**MEMORANDUM**

P020012

**SUBJECT**: Preclinical Review

**DEVICE:** Artecoll PMMA/Collagen Implant, P020012

**SPONSOR:** ARTES MEDICAL, INC.

> 4660 La Jolla Village Drive, Suite 825

San Diego, CA. 92122 USA

**INDICATIONS FOR USE:**

Artecoll is indicated for the correction of contour deficiencies of soft
tissue.

**Product Summary:**

Artecoll is a suspension of polymethylmethacrylate (PMMA) microspheres
suspended in a bovine collagen solution containing phosphate buffer,
sodium chloride, lidocaine-HCl and distilled water. The product is
supplied in a four pack of sterile 0.5 ml pre-filled syringes.

**Preclinical Studies:**

While there is no FDA-approved device using the microsphere form of
polymethylmethacrylate (PMMA) polymer, there is a long history of PMMA
use in medical devices. In support of the safety and biocompatibility of
Artecoll, the sponsor submitted the following preclinical studies in
Volume 10 of the original PMA application.

[Final Product Bench testing]{.underline}

-   *Extrusion force* -- The plunger force required to expel 0.5 ml of
    Artecoll through a 27 gauge needle from a standard 1.0 ml syringe
    was determined. The average force for 3 lots of product was 3.1 lbs.
    Based on published literature that suggests that the estimated
    maximum lateral finger/thumb forces for men and women are 23.2 lbs
    and 15.6 lbs, respectively, the force required to expel Artecoll was
    estimated to be approximately 13% and 20% of the average maximum
    lateral finger/thumb forces for men and women, respectively.

-   *Syringe leak* -- The integrity of the interface seal between the
    syringe tip and the syringe cap for Artecoll pre-filled syringes was
    assessed. In tests with 12 devices, each pre-filled syringe was
    found to hold a pressure of 20 pounds per square inch (PSI) for more
    than 5 minutes without a decrease in pressure greater than 1 PSI.
    This reflected a good integrity for the syringe tip/syringe cap
    seal.

[Biological Testing]{.underline} -- Biocompatibility testing for
Artecoll was performed with the final product.

-   *Cytotoxicity* -- A sample of Artecoll was tested based on the
    ISO-10993-5 guidelines. In this test, a positive control (i.e.,
    tin-stabilized polyvinylchloride), a negative control (i.e., high
    density polyethylene) and 0.1 ml of Artecoll were placed on agarose
    surfaces that directly overlayed a confluent monolayer of L-929
    mouse fibroblasts. After incubation at 37C for 24 hours, no evidence
    of cell toxicity (i.e., decolorization or lysis of cells) was
    observed with the control and Artecoll samples. In conclusion, the
    performance of the positive and negative controls demonstrated that
    the test was valid. Artecoll was also found to be less cytotoxic
    than a grade 2 (mild reactivity) material.

-   *Genotoxicity* -- The mutagenicity of Artecoll was assessed in a
    reverse mutation assay with strains of histidine-dependent
    *Salmonella typhimurium* (TA98, TA100, TA1535 and TA1537) and
    tryptophan-dependent *Escherichia coli* (WP2uvrA) in the presence
    and absence of S9 metabolic activation. The assay was conducted as
    described in ISO-10993 guidance. In this test, negative (i.e.,
    saline solution) and positive controls (i.e., p-dimethylaminobenzene
    diazosulfonic acid sodium salt) were also evaluated. The results
    demonstrated that the assay was valid and that Artecoll was
    non-mutagenic to the *Salmonella typhimurium* and *Escherichia coli*
    strains.

-   *Sensitization* -- A guinea pig maximization test was performed with
    Artecoll as described in ISO 10993 Part 10. In this analysis,
    Artecoll was injected i.d. and occlusively patched onto 10 guinea
    pigs, while the control (i.e., 0.9% NaCl USP solution) was applied
    similarly to 5 guinea pigs. After a recovery period, the test and
    control animals were challenged with patches of Artecoll or control.
    All sites were then scored at 24, 48 and 72 hours after patch
    removal. The results of this study demonstrate that Artecoll did not
    cause a delayed dermal contact sensitization reaction.

-   *Implantation in rabbits* -- Muscle tissue implantation studies were
    performed based on the recommendations in ISO 10993, Part 6. In this
    study, Artecoll and negative controls (polyethylene strips) were
    each implanted at 4 different sites in three New Zealand White
    rabbits. After 1 week the animals were euthanized and muscle tissues
    were examined macroscopically and microscopically. Under the
    conditions of this study Artecoll was not found to cause a
    significant macroscopic reaction compared to control.
    Microscopically Artecoll was judged to be a non-irritant.

-   *Implantation in humans and mice* -- In this study cross-linked
    collagen, hyaluronic acid, silicone oil, PMMA microspheres (ranging
    in size from 4-40 microns), PMMA microspheres in hyaluronic acid (40
    microns), polylactic acid microspheres (40 microns), dextran
    microspheres (40 microns), Trisacryl-gelatin microspheres, silicone
    particles, pyrrolytic carbon coated with ZrO beads (212 -- 500
    microns) suspended in 3% beta glucan and polyacrylamide were
    implanted into humans (s.c. into the volar forearm) or mice (see
    below). After injection the clinical appearance of each implant was
    evaluated every other week and excisions were made after 3, 6 and 9
    months. Biopsy samples were HE or Masson Trichrome stained and
    reviewed by a masked pathologist.

> In 44 female mice the same materials were administered either
> subdermally or intramuscularly into the cheek, axilla, groin, urethra
> and quadriceps muscle. After 1, 3, 6 and 9 months one mouse in each
> group was euthanatized and the implantation sites, adjacent lymph
> nodes and distant organs (liver, spleen, lings, muscle and urethra)
> were excised and histologically examined.
>
> The results of the human and mice implantation studies were similar
> with all implant sites apparent after 1 and 3 months and some
> undetectable after 6-9 months. In mice, no migration or transportation
> of any of injected particular filler substance to lymph nodes or
> filter organs could be detected. In these studies cross-linked
> collagen and hyaluronic acid were phagocytosized at 6 and 9 months,
> respectively. Microspheres of polylactic acid caused a mild
> inflammatory response and disappeared by 4 months. Dextran
> microspheres caused a pronounced foreign body response and disappeared
> at 8 months. Silicone particles caused the most pronounced foreign
> body reaction with a considerable amount of giant cell-persistence.
> Polyacrylamide injection was well tolerated, but disappeared
> clinically at 5 months. Trisacryl-gelatin microspheres caused little
> foreign body reaction, but were absorbed from the skin by 6 months.
> Polyvinylhydroxide microspheres were well tolerated and stable over 9
> moths like PMMA microspheres.

-   *Size Dependence of Spherical Tissue Fillers and* *Phagocytosis* --
    The phagocytosis of PLA and PMMA microspheres ranging in size from
    4.3 to 72 microns were determined by incubation with U-937
    macrophage, XS 106 and XS 52 Langerhans cells as well as HaCaT
    keratinocytes. The extent of phagocytosis was determined by light
    and confocal microscopy and also fluorescence activated cell
    sorting. The expression of TNFa after microparticle exposure was
    also determined.

> U-937 macrophages, keratinocytes and Langerhans cells phagocytosized
> PMMA microspheres smaller than 20 microns. Microspheres greater in
> size than 20 microns were not ingested by cells. Only collagen coated
> PLA microspheres stimulated TNFa secretion by U-937 cells.

[Other Biological Studies on the methylmethacrylate (MMA) monomer of
PMMA]{.underline}.

-   Pages 10-011 to 10-022 of Volume 10 summarize the published
    information on the toxicology and carcinogenicity of PMMA and its
    monomer (methylmethacrylate). This information includes summaries
    from "The World Health Organization International Agency for
    Research on Cancer" (i.e., a review of the chemical carcinogenicity
    of PMMA and MMA), "The National Toxicology Program" (i.e.,
    investigations on the dose-dependent carcinogenic effects of
    long-term daily systemic MMA exposure in rodents) and "Published
    reports on the safety of implant PMMA", (i.e., including cytotoxic,
    inflammatory and immunologic tissue responses to PMMA microspheres
    particles).

In conclusion, the biocompatibility tests and animal studies presented
show an acceptable response to PMMA. Tests on the biological response to
microspheres demonstrate that particles greater than 20 microns are not
phagocytised. The submitted data also demonstrate that MMA has been
removed by PMMA bead processing and the hence the risk of sensitization
is low.