FDA Review Team [1](#fda-review-team)

Team Leader Memo [2](#team-leader-memo)

Device Description [2](#device-description)

History [2](#history)

Report of Prior Investigations [3](#report-of-prior-investigations)

Clinical Review [13](#clinical-review)

Patient Population [14](#patient-population)

Treatment Parameters [16](#treatment-parameters)

Type of Material in Anastomoses [17](#type-of-material-in-anastomoses)

Safety Results [17](#safety-results)

Effectiveness Results [19](#effectiveness-results)

Limitations of the Study [20](#limitations-of-the-study)

Conclusion [21](#conclusion)

Immunology Review [21](#immunology-review)

Statistics Review [23](#statistics-review)

Chemistry Reviews [24](#chemistry-reviews)

# FDA Review Team

For P010003

CryoLife BioGlue®

## Team Leader Lisa Kennell

Clinical Reviewer Paul Chandeysson

Immunology Reviewer Katherine Merritt

Statistical Reviewer Hollington T. (T.C.) Lu

Chemist Reviewers Srilenka Das (BG3000) and

Ellen Chen (BG2000)

The memos that follow are from each of the reviewers on the review team.

# Team Leader Memo

### Device Description

The device is a two-part glue of purified bovine serum albumin (45% w/v
or 8 ml/10 ml) and glutaraldehyde (10% w/v or 2 ml/10 ml) in a
dispensing system that keeps the two parts separated until ready to use,
at which time they pass through a tortuous path applicator tip
(available in standard and extended lengths of 10, 20, and 27 cm) where
they are mixed in the appropriate proportions and dispensed onto the
tissue needing repair/bonding. The bonding is via a localized chemical
reaction resulting in covalent bonding (conjugated Schiff's bases or
amino bonds of the N-terminal amino acids and Є-amino groups of lysine)
of tissue surface proteins to the protein in the glue. The chemical
reaction begins immediately after the two compounds are mixed, setting
up in 20-30 seconds, and is completely reacted within 2 minutes. The
first 5 layers of the tissue are involved in the bond. It will bond the
tissue (closing the false lumen), reinforce tissue for facilitating
suturing of a graft, and seal the anastomoses. The compounds are in
separate plastic chambers in a dual-chamber cartridge, capped, double
pouched (foil pouch inside). A separate double pouch will contain the
twist ring tool (for tightening the tip) and a delivery device (glue
gun). All components are gamma irradiated (25 kGy dose with ^60^Co),
with the exception of the applicator tips with extended lengths, which
are sterilized by 100% ETO. Also available will be replacement tips and
refill cartridges. The glue has a 36-month shelf life date.

### History

This product has a regulatory history with the Agency, and has also
undergone some modifications in formulation, which are important to
understand in the review of the memos that follow. The sponsor first
submitted an Investigational Device Exemption (IDE) for a clinical study
for use of BioGlue® as an adjunct in repair of Type A (ascending) aortic
dissections. Once FDA approved the clinical study for that cohort, the
sponsor submitted a Humanitarian Device Exemption (HDE) for use of the
BioGlue® as an adjunct in repair of both Type A and Type B (descending)
aortic dissections. An HDE is a mechanism of granting limited marketing
approval for a restricted indication for treating a disease or patient
population that has an incidence rate of 4000 or less in the U.S. per
year. In granting such approvals, the Agency considers only the safety
and probable benefit of the device, but not the effectiveness. These
approvals are granted for devices when there are no alternatives, or the
alternatives are inadequate to treat this limited population. Since the
existing sealants and glues available (fibrin and thrombin based) are
slow-acting, FDA granted the approval in this case. However, should a
competitor gain marketing approval via the Premarket Approval (PMA)
process, in which both safety [and]{.underline} effectiveness are
established, then all existing products cleared via the HDE process
become null and void, since an alternative will then be available.

Once the HDE approval was granted for essentially the same patient
population as was being assessed in the IDE study, the sponsor had
difficulty enrolling patients into the IDE study, maintaining
randomization, and obtaining patient consent, and had protocol
violations with off-label use. FDA advised the sponsor that if they were
to seek a broader market for the BioGlue® that they would have to ensure
compliance with the protocol and regulations, or propose an alternative
study for the desired label. They then proposed the cardiac and vascular
study, for which the PMA is being considered today.

The formulation of the BioGlue® has undergone some modifications, which
should also be understood for your review. The sponsor began with the
BG1000 outside of the U.S. This was essentially the same as the BG2000,
but the user had to fill the syringes prior to using. The BG2000 was
supplied aseptically filled into the syringes for immediate use. The
BG3000 is gamma irradiated to achieve sterility, and also available for
immediate use. Both the BG2000 and the BG3000 are supplied in cartridges
that are loaded into a delivery system that has long, tortuous
applicators to pre-mix the two solutions in the correct ratios. The
BG2000 formulation gave toxic results in many of the biocompatibility
tests, but FDA decided to allow its use in the aortic dissection
indication, since the probable benefit of having a sealant that cures
immediately was believed to outweigh the risk of toxicity to this
patient population. However, when the sponsor decided to expand the
indication to sealing anastomoses, FDA's concern about the potential
toxicity was heightened. The gamma irradiated product was re-assessed
for the acute toxicity tests, and passed them all. We remain concerned
about the potential for an immune response, however.

### Report of Prior Investigations

Prior investigations of this material included an animal study,
biocompatibility testing, and bench studies on biodegradation, residual
glutaraldehyde, and visual, analytical, and functional testing,
described below.

##### Biocompatibility Testing

> The sponsor claims that ISO 10993-1/FDA Blue Book memo #G95-1 was
> followed for this testing, but some of the applicable tests were not
> performed, and no rationale was given for omission of the studies
> (specifically, no carcinogenicity or chronic toxicity testing was
> done, nor was biodegradation). Testing was done on the glue, and on
> the delivery system. The glue was allowed to cure prior to testing,
> which is probably not the preferred approach. FDA is developing a
> guidance for testing of cured products such as the BioGlue®, but it is
> not completed. The results of testing of the BG2000 formulation, the
> BG3000 formulation, and the delivery system components are tabulated
> on the next several pages.
>
> The BG2000 implantable glue gave a toxic response in many of the
> tests, specifically, in the cytotoxicity, genotoxicity, sub-chronic,
> hemolysis, and thrombogenicity tests. The components in the delivery
> system were not toxic, with the exception of the mixing chamber in the
> tip for cytotoxicity (this component was not assessed for 5 day
> implant). The sponsor does not provide a rationale to explain why
> these toxic results are acceptable, nor is there any rationale
> provided for not conducting the carcinogenicity, chronic, and
> biodegradation tests. Despite these omissions and the toxic results,
> as noted above, FDA granted permission to begin the aortic dissection
> study (1) because there is a history of use of similar products for
> the same indication in Europe, and (2) the formulations used in Europe
> contain similar amounts/doses of glutaraldehyde to that proposed in
> the BioGlue, and (3) the probable benefits (based on the European use
> and the animal study below) outweigh the disadvantages. We did not
> request further toxicological testing for this device for the
> dissection indication.
>
> The BG3000, assessed only for acute toxicity, passed all tests.

To justify omitting implant and chronic studies of the modified BG3000
formulation (gamma sterilized), testing was done in-house to compare the
extractables in the polymerized BG2000 series to the BG3000 series.
Testing involved pH, GC assay, UV-VIS scans, and HPLC analyses. The firm
concludes that there were no significant differences between the two
formulations. I referred this attachment to FDA chemists to verify that
the correct tests methods and conclusions were made. Dr. Chen indicated
that chemically, the BG2000 and BG3000 were not similar, but it was
agreed that since the physical and toxicological properties were not
altered, that we would allow this justification, and no further testing
was sought, with the exception of testing to assess the immunogenic
potential.

Table 1

Results of Toxicity Testing of the Implantable [BG2000]{.mark} Glue
Material

+--------+-------+--------------+-------+-----------------------------+
| **Test | **Ex  | **Extract    | *     | **Results/Comments** (units |
| P      | tract | conditions** | *Test | when appropriate)           |
| erform | (s)** | (time,       | and   |                             |
| ed/Lab | if    | temperature, | Co    |                             |
| Report | used  | area or mass | ntrol |                             |
| \#**   | (p    | to volume    | (s)** |                             |
| (lite  | olar, | ratio        | used  |                             |
| rature | non-p | compared to  |       |                             |
| cit    | olar) | in-use       |       |                             |
| ation, | or    | conditions)  |       |                             |
| st     | a     |              |       |                             |
| andard | nimal |              |       |                             |
| refere | model |              |       |                             |
| nce\*) |       |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| Cytoto | MEM   | 6 cm^2^/ml   | neat  | very cytotoxic reaction     |
| xicity |       | fresh cured  | and   | (Grade 4) after 48 hours    |
| (Chry  |       | at 37 °C for | 1:4   | with L-929 cells            |
| salis) |       | 24 hours     | dil   |                             |
|        |       |              | ution |                             |
|        |       |              | of    |                             |
|        |       |              | test  |                             |
|        |       |              | and   |                             |
|        |       |              | b     |                             |
|        |       |              | lank, |                             |
|        |       |              |       |                             |
|        |       |              | Cd    |                             |
|        |       |              | Cl~2~ |                             |
|        |       |              | 1000  |                             |
|        |       |              | µg/ml |                             |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | ve    |                             |
|        |       |              | hicle |                             |
|        |       |              | blank |                             |
|        |       |              | (-)   |                             |
+--------+-------+--------------+-------+-----------------------------+
| Cytoto | MEM   | 2.0 g air    | neat  | moderate reaction (Grade 3) |
| xicity |       | dried for 24 | test  | after 48 hours with L-929   |
| (To    |       | hours after  | ex    | cells                       |
| xicon) |       | curing in 10 | tract |                             |
|        |       | ml at 37 °C  |       |                             |
|        |       | for 24 hours | na    |                             |
|        |       |              | tural |                             |
|        |       |              | r     |                             |
|        |       |              | ubber |                             |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | sil   |                             |
|        |       |              | icone |                             |
|        |       |              | (-)   |                             |
+--------+-------+--------------+-------+-----------------------------+
| S      | s     | fresh cured  | DNCB  | 0.3 ml dermal doses, no     |
| ensiti | aline | 3 cm^2^/ml   | (+)   | sensitization observed      |
| zation |       | ratio at     |       |                             |
| (Chr   |       | 50°C for 72  | s     |                             |
| ysalis |       | hours        | aline |                             |
| B      |       |              | blank |                             |
| uehler |       |              | (-)   |                             |
| met    |       |              |       |                             |
| hod\*) |       |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| I      | sa    | ratio of 3   | ve    | No difference in the        |
| rritat | line, | cm^2^/ml at  | hicle | control and test extracts   |
| ion/In | 1:20  | 50 ° for 72  | b     | were observed at 24, 48, or |
| tracut | Et    | hours        | lanks | 72 hours for any vehicle.   |
| aneous | OH/sa |              | (-)   |                             |
| To     | line, |              |       |                             |
| xicity | PEG,  |              |       |                             |
| (Chry  | and   |              |       |                             |
| salis) | s     |              |       |                             |
|        | esame |              |       |                             |
|        | oil   |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| Sy     | sa    | ratio of 3   | ve    | No difference in the        |
| stemic | line, | cm^2^/ml at  | hicle | control and test extracts   |
| to     | 1:20  | 50 ° for 72  | b     | were observed at 24, 48, or |
| xicity | Et    | hours        | lanks | 72 hours for any vehicle.   |
|        | OH/sa |              | (-)   |                             |
|        | line, |              |       |                             |
|        | PEG,  |              |       |                             |
|        | and   |              |       |                             |
|        | s     |              |       |                             |
|        | esame |              |       |                             |
|        | oil   |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| Sub-c  | 90    | \~1.8 ml     | S     | Implant area remained       |
| hronic | day   |              | aline | raised and firm for most    |
| to     | su    |              |       | animals; some had erythema  |
| xicity | bcuta |              |       | and abrasions at the site.  |
|        | neous |              |       | Changes were noted in the   |
|        | im    |              |       | RBC and WBC values. No      |
|        | plant |              |       | effect on body weight or    |
|        | in    |              |       | weight gain, hematology,    |
|        | rat   |              |       | coagulation, clinical       |
|        | model |              |       | chemistry other than noted. |
|        | (n=20 |              |       | No gross pathology or organ |
|        | each  |              |       | weight differences. The     |
|        | test  |              |       | implant was encapsulated    |
|        | and   |              |       | with some mineralization    |
|        | con   |              |       | and evidence of a foreign   |
|        | trol) |              |       | body reaction.              |
+--------+-------+--------------+-------+-----------------------------+
| Genoto |       |              |       |                             |
| xicity |       |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| gene   | s     | 4 g/20 ml    | s     | no genotoxicity in TA 98,   |
| mu     | aline | ratio at 37  | aline | TA100, TA1535, and TA1537   |
| tation |       | °C for 24    | (-)   | strains of *S. typhimurium* |
| Ames   |       | hours        |       | with and without metabolic  |
|        |       |              | 2-    | activation                  |
|        |       |              | amino |                             |
|        |       |              | anthr |                             |
|        |       |              | acene |                             |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | S     |                             |
|        |       |              | odium |                             |
|        |       |              | Azide |                             |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | 2-Nit |                             |
|        |       |              | roflu |                             |
|        |       |              | orene |                             |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | 9-Ami |                             |
|        |       |              | noacr |                             |
|        |       |              | idine |                             |
|        |       |              | (+)   |                             |
+--------+-------+--------------+-------+-----------------------------+

+--------+-------+--------------+-------+-----------------------------+
| **Test | **Ex  | **Extract    | *     | **Results/Comments** (units |
| P      | tract | conditions** | *Test | when appropriate)           |
| erform | (s)** | (time,       | and   |                             |
| ed/Lab | if    | temperature, | Co    |                             |
| Report | used  | area or mass | ntrol |                             |
| \#**   | (p    | to volume    | (s)** |                             |
| (lite  | olar, | ratio        | used  |                             |
| rature | non-p | compared to  |       |                             |
| cit    | olar) | in-use       |       |                             |
| ation, | or    | conditions)  |       |                             |
| st     | a     |              |       |                             |
| andard | nimal |              |       |                             |
| refere | model |              |       |                             |
| nce\*) |       |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| gene   | s     | 6 cm^2^/ml   | s     | TA1535, TA1537, TA98, TA100 |
| mu     | aline | ratio at 50  | aline | and TA102 strains of *S.    |
| tation | and   | °C for 72    | (-)   | typhimurium* and WP2 *uvrA* |
| Ames   | DMSO  | hours in     |       | strain of *E. coli* with    |
|        |       | doses of     | 2-    | and without metabolic       |
|        |       | 1.67, 5.00,  | anthr | activation were assessed.   |
|        |       | 16.7, 50.0,  | amine | Possible genotoxic result   |
|        |       | 100, and 200 | (+)   | was found in strain TA1535  |
|        |       | µl/plate     |       | with activation in saline,  |
|        |       |              | S     | and for TA1535, TA 1537 and |
|        |       |              | odium | WP2 *uvrA* in DMSO.         |
|        |       |              | Azide | Retesting gave normal       |
|        |       |              | (+)   | results except for TA1537   |
|        |       |              |       | with S9, and thus           |
|        |       |              | 2-Nit | considered to be marginally |
|        |       |              | roflu | positive for strains TA1535 |
|        |       |              | orene | and TA1537.                 |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | 9-Ami |                             |
|        |       |              | noacr |                             |
|        |       |              | idine |                             |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | Mito  |                             |
|        |       |              | mycin |                             |
|        |       |              | C (+) |                             |
|        |       |              |       |                             |
|        |       |              | ENNG  |                             |
|        |       |              | (+)   |                             |
|        |       |              |       |                             |
|        |       |              | 2-ami |                             |
|        |       |              | noflu |                             |
|        |       |              | orene |                             |
|        |       |              | (+)   |                             |
+--------+-------+--------------+-------+-----------------------------+
| AS52   | Ham's | 120 cm^2^/40 | sham  | Genotoxic results           |
| C      | F12   | ml (3        | F12   | (dose-dependent increase in |
| hinese | trea  | cm^2^/ml)    | ve    | mutants) observed           |
| H      | tment | ratio for 24 | hicle |                             |
| amster | m     | hours at 37  | (-)   |                             |
| Ovary  | edium | ° C          |       |                             |
| Cel    |       |              | ethyl |                             |
| l/XPRT |       | Ten final    | m     |                             |
| Mam    |       | co           | ethan |                             |
| malian |       | ncentrations | esulf |                             |
| Cell   |       | ranging from | onate |                             |
| F      |       | 0.00333 to   | (+)   |                             |
| orward |       | neat for     |       |                             |
| Mu     |       | toxicity     | dime  |                             |
| tation |       | screen, then | thyln |                             |
| Assay  |       | between      | itros |                             |
|        |       | 0.0000500    | amine |                             |
|        |       | and neat     | (+)   |                             |
|        |       | with S9 and  |       |                             |
|        |       | 0.500 to     |       |                             |
|        |       | 66.7 with S9 |       |                             |
|        |       | for the      |       |                             |
|        |       | initial      |       |                             |
|        |       | assay, then  |       |                             |
|        |       | 0.500-83.3%  |       |                             |
|        |       | with and     |       |                             |
|        |       | without S9   |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| Chro   | RPMI  | 6 cm^2^/ml   | c     | Tox pre-screen revealed     |
| mosome | 1640  | at 37 ° C    | yclop | toxic effects, but not due  |
| aber   |       | for 24 hours | hosph | to pH or osmolality.        |
| ration |       |              | amide |                             |
| in     |       | Tox          | (+)   | Genotoxic effect (dose      |
| Human  |       | pre-screen   |       | dependent increase in       |
| Lymph  |       | in ten       | miti  | porportion of aberrant      |
| ocytes |       | co           | mycin | metaphase and               |
|        |       | ncentrations | C     | aberrations/cell) were      |
|        |       | with and     |       | noted in initial assay.     |
|        |       | without S9   |       |                             |
|        |       |              |       | Results of confirmatory     |
|        |       | Initial      |       | assay same as initial       |
|        |       | assay using  |       |                             |
|        |       | 5.00-75%     |       |                             |
|        |       | co           |       |                             |
|        |       | ncentration. |       |                             |
|        |       |              |       |                             |
|        |       | Confirmatory |       |                             |
|        |       | assay        |       |                             |
|        |       | involved     |       |                             |
|        |       | co           |       |                             |
|        |       | ncentrations |       |                             |
|        |       | of 5.00-50%  |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| DNA    |       |              |       |                             |
| damage |       |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| *in    |       |              |       |                             |
| vivo*  |       |              |       |                             |
| assay  |       |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| Implan | N/A   | N/A          | USP   | At 120 hours (5 days) the   |
| tation |       |              | po    | BioGlue gave results        |
|        |       |              | lyeth | similar to the negative     |
|        |       |              | ylene | controls (macroscopic       |
|        |       |              |       | assessment only)            |
|        |       |              | (-)   |                             |
+--------+-------+--------------+-------+-----------------------------+
| Hemoc  |       |              |       |                             |
| ompati |       |              |       |                             |
| bility |       |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+
| hem    | human | 1 cm x 1 mm  | USP   | No hemolysis of human       |
| olysis | and   | pieces in 5  | po    | blood, but monkey blood     |
|        | m     | ml           | lyeth | inconclusive, since         |
|        | onkey |              | ylene | negative control caused     |
|        | blood |              |       | hemolysis.                  |
|        |       |              | (-)   |                             |
+--------+-------+--------------+-------+-----------------------------+
| Comp   | rat   | subcutaneous | s     | 5 µl samples were analyzed  |
| lement | serum | implant of   | aline | for hemolytic complement    |
| acti   | sa    | \~1.8 ml     | inje  | with the EZ Complement CH50 |
| vation | mples |              | ction | assay kit on day 30 and day |
|        | from  |              |       | 90 after implant. No        |
|        | subch |              |       | difference was observed     |
|        | ronic |              |       | between test and control    |
|        | study |              |       | animals on day 30 or day 90 |
|        | an    |              |       |                             |
|        | imals |              |       |                             |
|        | below |              |       |                             |
+--------+-------+--------------+-------+-----------------------------+

  ----------------- ---------------- -------------- -------------- -----------------------------
  **Test            **Extract(s)**   **Extract      **Test and     **Results/Comments** (units
  Performed/Lab     if used (polar,  conditions**   Control(s)**   when appropriate)
  Report \#**       non-polar) or    (time,         used           
  (literature       animal model     temperature,                  
  citation,                          area or mass                  
  standard                           to volume                     
  reference\*)                       ratio compared                
                                     to in-use                     
                                     conditions)                   

  thrombogenicity   rabbit carotid   1.0-1.8 ml     N/A            aPTT, platelets, HCT, ACT,
                    artery (n=5)     injected into                 and bleeding time were
                                     carotid artery                measured before and after
                                                                   injection (at 15, 60 and 120
                                                                   minutes), as were systemic
                                                                   arterial pressure, heart
                                                                   rate, and lead II ECG (after
                                                                   30 minutes of stabilization,
                                                                   for 10 minutes). No effect on
                                                                   any parameter except for
                                                                   blood flow, which was 85%
                                                                   reduced in 2, and temporarily
                                                                   increased in 2.
                                                                   Microscopically the
                                                                   endothelial cells were
                                                                   affected (cells lost with
                                                                   thrombi formed on the wall,
                                                                   i.e., this material caused
                                                                   thrombus formation

  Pyrogenicity                                                     

  Chronic toxicity                                                 

  Carcinogenicity                                                  

  Biodegradation                                                   
  ----------------- ---------------- -------------- -------------- -----------------------------

Table 2

> Results of Toxicity Testing of the Delivery System

+--------+-------+-------------+--------+-----------------------------+
| **Test | **Ex  | **Extract   | **Test | **Results/Comments** (units |
| P      | tract | c           | and    | when appropriate)           |
| erform | (s)** | onditions** | Contro |                             |
| ed/Lab | if    | (time,      | l(s)** |                             |
| Report | used  | t           | used   |                             |
| \#**   | (p    | emperature, |        |                             |
| (lite  | olar, | area or     |        |                             |
| rature | non-p | mass to     |        |                             |
| cit    | olar) | volume      |        |                             |
| ation, | or    | ratio       |        |                             |
| st     | a     | compared to |        |                             |
| andard | nimal | in-use      |        |                             |
| refere | model | conditions) |        |                             |
| nce\*) |       |             |        |                             |
+--------+-------+-------------+--------+-----------------------------+
| Cytoto | MEM   | 6 cm^2^/ml  | v      | eight components tested     |
| xicity |       | ratio at 37 | ehicle | separately. All components  |
|        |       | ° C for 24  | (-)    | gave a mild or less         |
|        |       | hours       |        | reaction (non-toxic) after  |
|        |       |             | 2.00   | 48 hours of exposure,       |
|        |       |             | µl/ml  | except for the mixing tip,  |
|        |       |             | C      | which gave a moderate score |
|        |       |             | dCl~2~ | of 3.0 (toxic).             |
|        |       |             | (+)    |                             |
+--------+-------+-------------+--------+-----------------------------+
| S      | s     | "           | saline | None of the test or         |
| ensiti | aline | appropriate | (-)    | negative control animals    |
| zation |       | surfac      |        | had a sensitization         |
| (B     |       | e-to-volume | DNCB   | reaction; most of the       |
| uehler |       | ratios" at  | in     | positive control animals    |
| met    |       | 50 ° C for  | EtOH   | had a reaction.             |
| hod\*) |       | 72 hours    | (indu  |                             |
|        |       |             | ction) |                             |
|        |       |             | or     |                             |
|        |       |             | a      |                             |
|        |       |             | cetone |                             |
|        |       |             | (chal  |                             |
|        |       |             | lenge) |                             |
+--------+-------+-------------+--------+-----------------------------+

+--------+---+-------+--------------+--------+-----------------------------+
| **Test |   | **Ex  | **Extract    | **Test | **Results/Comments** (units |
| P      |   | tract | conditions** | and    | when appropriate)           |
| erform |   | (s)** | (time,       | Contro |                             |
| ed/Lab |   | if    | temperature, | l(s)** |                             |
| Report |   | used  | area or mass | used   |                             |
| \#**   |   | (p    | to volume    |        |                             |
| (lite  |   | olar, | ratio        |        |                             |
| rature |   | non-p | compared to  |        |                             |
| cit    |   | olar) | in-use       |        |                             |
| ation, |   | or    | conditions)  |        |                             |
| st     |   | a     |              |        |                             |
| andard |   | nimal |              |        |                             |
| refere |   | model |              |        |                             |
| nce\*) |   |       |              |        |                             |
+--------+---+-------+--------------+--------+-----------------------------+
| I      | s |       | all          | v      | No difference between       |
| rritat | a |       | components   | ehicle | results in the test and     |
| ion/In | l |       | tested       | co     | control animals at 24, 48   |
| tracut | i |       | separately   | ntrols | or 72 for any component,    |
| aneous | n |       | at 50 ° C    |        | except for the O-ring in    |
| To     | e |       | for 72 hours | (-);   | sesame oil (0.3 control vs. |
| xicity | , |       | extraction   |        | 1.3 test, repeat 0.5        |
|        | 1 |       | using the    | PEG    | control, 1.7 test)          |
|        | : |       | following    | blank  |                             |
|        | 2 |       | ratios:      | and    |                             |
|        | 0 |       |              | test   |                             |
|        | E |       | 1            | d      |                             |
|        | t |       | car          | iluted |                             |
|        | O |       | tridge/41.06 | 7:4    |                             |
|        | H |       | ml           |        |                             |
|        | : |       |              |        |                             |
|        | s |       | 10 pieces (5 |        |                             |
|        | a |       | each piston  |        |                             |
|        | l |       | A and        |        |                             |
|        | i |       | B)/23.5 ml   |        |                             |
|        | n |       |              |        |                             |
|        | e |       | 40 pieces/20 |        |                             |
|        | , |       | ml O-ring    |        |                             |
|        | P |       | (EPDM and ±  |        |                             |
|        | E |       | lubricant)   |        |                             |
|        | G |       |              |        |                             |
|        | 4 |       | 5            |        |                             |
|        | 0 |       | pieces/23.6  |        |                             |
|        | 0 |       | ml mixing    |        |                             |
|        | , |       | tip housing  |        |                             |
|        | s |       |              |        |                             |
|        | e |       | 24           |        |                             |
|        | s |       | pieces/20.2  |        |                             |
|        | a |       | ml mixing    |        |                             |
|        | m |       | tip mixing   |        |                             |
|        | e |       | chamber      |        |                             |
|        | o |       |              |        |                             |
|        | i |       | 1            |        |                             |
|        | l |       | piece/36.65  |        |                             |
|        |   |       | ml plunger   |        |                             |
+--------+---+-------+--------------+--------+-----------------------------+
| Sy     | s |       | same as      | same   | no difference between       |
| stemic | a |       | in           | as     | control and test animals    |
| to     | l |       | tracutaneous | above  | for any component           |
| xicity | i |       | test above   |        |                             |
|        | n |       |              |        |                             |
|        | e |       |              |        |                             |
|        | , |       |              |        |                             |
|        | 1 |       |              |        |                             |
|        | : |       |              |        |                             |
|        | 2 |       |              |        |                             |
|        | 0 |       |              |        |                             |
|        | s |       |              |        |                             |
|        | a |       |              |        |                             |
|        | l |       |              |        |                             |
|        | i |       |              |        |                             |
|        | n |       |              |        |                             |
|        | e |       |              |        |                             |
|        | : |       |              |        |                             |
|        | E |       |              |        |                             |
|        | t |       |              |        |                             |
|        | O |       |              |        |                             |
|        | H |       |              |        |                             |
|        | , |       |              |        |                             |
|        | P |       |              |        |                             |
|        | E |       |              |        |                             |
|        | G |       |              |        |                             |
|        | , |       |              |        |                             |
|        | s |       |              |        |                             |
|        | e |       |              |        |                             |
|        | s |       |              |        |                             |
|        | a |       |              |        |                             |
|        | m |       |              |        |                             |
|        | e |       |              |        |                             |
|        | o |       |              |        |                             |
|        | i |       |              |        |                             |
|        | l |       |              |        |                             |
+--------+---+-------+--------------+--------+-----------------------------+
| Sub-c  | N |       | 1 x 10 mm    | USP    | After 120 hours (5 days)    |
| hronic | / |       | strips of    | polyet | implant, sites were         |
| to     | A |       | the          | hylene | examined macroscopically    |
| xicity |   |       | cartridge,   | high   | only for capsule,           |
|        |   |       | pistons A    | d      | hemorrhage, infection, and  |
|        |   |       | and B,       | ensity | necrosis. There was no      |
|        |   |       | O-ring (EPDM | p      | encapsulation of any        |
|        |   |       | and ±        | lastic | component, and healing was  |
|        |   |       | lubr         |        | similar to the negative     |
|        |   |       | icant)mixing |        | control in all cases. No    |
|        |   |       | tip housing, |        | results were provided for   |
|        |   |       | , and        |        | the mixing tip mixing       |
|        |   |       | plunger      |        | chamber                     |
+--------+---+-------+--------------+--------+-----------------------------+

Table 3

Results of Toxicity Testing of the [BG3000]{.mark} BioGlue

+--------+-------+--------------+--------+-----------------------------+
| **Test | **Ex  | **Extract    | **Test | **Results/Comments** (units |
| P      | tract | conditions** | and    | when appropriate)           |
| erform | (s)** | (time,       | Contro |                             |
| ed/Lab | if    | temperature, | l(s)** |                             |
| Report | used  | area or mass | used   |                             |
| \#**   | (p    | to volume    |        |                             |
| (lite  | olar, | ratio        |        |                             |
| rature | non-p | compared to  |        |                             |
| cit    | olar) | in-use       |        |                             |
| ation, |       | conditions)  |        |                             |
| st     |       |              |        |                             |
| andard |       |              |        |                             |
| refere |       |              |        |                             |
| nce\*) |       |              |        |                             |
+--------+-------+--------------+--------+-----------------------------+
| Cytoto | MEM   | 9.6g in 48.0 | N      | L-929 cells gave a grade 2  |
| xicity |       | ml at 37 °C  | atural | (mild) reactivity score     |
| (ISO)  |       | for 24 hours | rubber | with test extract at 48     |
|        |       |              | (+)    | hours                       |
|        |       |              |        |                             |
|        |       |              | Si     |                             |
|        |       |              | licone |                             |
|        |       |              | tubing |                             |
|        |       |              | (-)    |                             |
+--------+-------+--------------+--------+-----------------------------+

+--------+-------+--------------+--------+-----------------------------+
| **Test | **Ex  | **Extract    | **Test | **Results/Comments** (units |
| P      | tract | conditions** | and    | when appropriate)           |
| erform | (s)** | (time,       | Contro |                             |
| ed/Lab | if    | temperature, | l(s)** |                             |
| Report | used  | area or mass | used   |                             |
| \#**   | (p    | to volume    |        |                             |
| (lite  | olar, | ratio        |        |                             |
| rature | non-p | compared to  |        |                             |
| cit    | olar) | in-use       |        |                             |
| ation, |       | conditions)  |        |                             |
| st     |       |              |        |                             |
| andard |       |              |        |                             |
| refere |       |              |        |                             |
| nce\*) |       |              |        |                             |
+--------+-------+--------------+--------+-----------------------------+
| S      | S     | 4g/20ml 37   | Saline | No sensitization was        |
| ensiti | aline | °C 72 hours  | and    | observed                    |
| zation | and   |              | CSO    |                             |
| (M     | CSO   |              | (-)    |                             |
| aximiz |       |              |        |                             |
| ation) |       |              | DNCB   |                             |
|        |       |              | (+)    |                             |
|        |       |              | with   |                             |
|        |       |              | and    |                             |
|        |       |              | w      |                             |
|        |       |              | ithout |                             |
|        |       |              | acti   |                             |
|        |       |              | vation |                             |
+--------+-------+--------------+--------+-----------------------------+
| In     | S     | 4g/20ml at   | Saline | No toxicity observed in     |
| tracut | aline | 37 °C for 72 | and    | either extract              |
| aneous | and   | hours        | CSO    |                             |
| To     | CSO   |              |        |                             |
| xicity |       |              |        |                             |
| (IS    |       |              |        |                             |
| O/USP) |       |              |        |                             |
+--------+-------+--------------+--------+-----------------------------+
| Sy     | S     | 4g/20ml at   | Saline | No signs of toxicity        |
| stemic | aline | 37 °C for 72 | and    |                             |
| To     | (IV)  | hours        | CSO    |                             |
| xicity | and   |              |        |                             |
| (IS    | CSO   |              |        |                             |
| O/USP) | (IP)  |              |        |                             |
+--------+-------+--------------+--------+-----------------------------+
| Hem    | S     | 4g/20ml at   | Saline | 4.45% hemolysis, considered |
| olysis | aline | 37 °C for 72 | (-)    | to be non-hemolytic         |
| (DHEW) |       | hours        |        |                             |
|        |       |              | Water  |                             |
|        |       |              | (+)    |                             |
+--------+-------+--------------+--------+-----------------------------+
| Rabbit | s     | 4g/20ml at   | Saline | \<0.5 °C rise in all        |
| p      | aline | 37 °C for 72 | (-)    | rabbits-non pyrogenic       |
| yrogen |       | hours        |        |                             |
| (ISO)  |       |              |        |                             |
+--------+-------+--------------+--------+-----------------------------+

##### Animal Study

> A previously reported sheep model for acute descending aortic
> dissection was used for this study, involving 30 sheep (30 enrolled to
> guarantee a minimum of 12 animals in both the test and control
> groups). Dissections were created and verified to meet criteria (false
> lumen volume \<50% of total aortic volume, length of \<7.1 cm, and
> width of \<2.6 cm). Animals were then randomized in a blinded fashion
> into either the surgical repair of the proximal flap alone, or repair
> by gluing the layers of the dissection together and surgical repair of
> the proximal flap. A total of 13 animals were randomized into each
> group (4 dissections did not meet criteria). A 40% mortality rate was
> anticipated either due to failure of the model to perform or failure
> of the treatment. Failure of the model to perform was defined as
> progression of the dissection past 7 cm, 50% of total aortic volume,
> spontaneous rupture, or other reason for instability of the
> dissection. At explant, the aortas were excised and gross and
> microscopic histopathology assessments made. Dissections varied from
> 5.0 to 7.0 cm long (mean 5.2 cm), 1.2-2.5 cm wide (mean 1.4 cm), and
> 20-50% (mean 35%) of the aortic volume (statistically similar among
> the two groups).
>
> In the [test]{.underline} group, 5 died (3 pneumonia/weather stress, 1
> tension pneumothorax, and 1 chronic aneurysm rupture at 51 days), all
> with healed or obliterated dissections except for the animal with the
> rupture (due to mycotic super-infection of a technical glue failure
> \[this event deemed related\]) In the 8 animals surviving to the 90
> day sacrifice period, the dissection was completely healed without
> signs of progression.
>
> In the [control/surgical]{.underline} group, 1 animal died of
> pneumonia/weather stress, and 4 of proximal or distal aneurysm
> progression and rupture \[all deemed related\]. Eight animals survived
> to 90 days. Of these, 2 dissections healed after 3-3.5 cm of
> progression; 2 had a chronic dissection formed at the distal suture
> line; in 4 a chronic dissection formed at re-entry points unrelated to
> the surgical repair site. The false lumen made up 25-75% of the total
> aortic lumen in these chronic cases, and the aortic wall was thinning
> and the diameter expanding.
>
> Histologically, various parameters were given a numeric score for
> comparison purposes (intimal hyperplasia, fibrin deposition, thrombus,
> acute inflammation, fibrosis, neovascularization, granulomatous
> inflammation, giant cells, lymphoid nodules, tissue necrosis,
> hemorrhage, and mineralization). Other parameters which were scored
> for statistical analysis included cross-clamp duration,
> intra-operative blood loss, complications, mortality, treatment
> failure, animal gender, age, weight, and hematocrit, dissection
> length, width, area. The early related mortality rate was
> significantly different using 1 sided Fisher's Exact test p values,
> but not for 2 sided (sample size too small), suggesting a trend toward
> lower mortality with BioGlue than surgical repair. All mortality
> combined was not statistically different between groups. Late death
> was not significantly different (related or overall). When treatment
> related mortality and morbidity were considered together, the
> difference between the groups was significant, with 12/13 animals in
> the control group experiencing either a related death or morbidity,
> compared to only one related death in the test group, whether or not
> animals not completing the study were considered in the analysis.
> Histological analysis revealed differences in the severity and
> frequency of lymphoid nodule formation; there was an inflammatory
> reaction associated with the BioGlue; mineralization and tissue
> necrosis was more frequent and severe in the BioGlue samples. In areas
> where the BioGlue was applied, there was fibrosis in the media, and
> thinner walls were observed. The conclusion by the pathologist was
> that these observations were consistent with a foreign body reaction,
> and that the wall thickness in the BioGlue group is consistent with
> the variation in the surgical procedure for creation of the
> dissection. Intimal hyperplasia was markedly reduced in the lesions
> treated with BioGlue.
>
> Additional animal studies were done to assess sealing at the
> anastomosis area (coagulopathic sheep model and sutureless porcine
> CABG model) in support of modifying the protocol for the cardiac and
> vascular study.

##### *In vitro* Testing of the BioGlue

> Tests were done on the components (albumin and glutaraldehyde), on the
> adhesive, and on the packaging, described below. Sample sizes were
> mostly chosen using the MIL Standard 105E.

1.  Tests on albumin component

-   physical inspection---is done throughout the manufacturing process.
    The entire lot is inspected after the BSA is filled into the vial
    for damage and volume, and portions of the lot are checked for
    integrity of the crimp and seal.

-   UV-Vis spectrophotometric profile---Each batch is checked in the
    range of 200-700 nm for its spectrophotometric profile at 10 mg/ml
    dilution. In order to be acceptable, peaks only at 212 (peptide
    bonds) and 280 (aromatic amino acid side chains) are acceptable.
    Additional peaks would mean impure albumin.

-   PH \[information redacted\]. The pH is monitored throughout the
    process (raw material, filtered bulk and heat treated vial).

-   Purity (SDS-PAGE---sodium dodecyl sulfate-polyacrylamide gel
    electrophoresis)---this method is used to determine the amount of
    polymerization/non-monomer form of the albumin after filtration and
    heat treatment \[information redacted\]. Bands forming in locations
    other than 66,000 kDa are indicative of impurities or polymers.
    \[information redacted\].

-   Protein concentration (Colorimetric)---this method verifies the
    concentration of the protein to ensure adequate mixing and
    dispensing (too high a concentration will result in too viscous a
    mixture which will not flow through the mixing tip; too low a
    concentration results in poor bonding strength). This method was
    done throughout the process at dilutions of 0.45 mg/ml. The
    specification is 45 ± 3% protein.

-   Bioburden---this is assessed by filtration of the 45% bulk material.
    \[Information redacted\] aerobic, anaerobic, mold and yeast. There
    is no mention of fungi.

-   Sterility of filled vials---the USP sampling procedure (10% of the
    lot, maximum 20) is used to verify sterility of the filled vials, by
    either filtration or direct inoculation.

2.  Tests on Glutaraldehyde component:

-   physical inspection---inspection similar to the albumin component is
    done throughout the process, both on the 50% and the 10% solutions
    in the filled vials.

-   glutaraldehyde concentration (hydroxylamine assay)---the adhesive
    properties needed are dependent on the appropriate amount of
    glutaraldehyde, \[information redacted\] so the specification chosen
    was 10%. This titration assay is used to verify the concentration of
    the raw stock solution \[information redacted\] and that in the
    vials and diluted stock solution \[information redacted\].

-   extent of autopolymerization (absorbance ratio)---The monomer has an
    absorbance peak at 280 nm, whereas polymerized material peaks at 235
    nm, so the ratio of the peak at 280 to 235 nm is measured to give an
    indication of the percentage of polymerization. \[Information
    redacted\].

-   Sterility of filled vials---This is similar to the testing for the
    albumin component; no aerobic or anaerobic growth after 14 days

3.  Functional testing of Adhesive

-   Shear strength---initial strength tests were done during muscle
    samples, but these samples were variable in their stretch, fiber
    orientation, and fat content, which led to variable results. A
    method using porcine dermal tissue was validated to give more
    reliable results, \[information redacted\]. Samples are tested for
    shear strength after applying glue to a 1 cm^2^ \[information
    redacted\].

4.  Packaging

-   Inspection (subassemblies and final)---packages are inspected for
    damage, component verification and seal closure. Sample size varies
    with lot. Also checked is the labeling, radiation dose (delivery
    system).

-   Applicator tip pouch sub-assembly---as above

-   BioGlue delivery tray package and label---as above

-   delivery tray bagged for irradiation---as above

-   inspection of irradiated tray---the count is verified to be the same
    as was sent out for sterilization

-   Adhesive package and label---as above, shelf life date also checked.

-   Seal Peel test---both the pouch and the tray are peel tested to
    verify that a minimum of 1.1 PSI is achieved (n=13/lot).

# Clinical Review

The sponsor has reported the results of a prospective, multicentered,
randomized, blinded clinical trial of BioGlue Surgical Adhesive in
patients undergoing cardiac or vascular surgical procedures. The purpose
of the study was to collect data to support the safety and effectiveness
of BioGlue when used as an anastomotic sealant to provide hemostasis. A
total of 151 patients who were to undergo a cardiac or vascular surgical
procedure were randomized in a 1:1 ratio to surgical treatment with or
without BioGlue. The patients were blinded as to which treatment they
had received until after the study was complete. Follow-up was at
discharge from the hospital, 1 month and 3 months. Enrollment of
patients began April 25, 2000 and was completed on September 15, 2000. A
total of 6 investigational sites participated.

The primary response variable was the proportion of vascular anastomoses
achieving hemostasis without the need for additional measures to control
bleeding. This was assessed just before skin closure after blood
pressure had been restored to the vessel. The study hypothesis was that
the proportion of anastomoses achieving hemostasis would be higher in
the patients treated with BioGlue. The required sample size was
estimated based on the assumption that the rate of continued bleeding at
the anastomotic site would be 5% in the patients treated with BioGlue
and 15% in the patients treated without BioGlue. Using a two-tailed test
and an alpha of 0.05, a sample size of 141 anastomoses per treatment
group was required to provide a statistical power of 80%. Assuming that
there would be 2 anastomoses per patient, this required 71 patients in
each treatment group. This number was increased by 10% to account for
patients who crossed over from one group to the other because of
uncontrolled bleeding. The number was increased by an additional 10% to
allow for patients lost to follow-up. The resulting estimated sample
size was 86 patients per treatment group.

The number of patients who crossed over was less than expected. Only one
patient, who had been randomized to treatment without BioGlue, had
uncontrolled bleeding. That patient was crossed over to the BioGlue
treatment arm. Enrollment in the study was stopped when 151 patients had
been treated and evaluated for the primary response variable. At that
time, 76 patients had been randomized to the test group treated with
BioGlue and 75 patients had been randomized to surgery without BioGlue.
These numbers exceeded the estimated required sample size of 71
patients.

The randomization of patients was controlled so that each clinical
center enrolled approximately the same number of patients into each
treatment group. The number of patients enrolled per clinical center
differed considerably, with one center enrolling 32% of the patients and
one center enrolling only 2% of the patients. Three of the six clinical
centers enrolled more than the expected average of 17% of the total;
therefore, the multicentered nature of the study was fairly well
maintained.

### Patient Population

Baseline demographic and clinical data are presented, allowing a
comparison of the treatment groups. The following table summarizes these
data. CAD, COPD, CRF, and CHF mean coronary artery disease, chronic
obstructive pulmonary disease, chronic renal failure, and congestive
heart failure. The numbers in parentheses are the percent of the total
number of patients in the treatment group.

###### Comparison of Treatment Groups

  --------------------------------- ------------ ------------ ------------
  Patient Parameter                 BioGlue      Control      Crossover
                                    Group (n=76) Group (n=74) (n=1)

  Mean Age (years)                  63.4         66.3         73

  Men                               49 (64%)     48 (65%)     0

  Women                             27 (36%)     26 (35%)     1

  Caucasian                         67 (88%)     66 (89%)     1

  Smoker                            60 (79%)     57 (77%)     1

  Diabetic                          12 (16%)     11 (15%)     0

  Herpertensive                     55 (73%)     57 (77%)     1

  Aortic aneurysm                   56 (74%)     49 (66%)     1

  CAD                               34 (45%)     30 (41%)     1

  Patient Parameter                 BioGlue      Control      Crossover
                                    Group (n=76) Group (n=74) (n=1)

  Visceral Occlusive Disease        4 (5%)       3 (4%)       0

  CHF                               16 (21%      11 (15%)     0

  CRF                               6 (8%)       7 (9%)       0

  Cerebrovascular Disease           7 (9%)       12 (16%)     0

  Peripheral Vascular Disease       15 (20%)     19 (26%)     0

  Stroke                            8 (11%)      12 (16%)     0
  --------------------------------- ------------ ------------ ------------

The patients in the treatment groups appear fairly well matched in terms
of these factors.

The preoperative indications for which the patients underwent surgery
are summarized in the following table. The numbers in parentheses are
the percent of the total number of patients in the treatment group.

###### Indications for Surgery

  ---------------------------------- ------------ ------------ ------------
  Parameter                          BioGlue®     Control      Crossover
                                     Group (n=76) Group (n=74) (n=1)

  Thoracoabdominal aneurism          16 (21%)     19 (26%)     0

  Ascending aortic aneurism          23 (30%)     19 (26%)     1

  Other aortic aneurism              16 (21%)     13 (18%)     0

  Peripheral occlusive disease       4 (%5)       7 (9%)       0

  Carotid occlusive disease          10 (13%)     9 (12%)      0

  Aortic root surgery                4 (5%)       3 (4%)       0

  Aortic valve disease               7 (9%)       14 (19%)     0

  Other surgery                      7 (9%)       7 (9%)       0

  Total                              87           91           1
  ---------------------------------- ------------ ------------ ------------

Since the total number of indications for surgery is greater than the
number of patients, some of the patients had more than one indication
for their surgery. Surgery for aortic aneurisms predominated in this
series of patients. The indications for surgery were fairly well
balanced between the treatment groups.

The preoperative laboratory values for the patients who underwent
surgery are summarized in the following table.

###### Preoperative Laboratory Values

  ---------------------------- -------------- -------------- -------------
  Parameter                    BioGlue® Group Control Group  Crossover
                               (n=76)         (n=74)         (n=1)

  Mean Hemoglobin (g/dl)       13.2           13.2           14.4

  Mean hematocrit (%)          39.7           39.6           42.0

  Mean WBC (10^3^/ul)          7.8            7.4            6.9

  Mean platelet count          238.1          227.5          212.0
  (10^3^/ul)                                                 

  Mean PT (sec)                11.9           11.9           12.7

  Mean PTT (sec)               29.0           28.9           23.2

  Mean fibrinogen (mg/dl)      384.2          383.6          320.0
  ---------------------------- -------------- -------------- -------------

The preoperative laboratory values appear fairly well balanced between
the treatment groups.

At least one laboratory value was abnormal in 28% of the patients
treated with BioGlue and 17% of the patients treated without BioGlue.
Any abnormal laboratory value was commented on as to its clinical
significance and whether it contributed to an adverse event. There were
only three instances of laboratory values contributing to an adverse
event; all of these were in the group treated without BioGlue. In one
patient, a preoperative hemoglobin of 9.1 g/dl and hematocrit of 27% was
thought to have contributed to an adverse event. Two patients had low
postoperative platelet counts which were thought to have contributed to
adverse events.

### Treatment Parameters

The surgical procedures were characterized in terms of the type of
procedure, the amount of BioGlue used, and the types of materials
bonded. The following table summarizes the types of surgical procedures.
The numbers in parentheses are the percent of the total number of
procedures in the treatment group.

###### Types of Surgery

  ---------------------------- --------------- -------------- ------------
  Parameter                    BioGlue® Group  Control Group  Crossover
                               (n=76)          (n=74)         (n=1)

  Cardiac Procedures           24 (23%)        25 (26%)       0

  Aortic Procedures            57 (54%)        47 (49%)       1

  Peripheral Vascular          25 (24%)        23 (24%)       0
  Procedures                                                  

  Total                        106             95             1
  ---------------------------- --------------- -------------- ------------

The proportion of patients having each of the three general types of
procedures seems to be similar in the two treatment groups. The cardiac
procedures included 4 aortic root replacements, 1 aortoplasty, 5 aortic
valve annuloplasties, 1 aortic valve resuspension, 23 aortic valve
replacements, 2 Bentall procedures, 8 composite valve conduit
procedures, 2 mitral valve replacements, 2 Ross procedures, and 1
coronary artery bypass grafting procedure. The aortic procedures
included 21 abdominal aortic aneurism repairs, 21 ascending aortic
aneurism repairs, 9 repairs of aneurisms of the ascending and aortic
arch, 12 repairs of aneurisms of the transverse aortic arch, 1 repair of
an aneurism of the aortic arch and descending aorta, 8 repairs of
aneurisms of the descending aorta, 32 thoracoabdominal aneurism repairs
and 1 repair of a type B aortic dissection. The peripheral vascular
procedures included 5 aorto-femoral bypasses, 2 aorto-iliac bypasses, 1
aorto-innominate artery bypass, 1 carotid bypass, 19 carotid
endarterectomies, 3 femoral-distal bypasses, 2 femero-femoral bypasses,
5 femero-popliteal bypasses, 1 hepato-renal bypass, 1 popliteal-dorsalis
pedis bypass, 1 profunda femoris endarterectomy, 6 renal artery
bypasses, and 1 renal artery endarterectomy.

The sponsor reports that the average number of cartridges of BioGlue
used per surgical repair was 2.6, with a range of from 1 to 4
cartridges. The types of material connected in the anastomoses are
summarized in the following table. The numbers in parentheses are the
percent of the total number of anastomoses in the treatment group. PTFE
means polytetrafluoroethylene.

### Type of Material in Anastomoses

  ---------------------------- -------------- -------------- ------------
  Parameter                    BioGlue® Group Control Group  Crossover
                               (n=76)         (n=74)         (n=1)

  Tissue-to-polyester          147 (73%)      127 (69%)      3

  Tissue-to-tissue             19 (9%)        28 (15%)       0

  Tissue-to-PTFE               14 (7%)        15 (8%)        0

  Other                        17 (8%)        13 (7%)        0

  Data Missing                 5 (2%)         1 (1%)         0

  Total                        202            184            3
  ---------------------------- -------------- -------------- ------------

The proportion of the various types of material connected in the
anastomoses seems to be similar in the two treatment groups. In most
anastomoses, the materials being connected were tissue-to-polyester,
with some anastomoses involving tissue-to-tissue and tissue-to-PTFE. The
other materials connected included 21 anastomoses of polyester to
polyester, 3 tissue to porcine tissue, 2 cryopreserved homograft to
tissue, 2 prosthetic valve to tissue, 1 polyester to PTFE, and 1
polyester to bovine pericardium.

### Safety Results

The safety data consisted of the number of patient deaths and the other
adverse events. There were 5 patient deaths in each of the two treatment
groups. There was one crossover patient who was originally randomized to
be treated without BioGlue. Because of uncontrolled bleeding, the
patient was crossed over and successfully treated with BioGlue. This
patient is considered to be part of the BioGlue treatment group for
safety purposes. She is one of the patients who died during the study.

A brief case history is presented for each of the ten patients who died
during the study. The following table summarized the patient deaths in
this study. TAA, AAA, and TAAA mean thoracic aortic aneurism, abdominal
aortic aneurism, and thoracoabdominal aortic aneurism. CHF is congestive
heart failure.

##### Patient Deaths

  ---------------- ----- ----- -------- ------ ----------------------- -----------
  AAAT1reatment1   Age   Sex   Lesion   Days   Cause of Death          Procedure
                                                                       Related

  BioGlue®         76    M     TAA      1      CHF, unable to wean     Yes

  BioGlue®         42    F     TAA      1      CHF, cardiac arrest     Yes

  BioGlue®         79    M     AAA      1      Bradycardia, asystole   Yes

  BioGlue®         78    F     TAAA     42     Stroke                  No

  Crossover        74    F     TAA      43     Sepsis                  Yes

  Control          70    F     TAAA     15     Multiorgan failure      Yes

  Control          72    F     AVR      109    Cardiac arrest          No

  Control          79    M     TAA      85     Respiratory failure     Yes

  Control          70    F     AAA      109    Cardiac arrest          No

  Control          75    M     TAA      17     Spinal infarction       Yes
  ---------------- ----- ----- -------- ------ ----------------------- -----------

Nine of the ten patients who died had aortic aneurisms, seven of which
involved the thoracic aorta. There were a variety of causes of death,
mostly related to the patient's comorbid condition exacerbated by the
surgery. None of the patients died of complications of surgical
bleeding.

The following table summarizes the other adverse events. The numbers of
patients having the adverse events are shown in order of their frequency
of occurrence in the total of the two treatment groups. The crossover
patient is included in the BioGlue treatment group. The numbers in
parentheses are the percent of the total number of patients in the
treatment group.

##### Other Adverse Events

  ----------------------- ----------------------- -----------------------
  Parameter               BioGlue® Group (n=77)   Control Groups (n=74)

  Pleural effusionn       20 (26%)                21 (28%)

  Cardiac arrhythmia      17 (22%)                14 (19%)

  Respiratory failure     13 (17%)                12 (16%)

  Infection               13 (17%)                10 (14%)

  Renal failure           13 (17%)                9 (12%)

  Neurological deficits   5 (6%)                  16 (22%)

  Atelectasis             6 (8%)                  3 (4%)

  Hemorrhage              3 (4%)                  3 (4%)

  Incisional pain         2 (3%)                  4 (5%)

  Anemia                  4 (5%)                  2 (3%)

  Ileus                   3 (4%)                  2 (3%)

  Organ system failure    3 (4%)                  2 (3%)

  Myocardial ischemia     3 (4%)                  2 (3%)

  Myocardial infarction   3 (4%)                  1 (1%)

  Dysphagia               4 (5%)                  1 (1%)

  Pneumothorax            2 (3%)                  2 (3%)

  Altered mental status   2 (3%)                  2 (3%)

  Stroke                  1 (1%)                  3 (4%)

  Paraplegia              1 (1%)                  2 (3%)

  Depression              3 (4%)                  0
  ----------------------- ----------------------- -----------------------

Only the rate of occurrence of neurological deficits appears to be
significantly different in the two treatment groups. The reason for this
is not clear, but is probably not related to the use of the BioGlue. The
safety data do not raise any significant concerns about the safety of
the use of BioGlue within the three month follow-up period.

### Effectiveness Results 

The primary response variable was the proportion of vascular anastomoses
achieving hemostasis without the need for additional measures to control
bleeding. The following table summarizes the hemostasis data on an
intent-to-treat basis.

##### Anastomoses Achieving Hemostasis

  ---------------------------- ----------------- --------------- ---------
  Parameter                    BioGlue® Group    Control Group   p-value

  Proportion of Anastomoses    164/202 (81.2%)   105/184 (57.1%  \<0.001

  Proportion of Patients       46/76 (60.5%)     29/74 (39.2%)   0.014
  ---------------------------- ----------------- --------------- ---------

The sponsor has calculated the P-value based on Fisher's Exact Test. The
proportion of patients and the proportion of anastomoses achieving
hemostasis are both statistically significantly greater in the group of
patients treated with BioGlue.

Secondary effectiveness endpoints including the intra-operative use of
blood products, additional hemostatic measures, and re-operation because
of bleeding at the anastomotic site. The following table summarizes the
results of the secondary effectiveness endpoints. CPB means
cardiopulmonary bypass. The procedure times are measured in minutes; the
blood products are measured in units.

##### Secondary Effectiveness Endpoints

  ----------------------------- ---------------- -------------- ----------
  Parameter                     BioGlue® Group   Control Group  p-value

  Blood products, mean value                                    N.S.

  Red blood cells               2.3              1.85           N.S.

  Platelets                     5.13             5.19           N.S.

  Fresh frozen plasma           3.79             3.29           N.S.

  Cryoprecipitate               4.32             2.03           N.S.

  Donor exposures               20.6             14.2           N.S.

  Pledgets used initially       53/202 (26%)     66/184 (36%)   0.047

  Mean CPB Time                 168.1            144.2          N.S.

  Mean cross-clamp time         74.0             69.1           N.S.

  Mean total operative time     237.7            228.7          N.S.

  Mean days in ICU              3.9              4.8            N.S.

  Mean days in hospital         9.5              10.9           N.S.

  Reoperation for bleeding      0                1              N.S.
  ----------------------------- ---------------- -------------- ----------

Only the proportion of anastomoses in which pledgets were initially used
to reinforce the anastomoses was statistically significantly different
between the two treatment groups.

### Limitations of the Study

A limitation of this clinical study was the choice of a surrogate
primary effectiveness endpoint. This endpoint was the proportion of
surgical anastomoses which achieved hemostasis without additional
measures to control bleeding. Although the study showed a statistically
significant improvement in the primary effectiveness endpoint, there was
no apparent improvement in most of the clinically significant secondary
effectiveness endpoints, particularly the use of blood products. This
may have occurred because the amount of blood saved by the improved
sealing of the surgical anastomoses was small compared to the
variability of blood loss between the different surgical procedures.
This would have made it difficult to demonstrate a reduction in the use
of blood products without a much larger sample size.

Another limitation of the study was the relatively small number of
anastomoses joining tissue-to-PTFE. The sponsor has submitted additional
clinical data from the commercial use of BioGlue in the international
market. This included 35 cases of the anastomosis of tissue-to-PTFE
grafts, 17 cases of tissue-to-tissue anastomoses, and 3 cases which
involved a combination of tissue-to tissue and tissue-to-polyester
anastomoses. When the users were asked whether BioGlue was effective,
the response was affirmative in all cases where a response was recorded.
This included 31 of the 35 cases of tissue-to-PTFE anastomoses, 13 of
the 17 cases of tissue-to-tissue anastomoses, and 2 of the 3 combined
anastomoses.

Another limitation of the study was that the follow-up period of three
months did not allow assessment of long term safety data.

### Conclusion

The sponsor has developed a surgical glue intended to be used as an
adjunct to standard methods of cardiac and vascular repair such as
sutures or staples by sealing and reinforcing anastomoses. They
conducted a multicenter, randomized, blinded clinical trial of the
surgical glue in 151 patients undergoing cardiac or vascular surgical
procedures. The data show that using the surgical glue caused an
improvement in the proportion of surgical anastomoses which achieved
hemostasis without additional measures to control bleeding; however, the
study did not demonstrate significant clinical benefits such as reduced
blood use or reduced operative time. No significant safety concerns were
evident during the 3 months of follow-up.

# Immunology Review

The studies submitted were appropriate and well done. However I disagree
with the interpretation of the results. The company does not consider
the inflammatory responses and the immune response significant. I
disagree with this. The studies indicate that Bioglue is a potent
immunogen. It has the potential to sensitize patients to bovine
products. This may result in hypersensitivity responses and immune
complex disease occurring a year or more after implantation. It may also
result in making the patient allergic or sensitive to other bovine
products. This Bioglue should be used with extreme care and the patient
adequately informed of the risk of allergy to bovine products, including
milk. The most likely consequences will be anaphylactic shock, immune
complex disease with liver, lung, and joints most commonly affected.
There also may be disturbances in the lymphoid tissue of the
gastrointestinal tract with various clinical signs and symptoms. These
may occur long after the initial surgery and should not be ignored or
forgotten.

Review of the document.

Attachment 4

The standard tests for delayed hypersensitivity reactions using the
Kligman maximization test in the guinea pig and the Buehler test in the
guinea pig showed no evidence of delayed contact hypersensitivity. A
study was then undertaken in Japan to assess humoral responses in the
guinea pig.

Animals were divided into 3 groups, one receiving extracts of Bioglue,
one receiving BSA, and one receiving saline all with Freund\'s complete
antigen. The humoral immunity was assessed by challenge for acute
anaphylactic shock. They also drew blood just prior to the shock
challenge. The results of the test showed that the negative control of
saline produced no signs of anaphylactic shock and no antibody titers.
The animals immunized with BSA had high titers of antibody and severe
anaphylactic shock. The animals receiving Bioglue did not have antibody
titers as high, nor anaphylactic responses as severe as those receiving
BSA, but nevertheless, they were significantly above the negative
control.

The testing lab commented that the Bioglue seemed to sensitize but not
induce the anaphylactic shock, but expressed concern that it could
sensitize to other bovine products.

The company indicates that this test indicates the product is not
immunogenic.

I have great concerns about this product and agree with the testing
house. Patients receiving Bioglue could have hypersensitivity responses
to other bovine products, including milk, sometime later after receiving
the Bioglue.

Attachment 5 is a peer review pathology study focusing on the skin sites
and the kidney. It was very important that they did the detailed review
of the kidney sections. One of the concerns in the biological response
to immunogenic materials is the formation of immune complex disease. The
kidneys are a prime target for the effects of this. The detailed study
showed no evidence of immune complex disease. The detailed analysis of
the skin sites showed some inflammatory cells.

Attachment 3.

Describes the Buehler test and results which showed no contact
sensitivity, the total complement assay which showed no differences, and
then the 90 day subcutaneous study in the rat for implantation toxicity.
The gross findings indicate that Bioglue is an irritant. The hematologic
and clinical chemistry vary some between the groups but is not
consistent. The pathology summary indicates that there is a lot of
inflammation at the site of the Bioglue. But they dismiss it as not
significant. The tabulated animal data shows a lot of sites where no
data was entered or the tissues were unavailable for evaluation.

The pathology summary is suspect since so many data points are missing.
However, it is evident that a chronic inflammatory response is mounted
against the Bioglue.

The study in the sheep is a little confusing to sort out. It would
appear that the inflammatory response to the Bioglue occurs late, about
the time that resorption is occurring. This is expected. However, since
the resorption occurs so late, the most severe immunologic potential may
not appear until a year or more after implantation.

# Statistics Review

Question 9

The logistic regression model for binary data was used to compare two
treatment groups while adjusting for study site and treatment by site
interactions:

Log ![](media/image1.wmf)intercept + trtgrp + site + trtgrp \* site

The hypothesis testing was clearly addressed.

This is no treatment by site interaction (p=0.6804). Therefore, the data
can be pooled across sites. This reviewer is satisfied with the
response.

Question 10

A logistic regression analysis was performed to determine if any
preoperative factors, had an influence on the primary outcome
(anastomotic hemostasis). Prognostic factors are listed on page 30. The
sponsor said that site 03 was not in the model is because site 03 did
not routinely collect fibrinogen values.

This clinical decision of fibrinogen values should be decided by a
physician.

Question 11

A Table... on page 31 depicts the observed adverse events. There were no
statistically significant differences between the BioGlue group and the
Conventional group except the Neurological depicts (p=0.0091). The ratio
is 6.5% (BioGlue group) vs. 21.6% (control groups). This reviewer is
satisfied with the response.

Question 12

For the purpose of documenting complication related to the surgical
procedure, the sponsor combined the data collected at hospital
discharge, at 30 days postoperatively, and 3 months postoperatively.
This reviewer is satisfied with the response.

# Chemistry Reviews

Review of comparative chemistry for BG2000 and BG3000

1\. This file has a technical defect to convince the claims made by the
CryoLife. The evidences that the company provided have revealed the
deficiency. The claimed equivalency of the new product of BG 3000 to the
marked product BG 2000 is very questionable. The evidences are:

-   The pH test results (page 5.1 Table 1)

> \[Information redacted\]. The ∆pH is the pH change after 72 hr
> extraction compared with 24 hr extractions. The pH is a sensitive
> issue in the protein chemistry area, especially for the value of pI
> (isoelectric point). As known the pI value for serum albumin is 4.9^1^
> and 5.89 is for tryptophan. N-acetyl-L-tryptophan is a stabilizer
> added to the BSA solution. The BSA reacts with glutaraldehyde (GLU) to
> form biopolymers. As extraction time increase from 24 hr to 72 hrs,
> the pH of the extraction solution from the BioGlue implant adhesive is
> expected to change, toward more basic. The polymerized product
> consists of Schiff base. The pH of the extract should be more basic
> than BSA. In any case, the pH is expected to change, either due to the
> leaching of the monomer, or the stabilizer. As documented in Table 1,
> \[information redacted\]. The unusual phenomena may explain the
> potential damage of the BioGlue through γ-irradiation. The
> denaturation of BSA and reduction of the activity of the GLU by light
> irradiation has been cited in the textbooks. The GLU consists of two
> aldehyde groups that are very easily oxidized by either oxidants or
> light irradiation, to become a carboxylic acid. Therefore, this pH
> experiment revealed the defect for equivalency claim between BG2000
> and BG 3000 with γ-irradiation.

-   The GC Analysis:

> The GC results from Table 2 Page 6 and the GC chromatogram has
> provided strong evidence that the BG3000 is NOT equivalent to BG2000.
> The peak intensity difference between BG2000 and BG3000 (without
> γ-irradiation) is \[information redacted\] for with γ-irradiation at
> 72 hr extraction. The explanation of the difference can not based on
> the assumption it is that due to age difference, without data being
> provided. As a matter of fact, the three GC chromatograms are not
> equivalent based on the number of peaks and the peak intensity. When
> GLU been deactivated by the γ-irradiation, and forms acid, the boiling
> point will be increased, therefore, less volatile, the peak intensity
> of the GLU will be 10 to 20 times reduced (Attachment 1). There is no
> consistency to confirm the conclusion that BG3000 is equivalent to
> BG2000 from the GC data.

-   UV-VIS Analysis:

> The UV spectrum (attachment II) has shown the typical tryptophan at
> 280 nm, which is the N-acetyl-L-tryptophan from both products. That
> only confirmed the stabilizer was added to both BG2000 and 3000.
> \[Information redacted\].~.~ This information revealed that the BG2000
> has more conjugated system with GLU and Lysine unit (BSA contains 59
> units) than BG3000. The peak intensity of BG2000 is 2 times of the
> BG3000. If the company added same amount of the stabilizer into BG2000
> and 3000, then the huge difference of the peak intensity and the shift
> of the λ~max~ can only confirm that BG 3000 is not equivalent to
> BG2000.

-   The HPLC Analysis:

The HPLC analysis data was confirmed again that the BG3000 is different
from BG2000. They are not even close to an equivalency claim. I do not
think that the HPLC is a good means for this claim. One reason is the
complexity of the peaks that have not been able to identify exclusively.
Second reason is the retention time that was changing.

2\. My Suggestions:

-   NMR analysis. The GLU has typical peaks at δ 9-10 if without light
    irradiation, otherwise the peak would have a very different δ if the
    GLU has been oxidized.

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-   IR analysis. The normal GLU has IR peak at 2720 and 1725 cm^-1^, if
    after γ-irradiation, then the peak positions are changed
    drastically.

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-   CE (Capillary Electrophoresis) analysis. This is a typical tool for
    analysis of proteins. The BSA forms Schiff base with GLU would have
    different R~f~ value for the one with aldehyde group and the one
    only has carboxyl groups if an oxidation occur during the
    irradiation. If self polymerized of GLU, then the R~f~ value should
    be totally different from either of other peaks. This method is very
    sensitive and economical.

^1^ Donald Voet and Judith G. Voet **Biochemistry,** second ed. John
Wiley & Sons, Inc, page 77, 1994.

Review for the change in shelf life due to the change in polymer to
monomer absorbance ratio.

The chemical constituents of BioGlue^®^ Surgical Adhesive are purified
bovine serum albumin (BSA), a protein and glutaraldehyde (GA), a
cross-linking agent. When activated by mixing and dispensed to target
tissue sites, GA covalently cross-links the surface-proteins of the
patient's tissue to the proteins of BSA. Thus, BSA acts as a protein
matrix for the cross-linking agent, GA.

In order to thoroughly review the Shelf Life of the device, parts of
Analytical and Functional testing have been evaluated.

Conclusions are:

1.  Adhesive cure rate (or set time approximately 2 minutes) and
    adhesive shear strength \[information redacted\] meets the
    acceptance criteria.

2.  The extent of autopolymerization (the final polymer to monomer ratio
    normalized against the initial polymer to monomer ratio) of
    glutaraldehyde also meets the acceptance criteria.

3.  In response to [question \# 3 of the initial scientific
    review]{.underline} of PMA application \# P010003, the company
    submitted an amendment \# 2, a detailed (step-by-step) mathematical
    approach to show how they obtained the Arrhenius shift factor
    \[information redacted\] for autopolymerization of glutaraldehyde at
    37 C relative to 25 C, the reference temperature. Hence, a 36 month
    shelf-life under the [specific storage condition]{.underline} (25 C)
    is justifiable.

**COMMENTS OF DR. HOMBERGER, A PANEL MEMBER FROM ANOTHER PANEL,
REGARDING IMMUNOLOGY:**

The following comments are in response to your request to review
information submitted to FDA relating to toxicological testing of
CryoLife BioGlue Surgical Adhesive. In particular, you have directed me
to review studies performed by the proponent to ascertain the potential
for "immunological reactions" that may attend the use of BioGlue in
broader clinical applications including cardiac and vascular repair; and
you have requested that I address 3 questions related to the information
presented in the PMA. Regarding the likelihood that a clinical
immunologic response may occur following the use of BioGlue, I believe
the data presented are not sufficient to reach a firm conclusion. The
data do indicate that the immune systems of test animals respond to
BioGlue with formation of specific antibodies which react with
glutaraldehyde-polymerized BSA (BioGlue) and BSA (cf. Study #I-00-082
Part II). Since these antibodies are of the IgG isotype, I believe it is
justifiable to conclude that there is recruitment of antigen-specific T
lymphocytes some of which are likely to persist for extended periods of
time as "memory cells" capable of responding to subsequent or persistent
exposures to BioGlue, BSA and structurally-related antigens. This
finding does not necessarily indicate an increased risk of a clinically
significant immunologic reaction. The presence of immunologic "memory"
is a prerequisite for having an ongoing or subsequent
immunologically-mediated reaction, but further exposure to the eliciting
antigen is usually required for such a reaction to occur in vivo. The
distinction here is subtle but important. A transitory immune response
to BioGlue is not likely to be clinically significant except as it may
prime the immune system to respond in a more vigorous way to subsequent
antigen exposure. In addition, it there is sufficient antigen available
at the surgical site to continuously stimulate an already responsive
immune system, there is at least a theoretical risk of developing a
clinically significant immune reaction locally or a systemic reaction
such as immune complex mediated disease. I do not believe the data
presented are sufficient to determine the risk of such an event
occurring clinically. In view of the foregoing, it is tempting to advise
that further animal studies be performed. I hesitate to make such a
recommendation because I believe it would be very difficult to design a
study that would yield data relevant to the issue of defining the
magnitude of risk in human subjects. Nevertheless, it is prudent and
advisable to provide extensive product labeling with specific references
to the risks of using this material in patients with known sensitivity
to bovine antigens, prior histories of immune-mediated diseases
including diseases characterized by vascular inflammation, or
generalized atopy. It is also prudent to caution against the repeated
use of this material in the same individual since animal studies show
that BioGlue does elicit an immune response in animal experiments.
Regarding the advisability of performing post-market studies, I would
recommend that human patients treated with BioGlue be tested at
appropriate times following surgical use of the material for the
presence of specific antibodies to BioGlue and BSA, and that in vitro
studies of delayed hypersensitivity to BioGlue also be performed. The
latter can be performed by measuring lymphocyte blastogenesis to several
concentrations of BioGlue in serum-free medium with sensitivity defined
by calculating a stimulation index. Please contact me if you have
questions about my comments or wish to discuss the recommendations for
product labeling or post-market studies.