimal erythema and tenderness, typi-
`cally 9 to 12 months’ duration of effect, and high
`patient satisfaction. (Aesthetic Surg J 2006;26:741–748.)
`
`In the United States, bovine collagen was essentially
`
`the only soft tissue filler on the market from the
`1980s until just a few years ago. In many other
`countries, however, a wide variety of injectable materi-
`als have been long utilized for soft tissue filling.1,2
`Perhaps the most widely used substance today is poly-
`merized chains of hyaluronan, hyaluronic acid (HA).
`Starting with the early 1996 Sweden experience, and
`spreading from Europe to the rest of the world, physi-
`cians have used cross-linked, non–animal source
`hyaluronic acid (NASHA).
`A large body of NASHA clinical experience has grown
`with generally excellent results. In the November/
`December 1999 issue of Aesthetic Surgery Journal,
`Troilius3 reported his initial favorable experience in more
`than 200 patients, using Restylane (QMed, Inc.,
`Eatontown, NJ), a NASHA preparation (mean particle
`size 525 μ) single cross-linked with ether bonds by 1,4-
`butanediol diglycidyl ether (BDDE).
`In December 2003, the Food and Drug Administration
`(FDA) approved Restylane, the first Restylane filler to be
`approved in the United States, and by January 2005, clinical
`use had become common. Advantages of Restylane include
`longer lasting effects than bovine collagen, improved con-
`touring and volume augmentation, increased patient satis-
`faction, and freedom from allergy testing. A major disadvan-
`tage of many HA preparations is the pain associated with
`injection and the need for several different preparations
`
`Brian M. Kinney, MD,
`MSME, Los Angeles, CA, is a
`board-certified plastic surgeon
`and an ASAPS member.
`
`based on particle size (300 to
`650 μ) to allow for injection
`at various tissue depths.
`Additionally, while duration
`of effect is longer than with
`bovine collagen, it still falls
`short of ideal. Restylane Fine
`Lines is recommended for
`superficial use, Restylane for
`deeper use, and Restylane
`SubQ and/or Perlane are rec-
`ommended for use deeper
`than the dermis. However, of
`these preparations, only
`Restylane is cleared for marketing in the United States. All
`of these products contain a concentration of 20 mg/mL.
`Hylaform (Allergan Inc., Irvine, CA) uses single cross-
`linking by divinyl sulphone (DVS), has a mean particle
`size of 692 μ, and has not gained significant market share
`in the United States. Juvaderm (Allergan Inc., Irvine, CA),
`a higher-concentration NASHA preparation with a mean
`particle size of about 594 μ, was approved by the FDA in
`June 2006 and is just coming to market. A major stated
`claim is that Juvaderm is not a gel-particle suspension
`but, instead, a malleable smooth gel that flows more easi-
`ly and with a higher level of control. There are several
`areas in which improved capabilities are desirable. Use of
`a single type of injectable at multiple tissue depths with
`only 1 syringe and 1 hypodermic skin puncture, little or
`no pain associated with the injection, and longer dura-
`tion of effect are important advantages.
`
`Materials and Methods
`
`The half-life of non–cross-linked, naturally occurring
`hyaluronan in the body is 2 to 4 days, and about one
`third is turned over per day. Alteration of the physical
`and chemical properties is required for duration of effect
`in the soft tissues. In creating a synthetic analog, one can
`categorize at least 5 different types for HA products:
`1. Liquid HA
`2.
`Syrup-like HA with higher viscosity
`3. A mix of syrup-like HA and weakly stabilized HA
`particles
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`H O T T O P I C S
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`4. A high concentration of HA particles with HA con-
`centration of heavily cross-linked HA molecules.
`Puragen (Mentor Corp., Santa Barbara, CA) falls
`into this category.5,6
`5. A high concentration of HA particles with high con-
`centration of minimally modified HA molecules. The
`Restylane family of fillers falls into this category.
`One method of increasing duration of effect is to
`vary particle size, as is demonstrated in the Restylane
`family. Another is to alter the chemistry of cross-link-
`ing by creating double-cross-linked chains with ether
`bonds and ester bonds. Puragen and Puragen Plus
`(Mentor Corp., Santa Barbara, CA) are double-cross-
`linked NASHA products. The ester bonds confer
`increased stability in vitro by resisting the enzymatic
`degradation by hyaluronidase and by protecting the
`ether bonds during sterilization. The ether bonds are
`hydrophobic and resist enzymatic degradation. The
`first chemical reaction is performed at high pH with 1,
`2, 7, 8–diepoxyoctane (DEO), a hydrophobic epoxide
`that builds an HA network through ether bonds
`between hydroxyl groups. The second chemical low-pH
`reaction, using the same agent (DEO), further cross-
`links carboxyl groups to form ester bonds. The
`increased chemical stability allows for the addition of
`lidocaine 0.3% for a relatively pain-free injection.
`Enhanced stability in vivo and slower degradation in
`vitro are achieved by double cross-linking. Solid C13
`nuclear magnetic resonance scanning in double-cross-
`linked HA shows a methylene bridge compared with
`standard HA.5 The ester bond is confirmed by Fourier
`transform infrared spectrometry (FT-IR).6
`Using a gel with a smaller average particle size (220 μ)
`may create a smoother injection (more continuous appli-
`cation of pressure). A gel with higher viscosity may
`require more pressure to inject. Depending on double
`cross-linking for duration of effect, instead of a varying
`particle size, may allow for use of 1 filler at all levels in
`the soft tissue.
`Puragen Plus biocompatibility studies were per-
`formed, including skin sensitization in the guinea pig (no
`positive responses) and intradermal implantation in the
`guinea pig (minor initial reactions of erythema seen clini-
`cally and anti-inflammatory and giant cells seen on
`histopathology, minimal or undetectable at 27 weeks,
`similar to 2 comparator products). Also, there was no
`cytotoxicity in the Ames test at concentrations up to
`5000 μg, no cytotoxicity in vitro in an agar overlay
`assay, no mutagenesis in vitro in a chromosomal aberra-
`tion test, no unscheduled DNA synthesis in vitro, no
`
`pyrogenicity (ISO 10993-11), and no hemolysis. No
`necrosis, fibrosis, or granuloma were observed.
`Puragen was introduced into the market in the
`European Union and many countries around the world
`in the spring of 2005. Its formulation is similar to
`Puragen Plus, except that it does not contain lidocaine.
`Puragen Plus has been undergoing FDA clinical trials
`since January 2005 in the United States. Each milliliter of
`Puragen Plus contains 20 mg of ether and ester cross-
`linked sodium hyaluronate 20 mg, sodium chloride 8.5
`mg, disodium hydrogen orthophosphate dehydrate 0.22
`mg, sodium dihydrogen phosphate dihydrate 0.045 mg,
`lidocaine HCl 0.3%, and water for injection.
`
`FDA Clinical Trials
`
`Five centers led by 3 dermatologists and 2 plastic sur-
`geons participated in the original FDA clinical trials. In
`the original (first) study group, patients were randomized,
`Restylane was injected into one nasolabial fold (NLF),
`and Puragen Plus, into the other. None of the 5 centers
`had enough patients to reach statistical significance and
`results have been blinded from center to center.
`In the continuing access (second) study, Puragen Plus
`was injected into both NLFs. Final data analysis has not
`been completed. Here, I report initial clinical impressions
`based on the experience of 1 center in the continuing
`access study. In future publications, I will present the
`compiled data. Study parameters included blood draw,
`photographs, 2 independent observers in the initial
`study, 1 observer (only) in the continuing access study, a
`pain assessment scale, and NLF scoring on a 6-part
`Lemperle scale. Follow-up took place at 14 days (when a
`second injection was allowed), and at 1, 4, 6, 9, and 12
`months in the first study group and at 14 days, and 1, 6,
`9, and 12 months in the second study group.
`In the initial study group each patient wore an eye
`mask, and in both studies each patient was injected after
`a skin prep of alcohol only. For both studies, only
`patients with a Lemperle scale of 2 to 4 were allowed to
`participate. Exclusion criteria included severe skin dis-
`ease (eg, eczema, psoriasis, severe acne, or rosacea), sys-
`temic diseases, history of NLF injection within 18
`months, use of tretinoin within 4 weeks, use of botu-
`linum toxin within 6 months, or laser resurfacing within
`12 months. Blood was drawn at baseline, 30 days after
`the last injection, and at the 6-month visit.
`Patients were injected in the NLF, deep and superfi-
`cially, from the alar base inferior to the oral commissure
`as dictated by fold anatomy. In the first study group, 2
`mL were allowed on each side during the first session,
`
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`H O T T O P I C S
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`B
`
`D
`
`F
`
`A
`
`C
`
`E
`
`Figure 1. A, Pretreatment view of a 41-year-old woman. B, Posttreatment view 1 month following Puragen Plus injection into the nasolabial folds.
`C, Posttreatment view after 3 months. D, Posttreatment view after 6 months. E, Posttreatment view after 9 months. F, Posttreatment view after 1 year.
`(Patient had transient postinjection erythema that lasted from 1 to 2 days.)
`
`Injecting Puragen Plus into the Nasolabial Folds:
`Preliminary Observations of FDA Trial
`
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`H O T T O P I C S
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`B
`
`D
`
`A
`
`C
`
`E
`
`Figure 2. A, Pretreatment view of a 39-year-old woman. B, Posttreatment view 1 month following Puragen Plus injection into the nasolabial folds.
`C, Posttreatment view after 3 months. D, Posttreatment view after 6 months. E, Posttreatment view after 9 months. The patient had deep folds; due to
`volume limitations she did not undergo complete correction in either fold; however, there is broad persistence of HA at 6 months.
`
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`Volume 26, Number 6
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`H O T T O P I C S
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`B
`
`D
`
`A
`
`C
`
`Figure 3. A, Pretreatment view of a 57-year-old man. B, Posttreatment view 1 month after Puragen Plus injection into the nasolabial folds. C,
`Posttreatment view after 3 months. D, Posttreatment view after 6 months. Challenging treatment of a thin male smoker who uses indoor tanning
`equipment 2 to 3 times weekly; results are below average.
`
`and 1 mL was allowed on each side during the second
`session at 14 days. In the second study group, 3 mL were
`allowed on each side during the first session, and 2 mL
`were allowed on each side during the second session at
`14 days. There were no further injections in either study
`group, and follow-up continued for 1 year. Full face and
`close-up photos were taken of each patient at each visit
`in the anterior-posterior, oblique, and lateral views on
`each side. Photos were taken under the same lighting
`conditions, against a Canfield-issued blue textured felt
`background with a Canon G2 Digital camera set on a
`fluorescent 2 white balance setting.
`There were 11 patients in the first group and 25 in the
`second group. In the first group, 1 patient was lost to fol-
`low-up at 6 months when her boyfriend suffered severe
`medical problems requiring hospitalization and intensive
`care. In the second group, 1 patient was lost to follow-up
`
`due to the inconvenience of attending postinjection
`appointments.
`All patients were injected by linear threading, and 1
`or 2 received supplementation by isolated serial puncture
`in inadequately filled areas. Because the length of the
`needle was less than the length of the NLF, more than 1
`puncture was required in all patients. All patients under-
`went massage immediately after injection until smooth
`contours were achieved.
`
`Results and Discussion
`
`Without formal statistical analysis, I can make only
`general clinical observations. Overall satisfaction among
`patients was very high. No patient had a visible nodule
`at any visit, perhaps due to immediate massage on initial
`injection. The gel can be manipulated and contoured eas-
`ily, although on no occasion was it expressed from the
`
`Injecting Puragen Plus into the Nasolabial Folds:
`Preliminary Observations of FDA Trial
`
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`H O T T O P I C S
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`B
`
`D
`
`A
`
`C
`
`Figure 4. A, Pretreatment view of a 44-year-old man. B, Posttreatment view 1 month after Puragen Plus injection into the nasolabial folds.
`C, Posttreatment view after 3 months. D, Posttreatment view after 6 months. Challenging treatment of a heavy-set male with an excellent result.
`
`skin through the puncture site. Not surprisingly, there
`was minimal to no pain in essentially every patient
`injected with Puragen Plus, and less pain in every patient
`injected with Puragen Plus compared with patients
`injected with Restylane. In both groups, mild injection
`erythema of a similar magnitude and similar to that seen
`with other HA preparations occurred in about one third
`of patients and lasted 1 to 2 days. No patient expressed
`regret at undergoing the initial injection in any location.
`On second injection in the initial group, all patients
`expressed a preference for a lidocaine-containing HA.
`Due to volume limitations, complete correction of the
`NLF was not possible in every patient. However, photo-
`graphic documentation allowed comparison of initial
`injection results and late visits at 6, 9, and 12 months.
`In my hands, judging subjectively, Puragen feels
`slightly more viscous on injection and requires more
`
`pressure than injecting Restylane, but I was able to adapt
`to this difference quickly. From the first patient, on ini-
`tial inspection, injection results on either side were simi-
`lar. Of the 36 patients at 6 months, all expressed interest
`in undergoing additional injection with the lidocaine-
`containing HA, if it were available at the completion of
`the study or cleared by the FDA.
`One patient undergoing lip injection had a 2-day
`episode of lip swelling documented by photography 2
`months after injection. Sulfa allergic, she attributed it to
`drinking red wine containing sulphites. No treatment
`was given, and she recovered from the episode without
`sequellae. Afterwards, she expressed a desire for addi-
`tional injection in the lips and NLF should injections
`become available. In the second study group, 2 patients
`reported transient episodes (1 to 2 days) of minor
`swelling between the 1- and 6-month visits, but chose
`
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`Volume 26, Number 6
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`

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`H O T T O P I C S
`
`B
`
`D
`
`A
`
`C
`
`E
`
`Figure 5. A, Pretreatment view of a 57-year-old woman. B, Posttreatment view 1 month after Puragen Plus injection into the nasolabial folds.
`C, Posttreatment view after 6 months. D, Posttreatment view after 9 months. E, Posttreatment view after 12 months.
`
`Injecting Puragen Plus into the Nasolabial Folds:
`Preliminary Observations of FDA Trial
`
`Downloaded from
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`H O T T O P I C S
`
`3. Troilius C. Soft tissue fillers—what options are available today?
`Aesthetic SurgJ 1999;19:505-507.
`
`4. Verpaele A, Strand A. Restylane SubQ, a non-animal stabilized
`hyaluronic acid gel for soft tissue augmentation of the mid- and lower
`face. Aesthetic SurgJ2006;26(suppl):S10-S17.
`
`5. Zhao XB, Fraser JE, Alexander C. Synthesis and characterization of a
`novel double crosslinked hyaluronan hydrogel. J Mater Sci Mater
`Med. 2002;13:11-16.
`
`6. Zhao XB. J Biomater Sci, Polymer ed. In press.
`
`Reprint requests: Brian M. Kinney, MD, MSME, 2080 Century Park E.,
`Suite 1110, Los Angeles, CA 90067-2009.
`Copyright © 2006 by The American Society for Aesthetic Plastic Surgery,
`Inc.
`1090-820X/$32.00
`doi:10.1016/j.asj.2006.10.008
`
`not to report it until the 6-month visit because they con-
`sidered it to be inconsequential. One of these patients
`suggested the swelling was due to gardening outdoors in
`hot weather and had 2 more episodes between the 6- and
`9-month visit. In addition, 1 day after the 9-month visit,
`she experienced an additional episode in the absence of
`exertion in hot weather (documented by photographs)
`that lasted 2 days. Transient nodularity, erythema, and
`tenderness surfaced overnight and dissipated without
`intervention in 2 days. This patient changed her mind,
`now stating that she would not undergo additional injec-
`tions after a 9-month experience.
`More than half of the patients had excellent persis-
`tence at 9 months, and about one quarter had moderate
`persistence even at 12 months. Only 1 or 2 patients
`resorbed the Puragen as early as seen in the 6-month pic-
`tures. Reasonable informed consent would be to predict
`9 to 12 months of persistence (Figures 1 to 5).
`
`Conclusions
`
`I can make a few preliminary observations before the
`formal data analysis; however, much is yet to be deter-
`mined. Injection is minimally painful due to the presence
`of lidocaine. Nine to twelve months of duration of effect
`can generally be expected based on my clinical experi-
`ence. Erythema, tenderness, and pain are minimal. Patient
`acceptance is excellent and satisfaction is extremely high.
`Likelihood of nodularity and other potential complica-
`tions await large-scale follow-up studies.
`The particle size is smaller than in other HA prepara-
`tions and this subjectively allows use of a single syringe
`to inject superficially, in intermediate depths, and deeply
`without expectation of lumpiness, protrusion, or promi-
`nence. Because it is the cross-linking, and not the particle
`size, that dictates the duration of effect, only 1 type of
`syringe is required for injection at various depths for cor-
`rection. The small particle size may allow for better mas-
`saging and manipulation of the material immediately
`after injection, but no blinded control studies have been
`performed to confirm this. If and when there is FDA
`approval, further clinical experience will reveal the com-
`plete picture. ■
`
`The author was paid by Mentor as a consultant to perform
`this clinical trial.
`
`References
`1. Kinney BM, Hughes CE. Soft tissue fillers—an overview. Aesthetic
`SurgJ2001;21:469-471.
`
`2. Bergeret-Galley C. Comparison of resorbable soft tissue fillers.
`Aesthetic SurgJ2004;24:33-46.
`
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`Volume 26, Number 6
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